Editor-In-Chief's Message

Editor-in-Chief’s message

Welcome to the second issue of Research and Knowledge. As with the ﬁ rst issue, the editorial team has worked very hard to get everything ready for publication.

However, as usual the journal could not have been published without the manuscripts submitted by the authors. We had many excellent papers to choose from when selecting the work to publish this issue. The issue starts with a review of the use of ﬁ broin and sericin-derived bioactive peptides and hydrolysates and their use for health promotion; this is a comprehensive paper that will enable anyone to understand the topic. Following this there is a paper related to the challenges of using genetic improvement in livestock and aquatic animals.

The next part of this issue then has two papers related to paleontology and new ﬁ ndings from Thailand. Then there are two papers about automotive engineering, with the ﬁ rst being about the design of an intake manifold for a student formula race car and the second about the initial development of a new predictive suspension control system.

Then the fi nal two papers relate fi rst to the optimization of a power plant so that it could potentially use its natural gas more effi ciently and second to the identifi cation of the best inclination angles for the transfer of heat by a thermo-syphon. These last four engineering papers were formerly presented at the International Conference on Science, Technology and Innovation for Sustainable Well-Being (STISWB VII). The event was organized by the Faculty of Engineering and Industrial Technology, Silpakorn University together with the Graduate School, Mahasarakham University during 30th July - 2nd August 2015.

I wish to thank all the authors for submitting their work for publication as well as all the reviewers who gave their time to consider each of the submissions. The editorial team is already starting to prepare the next issue so if you have any work that is suitable for publication please submit it so we can consider it.

Professor Wichian Magtoon, PhD. Dean, Faculty of science, Mahasarkham University Editor-in-Chief, Research and Knowledge CONTENTS

Fibroin and sericin-derived bioactive peptides and hydrolysates as alternative sources of food additive for promotion of human health: A review Motoyuki Sumida and Vallaya Sutthikhum ...... 1

Challenges for genetic improvement of livestock and aquatic animals Uthairat Na-Nakorn and Skorn Koonawootrittriron ...... 18

Upper Devonian (Famennian) conodonts from radiolarian cherts, Loei Terrane, Loei Province, Northeast Thailand Clive Burrett, Hathaithip Thassanapak and Mongkol Udchachon ...... 26

New fossil woods from the Mesozoic Khorat Group of Thailand and their palaeoecological implications Marc Philippe, Pratueng Jintasakul and Nareerat Boonchai ...... 33

Development of intake manifold for Student Formula CMU F-914 using theoretical and simulated designs Niti Kammuang-lue and Worawit Borikhut ...... 39

Predictive suspension control system by inclinometer and accelerometer Jarut Kunanoppadol ...... 46

Power plant efﬁ ciency improvement by optimizing main inﬂ uential control parameters Polachate Tosomboon and Thosapon Katejanekarn ...... 52

Heat transfer characteristics of thermo-syphon: Case study of modifying inclination angle of evaporator and condenser sections Kittipon Khangam and Narong Srihajong ...... 57 Instructions for Authors

Aim & Scope Research & Knowledge is a peer-reviewed International Journal with open access that is published by the Research Center of Mahasarakham University. The aim of the journal is to provide a platform for researchers, academics, professionals, practitioners and students to publish and share knowledge in the form of high quality original research and review papers. The journal publishes in both hardcopy and online versions. It publishes papers in the ﬁ elds of Science and Technology such as Biology, Biotechnology, Botany, Chemistry, Ecology, Engineering, Environmental Science, Ichthyology, Geology, Genetics, Mathematics, Microbiology, Molecular Biology, Organic Chemistry, Paleontology, Physics, Plants and Animal Science, Statistics, Taxonomy and Zoology. All manuscripts can be submitted through online submission or email. For further information, please visit: http://www.researchandknowledge.msu.ac.th

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Journal Kofukuda, D., Isozaki, Y. and Igo, H. 2014. A remarkable sea-level drop and relevant biotic responses across the Guadalupian-Lopingian (Permian) boundary in low latitude mid-Panthalassa: Irreversible changes recorded in accreted paleo-atoll limestones in Akasaka and Ishiyama, Japan. Journal of Asian Earth Science 82, 47–65.

Conference proceedings Bunopas, S. 1983. Palaeozoic succession in Thailand. In: Nutalaya, P. (Ed.), Proceedings of the Workshop on Stratigraphic Correlation of Thailand and Malaysia: Haad Yai, Thailand, vol. 1, pp. 39–76.

Book Flügel, E. 2004. Microfacies of carbonate rocks: Analysis, interpretation and application. Springer-Verlag, Berlin, pp. 976. Aigner, T. 1982. Calcareous tempestites, storm-dominated stratiﬁ cation in Upper MuschelkalkLimestones (Middle Triassic, SW-Germany). In: Einsele, G. and Seilacher, A. (Eds.), Cyclic and Event Stratiﬁ cation. Springer-Verlag, Berlin, pp. 180–198.

PhD thesis Thambunya, S. 2005. Lithofacies and diagenesis of the KhaoKhad Formation in the vicinity of ChangwatSaraburi, Central Thailand. Ph.D. dissertation, Department of Geology, Chulalongkorn University, Thailand.

...... All correspondence should be directed to: The Editor, Research & Knowledge, Division of Research Facilitation and Dissemination (DRFD) Mahasarakham University, Kantarawichai, Mahasarakham, 44150 Thailand Phone: (66) 437-54322 ext. 1757 / 1173 or 437-54416 / 437-54247 Fax: (66) 437-54416 or 437-54247 E-mail: [email protected] Website: https://rk.msu.ac.th Vol. 1 No. 2 page 1-17 2015 DOI:10.14456/randk.2015.16 RESEARCH & KNOWLEDGE Review Article Fibroin and sericin-derived bioactive peptides and hydrolysates as alternative sources of food additive for promotion of human health: A review

Motoyuki Sumida1, * and Vallaya Sutthikhum2 1 Center of Excellence for Silk Innovation (Formerly Silk Innovation Center) Mahasarakham University, Tambon Khamriang, Khantarawichai District, Maha Sarakham 44150, Thailand 2 Department of Chemistry, Faculty of Science, Mahasarakham University, Tambon Khamriang, Khantarawichai District, Maha Sarakham 44150, Thailand

(Received 19 May 2015; accepted 13 December 2015)

Abstract - Clothing, food and housing are the fundamental three items for human beings to live humanly. Sericulture has contributed to clothing and housing for more than four thousand years via the production of clothes and goods for houses, such as curtains and bed covers, but has contributed rather little to food, except entomophagy or eating larvae or pupae inside cocoons. When we consider ingestion of silk proteins, fi broin and sericin, from cocoons in the form of bioactive peptides and hydrolysates as in bioactive peptides and hydrolysates from food proteins, such as soy, fi sh, meat, milk, egg, wheat, broccoli and rice, which are known to be benefi cial for the promotion of human health, modern sericulture should contribute to food, and therefore contribute to clothing, food and housing equally. For the preparation of bioactive peptides and hydrolysates from fi broin and sericin, enzymatic hydrolysis is a powerful tool. Based on our experience of the study of silk digestion enzyme for more than twenty years, in this review we summarize current knowledge of bioactive peptides and hydrolysates prepared from fi broin and sericin from domesticated silkworm, Bombyx mori, as well as from wild silkmoths, by proteases and their potency for the promotion of human health. Although the number of bioactive peptides and hydrolysates from fi broin and sericin is currently limited, we believe more products will be added in the future from fi broin and sericin and the contribution of modern sericulture to the promotion of human health from this aspect is likely to be assured. We encourage researchers related to silk proteins, fi broin and sericin, to perform further comprehensive studies on bioactive peptides and hydrolysates from fi broin and sericin from domesticated silkworm and wild silkmoths, both of which should provide fruitful resources for the welfare of human beings.

Keywords: Fibroin, sericin, bioactive peptides, Bombyx mori, wild silkmoths

1. Introduction Shantibala, 2010). When we consider the possibility of Clothing, food and housing are the fundamental three items ingestion of silk proteins, fi broin and sericin, from cocoons, needed for human beings to live humanly. Sericulture has as food additives, the potential contribution of modern contributed to clothing and housing for more than four sericulture to food becomes tremendous. Accordingly, thousand years (Good et al., 2009). This is because sericulture should contribute equally to clothing, food and sericulture is a systematic human activity: starting from housing. cultivating mulberry trees in the fi eld for harvesting fresh To our knowledge, the fi rst time that a method of mulberry leaves, rearing silkworm larvae on fresh ingesting the silk protein, fi broin, was devised was in 1989 mulberry leaves for harvesting cocoons, into reeling silk (Hirabayashi et al., 1989). In the 1980s the preparation of yarns from harvested cocoons and dyeing silk yarns for a food additive from fibroin that was prepared from weaving silk fabrics for making clothes and goods for extremely low cost silk, such as thin shell cocoons, which houses, such as curtains, covers of cushions and furniture, were no longer utilized for silk reeling, and discarded silk table cloths and bed covers. As for food, sericulture has fabrics, was proposed. The authors solubilized solid fi broin contributed rather little except larvae or pupae inside with a calcium chloride solution to make a fi broin solution cocoons as sources of protein nutrition (Lokeshwari and after degumming the cocoons and silk fabrics or removing

* Author for correspondence: [email protected] Former Professor of Insect Physiology, Insect Biochemistry and Sericultural Science, Center for Bioresources Field Science, Kyoto Institute of Technology, Ippon-gi Town, Saga, Kyoto City 616-8354, Japan 2 Motoyuki Sumida and Vallaya Sutthikhum R & K sericin from them, studied the gelation of the fi broin hydrolysis or by the digestion of protease, are consisted to solution to prepare fi broin powder and determined the be a mixture of amino acids and oligopeptides, and they effect of feeding fi broin powder to the assay subject. The are known to have benefi cial effects on animal subjects research group obtained benefi cial effects from the fi broin (See Table 3 in Section 8) and they could show similar powder. Fibroin powder prepared by the above method is benefi cial effects on humans. currently available as a health food or food additive, by the name of ‘Kaya silk powder’ from a company in Yosano 2. Cocoon fi lament and fi broin and sericin Town (formerly Kaya Town), Yosano County, Kyoto A single cocoon fi lament from a spinneret of a matured Prefecture, Japan. Various food products containing fi broin larva of the domesticated silkworm, B. mori, consists of powder are available on the market, e.g., Japanese noodle, two silk proteins, fi broin and sericin (Mondal et al., 2007; Japanese soba noodle, bread, cake, tofu, soft drinks and Vepari and Kaplan, 2007; Kundu et al., 2008). In a cross cooked rice. Subsequently, four forms of fi broin (fi broin section of a single cocoon fi lament, there are two columns solution, fibroin gel, bubbled fibroin gel and fibroin of fi broin fi ber with sericin around them. This is because powder) were prepared and the application of these a silkworm larva has a pair of silk glands and each of the products as food additive and bioactive peptides from silk gland connects to a single spinneret in the anterior part. fi broin on animal subjects were studied (Hirao and Igarashi, Accordingly two columns of fi broin fi ber come from a 2013), which clearly demonstrated the usefulness of fi broin spinneret. products as food additives to various types of foods and of A silk gland is a tubular organ that consists of three fibroin-derived bioactive peptides to hypertensive rat parts: from posterior to anterior, the posterior silk gland, (Igarashi et al., 2006). Fibroin solution is used to make middle silk gland and anterior silk gland (see fi gure 3 of bread containing 30% Japanese willet powder, blancmange silk gland in Mondal et al., 2007). The cells of the and yogurt jelly. Fibroin gel is used to cook sticky rice and posterior silk gland synthesize and secrete fi broin into the for making rice cakes, steamed buns, soft adzuki-bean lumen. The distal part of the posterior silk gland is made jelly, noodles, sponge cakes and sticky rice powder balls. of a blind tube, and fi broin secreted into the lumen of the Bubbled fi broin gel is used for making meringues, bubbled posterior silk gland is extruded gradually in the anterior soft adzuki-bean jelly and sponge cakes. Fibroin gel direction, i.e., to the lumen of the middle silk gland. Cells prepared by the method of Hirao and Igarashi (2013) for of the middle silk gland synthesize and secrete sericin into food is available from Matsuoka Co. Ltd., Tsuruoka City, the lumen. As a result, in a cross section of the middle silk Yamagata Prefecture, Japan. gland, a column of fi broin in the center can be seen that is Fibroin-derived bioactive peptides and hydrolysates surrounded by several layers of sericin. The anterior silk have been prepared by several methods, among which the gland is the path of the fi broin and sericin when a mature typical ones were (1) acid hydrolysis of solid fi broin using larva spins silk thread. hydrochloric acid solution or (2) solubilization of solid The protein structure of fibroin and sericin in fibroin with calcium chloride solution, followed by domesticated silkworm, B. mori, and other insect species enzymatic hydrolysis using protease or protein digestion is summarized in a review by Sehnal and Sutherland enzyme. The latter method has a link to our study and is (2008). Fibroin is secreted from the cells of the posterior the reason why we wrote this review; we have been studying silk glands into the lumen as an assembled form of a high silk digestion enzyme from silk glands of insects for more molecular mass elementary unit consisting of H-chain, than twenty years (Sumida, 2010). The silk digestion L-chain and P25 with a 6:6:1 molar ratio (Inoue et al., enzyme is synthesized by silk gland cells, secreted into the 2000). Molecular masses of H-chain, L-chain and P25 are lumen of silk glands at each molt period in the larva 350, 25 and 27 or 30 kDa due to differential glycosylation (Sutthikhum et al., 2004a; Watanabe et al., 2004) and it of P25 (Tanaka et al., 1993), respectively. Gene sequence digests the fi broin and sericin stored in the silk glands of the H-chain has been reported (Zhou et al., 2000). (Watanabe et al., 2007), and it is found in domesticated Cloning of the cDNA of the L-chain and its primary silkworm, Bombyx mori and in wild silkmoth, Samia structure has been reported (Kimura et al., 1985; cynthia ricini (Watanabe and Sumida, 2006; Watanabe Yamaguchi et al., 1989). Amino acid sequence of P25 was et al., 2006c). This is done so the larva of the silkworm or reported (Chevillard et al., 1986). Sericin mainly consists wild silkmoth ensures that the lumen contents of the silk of three proteins (A, M and P) that are synthesized and glands are vacant so that the next instar larva can synthesize secreted into the anterior, middle and posterior subparts of fi broin and sericin in the cells of the silk glands and secrete the middle silk gland, respectively (Takasu et al., 2002). them into the lumen of the silk glands. Thus, silk digestion The molecular masses of sericin A, M and P are 250, 400 enzyme is a good candidate to prepare bioactive peptides and 150 kDa, respectively, as estimated by SDS-PAGE from fi broin and sericin. In the classifi cation of proteinases, (Takasu et al., 2002). Three genes for sericin are known, the silk digestion enzyme belongs to cysteine proteinase namely Ser1 (Okamoto et al., 1982), Ser2 (Michaille et al., (Watanabe et al., 2006d). It functions optimally at an 1990) and Ser3 (Takasu et al., 2007). The Ser1 gene acidic pH of 4. The important implication of our study was encodes sericins M and P. The Ser2 gene encodes unique that it gave a rational to use cysteine proteinase for the sericin proteins with molecular masses of 230 and 120 kDa, digestion of fi broin and sericin in vitro. Hydrolysates of which are different from sericin M, P and A, and which are fibroin and sericin, which can be prepared by acid not incorporated into the cocoon silk (Kludkiewicz et al., Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 3 for promotion of human health: A review

2009). The Ser3 gene encodes sericin A (Takasu et al., of peptides have been identifi ed in bioactive peptides from 2005; 2007). Four species of small molecular mass proteins food sources (see Hong et al. (2008)). For the various kinds have been identifi ed in the silk of B. mori (Nirmala et al., of bioactive peptides identifi ed from food proteins, the 2001). They are seroin 1 and seroin 2, with molecular following physiological functions on four kinds of human masses of 9.9 and 10.3 kDa, respectively, and Kunitz-type systems are known (Hartmann and Meisel, 2007): (1) and somewhat unusual Kazal-type proteinase inhibitors cardiovascular system, hypocholesterolemic, antioxidative with molecular masses of 6 and 4.7 kDa, respectively. It is and antithrombotic functions; (2) nervous system, opioid assumed that seroins and proteinase inhibitors function in agonist and opioid antagonist functions; (3) gastrointestinal cocoon protection against predators and microbes. Interestingly, system, mineral binding, opioid agonist, opioid antagonist fi broin and sericin are different in different insect species and antimicrobial functions; and (4) immune system, in protein composition, protein structure and amino acid immunomodulatory, opioid agonist, opioid antagonist and sequence (Sehnal and Sutherland, 2008). This will open an antimicrobial functions. Erdmann et al. (2008) added opportunity to produce a wide variety of bioactive peptides antihypertensive, hypotriglyceridemic and antiobesitic and hydrolysates from fi broin and sericin from insect functions to those described above. Compared to these species distributed worldwide. functions known in bioactive peptides derived from food proteins, in bioactive peptides derived from fi broin, only 3. Bioactive peptides derived from fi broin of domesticated an antihypertensive function is known. The reason for this silkworm, B. mori may be due to the short history of research on bioactive Bioactive peptides derived from fi broin are summarized in peptides derived from fibroin, and also due to the Table 1. Three peptides are known, with the sequences of comparatively smaller number of researchers to study GY, GVGAGY and GVGY. For their functions, GY has fi broin derived bioactive peptides. Taking into consideration both ACE inhibitory activity and an antihypertensive the unique amino acid sequences in bioactive peptides from function. For GVGAGY, ACE inhibitory activity is known. fi broin, such as GY and GVGAGY with ACE inhibitory For GVGY, there is an antihypertensive function. It is activity and GY and GVGY with antihypertensive function, interesting to note that the three peptides described above further studies on bioactive peptides from fi broin will are unique peptides that are derived from fi broin with elucidate more unique bioactive peptides with various antihypertensive functions, because no similar sequences benefi cial functions on human health.

Table 1. Bioactive peptides derived from ﬁ broin and sericin of domesticated silkworm, Bombyx mori and function on assay subject.

Function Precursor Peptide Origin Reference protein sequence ACE* inhibitory Fibroin GY Cocoon Ni et al. (2001) ACE inhibitory Fibroin GVGAGY Cocoon Igarashi et al. (2006) Antihypertensive Fibroin GVGY Cocoon Igarashi et al. (2006) Antihypertensive Fibroin GY Cocoon Zhou et al. (2010) Protection from death of Sericin SGGSSTYGYS Chemical Takahashi et al. (2005) Sf 9 cells from serum synthesis** deprivation Sericin SGGSSTWGWS Chemical Takahashi et al. (2005) synthesis***

ACE*, Angiotensin I-converting enzyme **, chemically synthesized based on amino acid sequence of a peptide of 38 amino acids obtained from sericin hydrolysate which showed activity of cellular protection. ***, chemically synthesized based on SGGSSTYGYS by replacing Tyr (Y) with Trp (W).

4. Bioactive peptides derived from sericin of domesti with reference to the former peptide and by replacing Tyr cated silkworm, B. mori (Y) with Trp (W). In the case of sericin-derived bioactive Bioactive peptides derived from sericin are summarized peptides, only two peptides are known. The reason may be also in Table 1. Two peptides of biological activity that similar to the case of ﬁ broin-derived bioactive peptides, support the survival of cultured cells are known, for which i.e., due to a short history of research and smaller number the sequences are SGGSSTYGYS and SGGSSTWGWS of researchers. More bioactive peptides derived from (Takahashi et al., 2005). The former peptide was chemically sericin, which is another silk protein, will be added to the synthesized with reference to a characteristic amino acid list by further studies. sequence of sericin. The latter was chemically synthesized 4 Motoyuki Sumida and Vallaya Sutthikhum R & K

5. Enzymes used for preparation of bioactive peptides became available, and we tested if purifi ed fi broinase and hydrolysates from fi broin and sericin digested sericine (Watanabe et al., 2007), and it was found Alcalase was used in the preparation of bioactive peptides out that fi broinase digests sericin. We re-named fi broinase from fi broin (Ni et al., 2001; Igarashi et al., 2006; Zhou et as silk digestion enzyme since it digests both fi broin and al., 2010). Chymotrypsin (Chen et al., 1991), alcalase, sericin (Watanabe et al., 2006d). Accordingly, the presence trypsin and pepsin (Park et al., 2002) were used for the of fi broinase alone in the lumen contents of silk glands is preparation of fi broin hydrolysate. Protease N “amino”, suffi cient for the digestion of both the fi broin and sericin protease P “amino” 6, alcalase 2.4L, neutrase 1.5MG and stored in the silk glands at each molt period in the larva 1.398 neutral protease were used for preparation of sericin and early pupa in B. mori. We demonstrated that fi broinase hydrolysate (Wu et al., 2008). Since the preparation of food in the silk glands of the eri silkworm, S. cynthia ricini, additives from fi broin and sericin is the objective, enzymes shows extremely high activity at the end of the spinning that have a long history of use for preparation of bioactive period, some 38.3-fold higher activity per individual insect peptides from foods and enzymes that are guaranteed for than the maximum activity of the silkworm, B. mori, at the their safety to humans should be chosen, as in the production fourth molt period in the fourth instar larva or day one pupa of milk-derived antioxidative peptides in which trypsin, (Watanabe et al., 2006c), both of which show similar high chymotrypsin, validase, pepsin, bacterial and plant food activity, and the enzymatic entity of the eri silkworm is grade enzymes, alcalase, protamex, neutrase, thermolysin highly likely to be slightly different from that of B. mori, and corolase PP were used (Power et al., 2013). such as a slightly different N-terminal amino acid sequence of the enzyme (Watanabe and Sumida, 2006). This suggests 6. Silk digestion enzyme that variation in the silk digestion enzyme among different Silk digestion enzyme or fi broinase, the name given in our insect species exists in the developmental expression initial study, was, at fi rst, detected by its ability to digest profi le as well as in the quantities produced during development liquid fi broin in vitro, visualized by SDS-PAGE, and it was and probably in the molecular structure of the enzyme found from degenerating silk glands from day one pupa, itself. The high activity of the silk digestion enzyme in the of the silkworm, B. mori (Sumida et al., 1993a). The question, silk glands of S. cynthia ricini at the end of spinning in the ‘Do silk glands produce silk digestion enzyme or fi broinase?’ fi fth instar larva opens an application for this enzyme as a based on the experience from the enzyme study in B. mori, source of a degumming agent as well as for an enzyme to and it was tested using, at fi rst, a homogenate of silk glands prepare fi broin and sericin-derived bioactive peptides and remaining in a day one pupa of B. mori as the enzyme hydrolysates. On the other hand, from an academic point source, since the silk glands remaining in the pupa become of view, we believe that if the molecular features of the silk slender from day zero to day one pupa, which suggested digestion enzyme in each insect species would be revealed, that fi broin and sericin remaining within the pupal silk the mechanism of co-evolution of the substrate and glands must be digested by some protease within the silk enzyme, or between fi broin or sericin and silk digestion glands from day zero to day one pupa. The history of the enzyme, will be elucidated in future studies. Anyhow, silk study of silk digestion enzyme will be reported elsewhere digestion enzyme digests both fi broin and sericin; and as a case study (Sutthikhum and Sumida, in preparation). accordingly, silk digestion enzyme is a candidate enzyme The literature survey, then, showed that Akai (1965) to prepare bioactive peptides and hydorlysates from fi broin described observations by light microscopy in which the and sericine. Incidentally, another physiological function complete digestion of fi broin and sericin occurs in the of silk digestion enzyme was found in the feeding period lumen contents of the silk glands of B. mori at the fourth at each larval stage and at the spinning period in the last molt period in the fourth instar larva. This suggested the fi fth instar larva in B. mori (Sutthikhum et al., 2004a, b). presence of silk digestion enzyme or fi broinase in the silk The enzyme functions as a lysosomal enzyme in these glands at the fourth molt period in the fourth instar larva. developmental periods, and functions within lysosomes in We assayed fi broinase activity using silk glands from the the silk gland cells to digest obsolete proteins and organelles fourth molt period of the fourth instar larva as the enzyme transported into the lysosomes, such as endoplasmic source (Sumida et al., 1993b). Strong activity was reticulum and mitochondria that are no longer function to detected, which is comparable to the high activity in day regenerate highly functional protein synthesizing machinery one pupa. The result suggested that fi broinase is involved of the silk gland cells. in the physiological function in silk glands to prepare There is no guarantee that we can obtain bioactive vacant lumen contents at each molt period in the larva for peptides from fi broin and sericin once we use silk digestion the next instar larva to synthesize fi broin and sericin in the enzyme as a hydrolysis agent. There is only the fact that cells of the silk glands and to enable the next instar larva silk producing insects synthesize silk digestion enzyme in to secrete them into the lumen of the silk glands. Subsequently, the cells of the silk glands themselves and utilize it for the we purifi ed fi broinase from the silk glands of B. mori from digestion of fi broin and sericin stored in the lumen of silk the fourth molt period in the fourth instar larva (Watanabe glands at each molt period in the larva and early pupa as et al., 2004) and from day one pupa (Watanabe et al., well as in the feeding period and spinning period within 2006a) and characterized the enzymatic properties. Fibroinase the lysosomes as a lysosomal enzyme. Various kinds of was found to be a cysteine proteinase and the properties of proteases have been utilized for production of bioactive the enzyme were similar to those of cathepsin L. When peptides from food proteins (Power et al., 2013), and they purifi ed by the method of Takasu et al. (2002), sericine should be tried and used in addition to silk digestion Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 5 for promotion of human health: A review

enzyme. We experienced high specifi city of protease in the and sericin with ACE inhibitory, antioxidant, antithrombotic, hydrolysis of the silk protein sericin. Hydrolysis of the hypocholesterolemic and antiobesity activities. In their sericin A fraction by the silk digestion enzyme produced a comments on ACE inhibitory activities, they describe that sericin peptide, s-A12, of which the sequence was its structural element is a Tyr or Phe as the C-terminus. XPFPKASSXF… (Watanabe et al., 2007). On the other This corresponds to the case of GY from fi broin (Ni et al., hand, Takasu et al. (2005) obtained a peptide, 2-1, 2001) and GVGAGY also from fi broin (Igarashi et al., ASSSFDASSA…, as a sericin product that was hydrolyzed 2006) (See Table 1 in Section 3). Their remarks state that by lysyl endopeptidase. We speculated that XPFPKASSXF… dipeptides with a C-terminal Tyr produces a higher overlapped with ASSSFDASSA… in the region of antihypertensive effect compared to dipeptides with ASSXF… in our product and ASSSF in a product by C-terminal Phe. It is interesting to note that GY and Takasu et al. (2005), and lysyl endopeptidase cleaved the GVGAGY with antihypertensive effects have never been fi fth amino acid, lysine, of XPFPKASSXF…. This speculation found from peptides derived from food proteins, and yet may be true. We felt that enzymatic hydrolysis of sericin their remarks are valid for bioactive peptides from fi broin. was very precise. The same must be true of the hydrolysis Hong et al. (2008) provide a comprehensive summary of of fi broin by proteinase. In the case of silk digestion the antihypertensive effect of peptides with numerous enzyme from silk glands, the specifi city is fairly high, and sequences obtained from foods, such as milk, egg proteins, it cleaves preferentially the peptide bond between the Gly fi sh, globin hydrolysate and plants, which are a collection and Ala of the fi broin molecule (Watanabe et al., 2004) to of data handy to fi nd bioactive peptides from fi broin and produce two peptides fi nally from fi broin by prolonged sericin. Power et al. (2013) provided a comprehensive enzymatic hydrolysis, such as AGYG and AGAGAGYG summary of the enzymatic production of bioactive peptides (Watanabe et al., 2006b). that specifi cally had antioxidative functions. They also provided a summary of the in vitro assay system of 7. Food derived bioactive peptides as references for antioxidant properties. Raikos and Dassios (2014) preparation of bioactive peptides from fi broin and provided a summary of biofunctional peptides derived from sericin human and bovine milk proteins with effects on human There are good references related to food derived bioactive health with the sequences of the peptides. de Castro and peptides (Table 2). They are useful when we prepare Sato (2015) provided a summary of the production of bioactive peptides from fi broin and sericin. The food bioactive peptides with fermentation and proteases from sources are diverse: they are sardine muscle, sake lees, food proteins, such as soy, rice, casein, okara, wheat, cow wakame, chicken muscle, wheat germ, α-zein and α- milk, bean by fermentation, soy, purifi ed soy protein, lactalbumin, caseins, milk proteins, soy, fi sh, meat, eggs, bovine hemoglobin, bean, goby muscle, salmon and broccoli, rice, β-lactoglobulin, globin, marine animals and cuttlefi sh (Sepia offi cinalis) muscle by proteases. They also plants, whey proteins, American lobster, mushroom, provided a comprehensive summary of the analytical chlorella vulgarian 87/1, human milk, okara and bean. The methods for the purifi cation and identifi cation of bioactive diversity of foods indicates that a large variety in the peptides and major methods for measuring antioxidant amino acid sequence of food proteins provides a large activities of peptides in vitro and in vivo, and their respective number of bioactive peptides that are unique in biological mechanisms. functions. Related to this, we will propose the use of fi broin and sericin from a wide variety of species of wild silkmoths 8. Hydrolysate derived from fi broin of domesticated in addition to fibroin and sericin from domesticated silkworm, B. mori, and function on assay subject silkworm, B. mori, in the latter section of this review. The Fibroin hydrolysate from B. mori and its function on the preparation of bioactive peptides from foods is generally assay subject are shown in Table 3 in chronological order. carried out by two methods: one is enzymatic hydrolysis Fibroin hydrolysates were prepared by acid hydrolysis by and the other is fermentation. The functions of bioactive HCl solution except the one listed fi rst, in which fi broin peptides from foods are diverse. They are generally powder was directly tested (Hirabayashi et al., 1989) and classifi ed according to the functions on each of the human the one listed fourth, in which preparation of undegraded systems, such as (1) cardiovascular system, (2) nervous native molecular fi broin solution was described for use as system, (3) nutrition system or gastrointestinal system and a standard reference of fi broin molecule (Yamada et al., (4) immune system. Each reference in Table 2 has special 2001) in the study of fi broin hydrolysis. Various effects features. For example, Hartmann and Meisel (2007) were observed, which included lowering blood cholesterol provide a concise, comprehensive overview of the subject level, antigenotoxicity, enhancement of insulin sensitivity and it is especially handy to consider the functions of and glucose metabolism, inhibition of adipocyte differentiation peptides on human health. They also provide information and stimulation of osteoblastic differentiation. It is likely about commercially available functional foods or food that hydrolysate derived from fi broin contains putative ingredients carrying bioactive peptides. Erdmann et al. bioactive peptide/peptides or it is likely that amino acids (2008) provided many examples of ACE inhibitory in the hydrolysate showed a synergistic function. If the peptides with in vitro antihypertensive effects and the former is the case, efforts towards the purifi cation and sequences of the peptides. They describe the structural identifi cation of the putative peptide/peptides from fi broin properties of selected biofunctional peptides, which give hydrolysate are worthwhile. good hints to determine the bioactive peptides from fi broin 6 Motoyuki Sumida and Vallaya Sutthikhum R & K

Table 2. Bioactive peptides derived from foods.

Source Preparation Function Remarks Reference Sardine Enzymatic Angiotensin I- converting Development of a novel Li et al. (2004) muscle, hydrolysis enzyme (ACE) inhibitory functional food for Sake lees prevention of Wakame hypertension as well as Chicken for therapeutic purposes muscle, Wheat germ, α-zein α-lactalbumin Caseins Selective Cardiovascular system Biological signiﬁ cance, Silva and Malcata enzymatic Antithrombotic impact on human health (2005) hydrolysis Antihypertensive and manufacture of Nervous system novel functional food Opioid Agonistic ingredients Antagonistic Nutrition system Caseinophophopeptide Glycomacropeptide Immune system Immunomodulatory Antimicrobial Milk Fermentation Hypotensive Peptides resistant to FitzGerald and proteins ACE inhibitory further degradation by Murray (2006) gastro-intestinal and serum proteinase/ peptidase following oral ingestion Milk Enzymatic ACE inhibitory Preparation by (a) Korhonen and proteins hydrolysis, Antihypertensive enzymatic hydrolysis, (b) Pihlanto (2006) fermentation Opiod fermentation, Immunostimulatory (c) proteolysis by Antioxidative microbial proteases or combination of (a) and (b) or (a) and (c) Soy Enzymatic Antioxidant Prevention of age-related Wang and de Mejia hydrolysis, Hypotensive chronic diseases (2006) fermentation Antihypertensive Hypocholesterolemic Immunostimulating Antioxidative ACE inhibitory Soy * Cardiovascular system Comprehensive overview, Hartmann and Meisel Fish Hypocholesterolemic table of commercially (2007) Meat Antioxidative available functional foods Milk Antithrombotic or food ingredients Egg Nervous system carrying bioactive Wheat Opioid agonist peptides Broccoli Opioid antagonist Rice Gastrointestinal system Mineral binding Opioid agonist Opioid antagonist Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 7 for promotion of human health: A review

Table 2. Bioactive peptides derived from foods. (Cont.)

Source Preparation Function Remarks Reference Antimicrobial Immune system Immunomodulatory Opioid agonist Opioid antagonistAntimicrobial Milk * Reduction of cardio- ACE inhibitory peptides Erdmann et al. Fish vascular disease with in vitro (2008) Meat Antihypertensive antihypertensive effects Egg Antioxidative with sequences of Soy Antithrombotic peptides, table of Wheat Hypocholesterolemic commercially available Hypotriglyceridemic functional foods carrying Antiobesitic bioactive peptides, table of structural properties of selected biofunctional peptides, highly useful to fi nd out bio-active peptides with ACE inhibitory, antioxidant, anti-thrombotic, hypocholesterolemic and antiobesitic activities β-lacto- Enzymatic ACE inhibitory Structure, biological Hernandez-Ledesma globulin hydrolysis Antihypertensive signifi -cance and et al. (2008) Antioxidant mechanism of action of Antimicrobial bioactive peptides derived Immunomodulatory from b-lactoglobulin Opioid Hypocholesterolaemic Milk * ACE inhibitory Comprehensive summary Hong et al. (2008) Fish Antihypertensive of antihypertensive effect Egg of peptides with Globin numerous sequences from Plants foods. Milk Enzymatic ACE inhibitory Reduction of mild Korhonen (2009) hydrolysis, Immunostimulatory hypertension microbial Antimicrobial fermentation Antihypertensive Casein Enzymatic Antioxidant Selected biological Phelan et al. (2009) hydrolysis, Cytomodulatory effects, application in fermentation, Immunomodulatory industry and safety proteolysis by Antimicrobial aspects and regulations enzymes from relating to the use of micro-organisms or bioactive peptides plants Marine Enzymatic Antioxidant Bioactive peptides Kim and Wijesekara animals hydrolysis Anticoagulant in functional foods or (2010) and Antimicrobial nutraceuticals and plants pharmaceuticals Whey Enzymatic Immune system Specifi c physiological Mandureira et al. proteins hydrolysis, Antimicrobial effects, achieved (2010) microbial Immunomodulatory mechanisms, stabilities of fermentation Cytomodulatory peptides in Cardiovascular system gastrointestinal route 8 Motoyuki Sumida and Vallaya Sutthikhum R & K

Table 2. Bioactive peptides derived from foods. (Cont.)

Source Preparation Function Remarks Reference Antihypertensive Antioxidative Antithrombotic Hypocholesterolemic Nervous system Opioid agonist Opioid antagonist Gastrointestinal system Mineral-binding Anti-appetizing Chicken Enzymatic ACE inhibitory Industrial-scale manufac- Agyei and Danquah muscle, hydrolysis Antimicrobial turing of pharmaceutical (2011) a-zein Antiviral grade peptides American Immunomodulatory lobster, Opioid antagonist Soy hydro- Antithrombotic lysates, Hypocholesterolemic Bovine milk Mushroom Egg Rice Chlorella vulgarian 87/1, Human milk Muscle Enzymatic Antihypertensive Incorporation of bioactive Ryan et al. (2011) proteins hydrolysis Antioxidant peptides into functional from Antimicrobial foods and neutraceuticals meat Antiproleferative and fi sh Milk Enzymatic Antimicrobial Further insightful Agyei and Danquah Mush- hydrolysis Immunomodulatory research needed on the (2012) room, pharmaco-kinetics of Soy beans immunomodulatory Egg peptides in vivo, clinical Wheat studies needed Chlorella vulgarian 87/1 Marine Protease Antihypertensive Strategic production and Udenigwe and Aluko proteins hydrolysis Antioxidant processing methods, (2012) Hypolipidemic elucidation of in vivo Hypocholesterolemic molecular mechanisms of Anticancer action, safety at various Immunomodulatory doses and pharmacologi- Anti-infl ammatory cal activity in maintaining Multifunctional homeostasis during aberrant health conditions in human subjects Milk Enzymatic Antioxidative Comprehensive Power et al. (2013) hydrolysis enzymatic production of bioactive peptides specifi cally antioxidative Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 9 for promotion of human health: A review

Table 2. Bioactive peptides derived from foods. (Cont.)

Source Preparation Function Remarks Reference function, table of in vitro assay to evaluate food protein/peptide with antioxidant properties Human milk * Antihypertensive Peptides with effects Raikos and Dassios Bovine milk Antioxidant on human health with (2014) Antithrombotic Opiod sequences, table of Immunomodulatory in vitro assay system of Mineral binding antioxidant properties Antimicrobial Soy Fermentation, Antihypertensive Production of de Castro and Sato Rice enzymatic hydrolysis Antioxidant bioactivepeptides with (2015) Casein Immune-regulatory fermentation and protease Okara Immunomodulatory from food proteins, table Wheat Antiadipogenesis of analytical methods of Cow’s milk Antimicrobial purifi cation and Bean Anti-infl ammatory identifi cation of Salmon Anticoagulant bio-active peptides, and methods for measuring various activities

*, not speciﬁ cally mentioned.

Table 3. Fibroin and sericin hydrolysates from cocoons of domesticated silkworm, Bombyx mori and function on assay subject.

Function Assay subject Preparation Results or potencies Reference Fibroin hydrolysates Enhanced Rat 0.5 g fi broin powder in 2 Light coma but full Hirabayashi et al. alcohol ml PBS + 3 ml 50% recovery, no dead rats (1989) metabolism ethanol in PBS Digestibility Rat Acid (2N HCl for 48 h), 90% digestibility Chen et al. (1991) of fi broin Chymotrypsin 70% digestibility, higher digestibility in acid hydrolysed fi broin

Lowering Rat Acid (3N HCl for 48 h) Prevention of Chen et al. (1993) blood cardiovascular disease cholesterol and brain blood vessels level disease Undegraded Ajisawa’s Standard methods to Yamada et al. (2001) native method, prepare native ﬁ broin molecular LiSCN solution from silkworm ﬁ broin method cocoons solution Antigenoto- Mouse Acid (2NHCl for 4 h) Chemopreventive Park et al. (2002) xicity Embryo Enzymatic hydrolysis functional peptides 3T3 (Alcalase, trypsin, cells pepsin) Enhancement Adipocyte Acid (2M HCl for 4 h) Improvement of diabetic Hyun et al. (2004) of insulin 3T3-L1 hyperglycemia sensitivity cells and glucose metabolism 10 Motoyuki Sumida and Vallaya Sutthikhum R & K

Table 3. Fibroin and sericin hydrolysates from cocoons of domesticated silkworm, Bombyx mori and function on assay subjoet. (Cont.)

Function Assay subject Preparation Results or potencies Reference Inhibition of Adipocyte Acid (6M HCl for 5 h) Inhibition of Notch Jung et al. (2011) adipocyte 3T3-L1 cells signaling pathway, differentiation treatment of obesity and related metabolic diseases Stimulation Osteoblast cell Acid (6M HCl for 5 h) Suppression of Notch Jung et al. (2013) of osteoblastic C3H10T1/2 signaling pathway, differentiation M2-1084 promotion of bone healing, therapeutic intervention for bone fractures and osteoporosity Sericin hydrolysate Antioxidant, Rat Enzymatic hydrolysis Valuable ingredients Wu et al. (2008) Tyrosinase- (Protease N “amino”, proin the food, cosmetic inhibitory tease P “amino” 6, alcalase and medicine 2.4 L, neutrase 1.5 MG, industries 1.398 neutral protease)

9. Hydrolysate derived from sericin of domesticated and Mizuno, 2001). Also in Caddisfl y listed 12th from the silkworm, B. mori, and function on assay subject top (Yonemura et al., 2006) fi broin is devoid of P25. In S. Sericin hydrolysate from B. mori and its function on the cynthia ricini listed at the bottom of Table 4 (Sezutsu et assay subject are shown also in Table 3. Sericin hydrolysate al., 2014), fi broin gene contains repetitive polyalanine was prepared by enzymatic hydrolysis using protease N block, which is devoid in fi broin in B. mori. (See also Craig “amino”, protease P “amino” 6, alcalase 2.4L, neutrase and Riekel (2002) listed sixth from the top of Table 4 and 1.5MG and 1.398 neutral protease. Antioxidant activity Fedic et al., (2002) listed seventh from the top of Table 4.) and tyrosinase-inhibitory activity were observed. Similar Therefore the fi broin proteins seemed to be vastly diverged mechanism of action on the assay subject as with fi broin among wild silkmoths. These facts show that bioactive hydrolysate are likely in the case of sericin hydrolysate, peptides and hydrolysates produced from fi broin, sericin i.e., (1) presence of bioactive peptide/peptides in sericin and other silk proteins of wild silkmoths are highly likely hydrolysate or (2) synergistic function of amino acids in to be unique. To our knowledge, no information is cur- sericin hydrolysate. rently available on the sericin gene from wild silkmoths. Fibroin isolated from wild silkmoths is shown in 10. Fibroin, sericin and other silk proteins from wild Table 5. Considerable differences among the fi broins from silkmoths as another source of wide variety of wild silkmoths are now known in the structural assembly bioactive peptides and hydrolysates of the fi broin subunits, amino acid sequence and molecular Table 4 shows references for the genes of fi broin and masses. For example, in P. ricini listed fourth from the top other silk proteins from a wide variety of species of wild of Table 5 (Ahmad et al., 2004) and in A. assama listed silkmoths in chronological order, except Gonometa fi fth from the top of Table 5 (Ahmad et al., 2004) fi broin postica (Mhuka et al., 2013), which is from a paper on is consisted of H-chain and L-chain but the molecular mass fi broin protein. It is included here for convenience to show of each subunit was different, H-chain in P. ricini, 97-kDa, a list of the variety of species of wild silkmoths that produce H-chain in A. assama, 220-kDa, L-chain in P. ricini, silk proteins. The table shows that genes for fi broin and 45-kDa, L-chain in A. assama, 20-kDa. In A. mylitta listed other silk proteins from wild silkmoths are vastly diverged. second from the top of Table 5 (Datta et al., 2001) and For example, the paper listed fourth from the top (Tanaka listed second from the bottom of Table 5 (Mandal and and Mizuno, 2001) showed that in D. spectabilis and P. Kundu, 2008), although the molecular mass is not fi xed xuthus, homologues of L-fi broin and P25 of B. mori were yet, fi broin seemed to be consisted of dimeric fi broin clearly identifi ed as in G. mellonella in the papers listed molecules, which is different from the composition of fi rst and second in Table 4, but in A. yamamai, homologues fi broin of B. mori, which is consisted of H-chain , L-chain of L-fi broin and P25 of B. mori were not identifi ed. In A. and P25, with a 6:6:1 molar ratio (Inoue et al., 2000). yamamai, fi broin was consisted of H-fi broin alone (Tanaka Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 11 for promotion of human health: A review

Table 4. Genes for ﬁ broin and other silk proteins from wild silkmoths.

Species Fibroin Characteristics Reference Galleria mellonella Light chain fi broin L-fi broin (25 kDa) Zurovec et al. (1995) occurs in two isoforms, shorter one lacking the Ala-Pro dipeptide residue at its N-terminus Galleria mellonella P25 gene 29-and 30-kDa proteins Zurovec et al. (1998) from a single gene of P25 with post-trans-lational modifi -cation Antheraea pernyi Fibroin gene characterization of full Sezutsu and Yukuhiro length gene of fi broin, 80 (2000) tandemly arranged polyalanine-units Dendrolimus spectabilis L-fi broin P25 homologues of L-fi broin Tanaka and Mizuno (2001) Papilio xuthus and P25 of B. mori in Antheraea yamamai D.spectabilis, P. xuthus but not in A. yamamai Galleria mellonella H-fi broin 500 kDa H-fi broin, over Zurovec and Sehnal (2002) 95% of the protein consists of highly ordered repetitive structures, unmatched in other species Spiders MA and Flag silk proteins, comparative architecture of Craig and Riekel (2002) Bombyx mori H-fi broin, sericin protein, silk, fi brous proteins and Antheraea pernyi Balbiani Ring gene their encoding genes Chronomus tentans proteins Lepidopteran insects H-fi broin, L-fi broin, P25, overview of the silk of Fedic et al. (2002) sericin, seroin, protease Lepidoptera inhibitor Galleria mellonella H-fi broin gene correlation between fi broin Fedic et al. (2003) Ephestia kuehniella amino acid sequence and Plodia interpunctella physical silk properties Bombyx mori H-fi broin intron gene functional signifi cance Martinez et al. (2004) Bombyx mandarina on the regulation of transcription Gelleria melonella H-fi broin, L-fi broin, P25 construction of silk fi ber Sehnal and Zurovec (2004) core Yponomenta evonymella H-fi broin design of silk fi ber Yonemura and Sehnal composition conserved for (2006) more than 150 million years Caddisfl yH-fi broin, L-fi broin H-fi broin (>500 kDa), Yonemura et al. (2006) L-fi broin (25 kDa), no P25 Vespa simillima Four major hornet complex of alanine-rich Sezutsu et al. (2007) xanthoptera Cameron silk genes and serine-rich sequences Various insect species Fibroin and sericin genes, silks produced by insect Sehnal and Sutherland proteins labial glands (2008) Rhodinia fugax Fibroin gene Leucine-rich fi broin Sezutsu et al. (2008) Bombyx mori Antheraea H-fi broin gene pro-moter fi broin gene promoter Sezutsu et al. (2009) yamamai conserved 12 Motoyuki Sumida and Vallaya Sutthikhum R & K

Table 4. Genes for ﬁ broin and other silk proteins from wild silkmoths (Cont.).

Species Fibroin Characteristics Reference Corcyra cephalonica L-fi broin gene, P25 gene 20E regulates expression of Chaitanya and Dutta-Gupta L-fi broin gene and P25gene (2010) Hepialus californicus L-fi broin, H-fi broin L-fi broin, a pro-missing Collin et al. (2010) molecular marker for the study of evolutionary processes Antheraea mylitta Seroin 1 gene transcript analysis of whole Maity et al. (2010) Seroin 2 gene bodies and silk glands Caddisfl yH-fi broin H-fi broin serines Stewart and Wang (2010) phos-phorylated Stenopsyche marmorata H-fi broin characterization of unique Wang et al. (2010) H-fi broin Cricula trifenestrata Fibroin gene characterization of partial Suriana et al. (2011) coding region Gonometa postica Fibroin protein chemical, structural and Mhuka et al. (2013) Gonometa rufobrunnea thermal proper-ties of fi broin Samia cynthia ricini Fibroin gene complete nucleotide Sezutsu et al., (2014) sequence, a large region of repetitive arrays, common poly-alanine block and variable nonpoly-alanine block, fi broin dimers

Table 5. Fibroin from wild silkmoths.

Species Fibroin Preparation Potencies Reference Antheraea pernyi Fibroin 4 N HCl for 48 h Preparation of fi broin Lu et al. (1996) from cocoons powder, soluble in water, with characteristic amino acid composition Antheraea mylitta Fibroin from larval 1% lithium LDS Homodimeric protein Datta et al. (2001) silk glands of two similar sized polypeptides of 197 kDa Cricula trifene-strata Yellow pigment is Yamada et al. (2001) mainly present in fi broin core of cocoon thread Philosamia ricini 60% LiSCN 97-kDa (H-chain) Ahmad et al. (2004) in water for 2 h and 45-kDa (L-chain) H-fi broin constituted fribroin protein Antheraea assama H-fi broin 60% LiSCN 220-kDa (H-chain) Ahmad et al. (2004) L-fi broin in water for 2 hr and 20-kDa (L-chain) Antheraea mylitta fi broin 1% SDS 395 kDa and 197 kDa Mandal and Kundu (2008) Antheraea assama fi broin * Silk-based nonwoven Kasoju et al. (2009) fi broin scaffl old for application in tissue engineering and regenerative medicine *, Data are not available. Fibroin and sericin-derived bioactive peptides Vol 1. No 2, July-December 2015 and hydrolysates as alternative sources of food additive 13 for promotion of human health: A review

Sericin isolated from wild silkmoths is shown in silkmoths, and the thermostability of protease is actually Table 6. Differences among the sericins from wild important in industrial processing of fi broin and sericin. silkmoths are now known in the amino acid sequence and Prasad et al. (2012) studied Antheraea mylitta cocoonase molecular masses. Information about fi broin, sericin and for its use in cocoonase cooking. Their idea was to utilize other silk proteins from wild silkmoths indicates that the natural protease cocoonase for degumming of silk, and we vast diversity in fi broin, sericin and other silk proteins from appreciate it as a good proposal. Cocoonase is synthesized wild silkmoths should provide an indispensable resource in the galea of the pupa and extruded onto the surface of for the preparation of bioactive peptides and hydrolysates the galea to form protein crystals of pure cocoonase. When with unique biological functions. a wild silkmoth adult ecdysed from the pupa in the cocoon, Useful information that is available for the preparation it secretes saliva from the mouth and dissolves the cocoonase of bioactive peptides and hydrolysates from fi broin and crystal to make a cocoonase solution. The adult moth puts sericin of wild silkmoths is as follows. Gheysens et al. the solution onto the inner surface of the cocoon, and the (2011) demonstrated that demineralization enables the sericin is hydrolyzed but the thin fi lament of fi broin of the reeling of the cocoons of wild silkmoth, Gonometa postica, cocoon thread remains. The adult moth pushes its head to which is very useful and signifi cant for obtaining intact the fi broin fi laments to make a hole, and the adult moth yarns from this species as well as for obtaining undamaged emerges from this hole. Cocoonase belongs to the serine biopeptides and hydrolysates after obtaining intact yarns. proteinases and functions at a weak alkaline pH and Rajasekhar et al. (2011) discussed a thermostable bacterial hydrolyzes sericin only. In this sense, cocoonase is a good protease as a new way for obtaining quality silk production. degumming enzyme (see cocoonase section in Law This is useful for enzymatic production of bioactive (2015)). There is no danger of cocoonase hydrolyzing the peptides and hydrolysates from B. mori and from wild fi broin of the silk thread.

Table 6. Sericin from wild silkmoths.

Species Sericin Preparation Results or potencies Reference Antheraea assama from cocoon * 66 kDa Ahmad et al. (2004) Philosamia ricini from cocoon * 66 kDa Ahmad et al. (2004) Antheraea mylitta from cocoon 8 M urea, 2% 200 kDa sericin in Dash et al. (2006) peduncles β-mercapto-ethanol, the peduncle, 2% SDS, 80ºC for 5 different from sericin min of B. mori in amino acid composition Antheraea mylitta 70 kDa sericin 8 M urea,2% Improved Dash et al. (2007) β-mercapto-ethanol, understanding of role 2% SDS, 80ºC for 5 of sericins in forming min or 1% NaOH stable ﬁ broin overnight at room ﬁ ber-sericin temperature followed composite by anion exchange chromatography Antheraea mylitta from peduncle * 200 kDa Kundu et al. (2008) Antheraea assama from cocoon * 66 kDa Kundu et al. (2011) Philosamia ricini from cocoon * 66 kDa Kundu et al. (2011)

*, Data are not available. 11. Conclusion foods from silk proteins have been neglected in sericulture Modern sericulture deals with products from domesticated for a very long time. silkworm, B. mori, as well as from a wide variety of species of wild silkmoths that include typically eri silkworm, S. Acknowledgements cynthia ricini, for the preparation of a wide variety of We thank Prof. Dr. Hiromu Akai, Tokyo University of ﬁ broin and sericin products that include bioactive peptides Agriculture; Prof. Okitsugu Yamashita, Nagoya University; and hydrolysates for the promotion of human health. This Prof. John H. Law and the late Prof. Michael A. Wells, is undoubtedly one of the most important objectives to be Insect Science Center, The University of Arizona; and Prof. achieved in sericulture. In this sense, modern sericulture Dick J. Van der Horst, Utrecht University for encouragement should contribute to clothing, food and housing: all the and discussion on the study of silk digestion enzyme. We more so because aspects of the preparation of functional thank Mahasarakham University for support for this study. 14 Motoyuki Sumida and Vallaya Sutthikhum R & K

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Uthairat Na-Nakorn1,* and Skorn Koonawootrittriron 2 1Department of Aquaculture, Faculty of Fisheries, Kasetsart University, Chatuchak, Bangkok, Thailand 2Department of Animal Husbandry, Faculty of Agriculture, Kasetsart University, Chatuchak, Bangkok, Thailand

(Received 7 September 2015; accepted 11 December 2015)

Abstract-This article reviews the challenges for genetic improvement of livestock and aquatic animals to produce sufﬁ cient qualities and quantities of food supplies for the increasing world population. Both sectors are threatened by various factors, and genetic improvement is a promising way to achieve the goals. For livestock genetic improvement, it is crucial to enhance contributions to small scale farmers, probably through on-farm genetic improvement programs, while minimizing the impact on extinction of local breeds. Genetic improvement can mitigate climate change by improving traits that indirectly result in less emissions of greenhouse gases (GHG) throughout the production period. It also provides adaptation measures by improving animals with adaptability (e.g., heat tolerance, disease resistance, etc.) traits. Genetic improvement for stress tolerance and other related traits would cope with issues on animal welfare while disease resistant traits would partially solve problems enhanced by globalization (e.g., trans-boundary animal translocation). While in the aquaculture sector, genetic improvement also has high potential to overcome the upcoming limitations and threats, particularly climate change (e.g., improving salt tolerance traits to cope with saline water intrusion, stress tolerance traits to withstand extreme environments, etc.), concern about animal welfare (e.g., improving stress tolerant traits), and globalization (e.g., improving disease resistant traits). However, due to a large number of species to work with and short history of development, there are more limitations to tackle on the aquaculture side. To attain a successful genetic improvement program, the studies on the following issues are at priori, characterization of gene pools, development of efﬁ cient breeding techniques, development of cost effective tagging techniques, and characterization of target traits for genetic improvement.

Keywords: Genetic improvement, food security, climate change, animal welfare

1. Introduction A shortage of water supply is one major limitation for World food security is jeopardized because of the enormous enhancement of agricultural products. Agriculture is the increases in food demand while the food production biggest sector (70%) utilizing world water resources (FAO, potential is declining. FAO forecasts that the world 2013a). It is estimated that about 1 m3 of water is required population will reach 9 billion in 2050, and thus will to produce 1 kg of cereal grains, while production of 1 kg demand an increase in food production by 70% (FAO, of beef needs about 13.5 m3 water (Rijberman, 2006). 2009). Livestock and ﬁ sheries products comprise at least While demand for water is increased, water available for 50% of total agriculture products, for which the demands agriculture and human consumption is dramatically are apparently increased. This is not only to cope with the decreased (Ayibotele, 1992). At present many areas of the demand from the hungers but also the expanding popula- world have faced water scarcity, but the situation will be tion. The rapid expansion of urbanization and increasing worse in the future due to many reasons, for example, per capita income have triggered people to change the type population growth, pollution, land reclamation, and of food they consume to more animal protein (FAO, 2009). reduced precipitation (FAO, 2009). There are many constraints towards increasing Lastly, global warming is another important factor agricultural products. For instance, agricultural land is that hampers enhancement of livestock and fisheries gradually diminishing due to various factors. Lambin and products, probably through increased temperature and Meyfroidt, (2011) predicted that within the next 30 years changing amount of precipitation, which will both directly urbanization will result in the loss of agriculture land by affect the animals and indirectly trigger expansion of diseases 1.6 – 3.3 million hectare (Mha)/year while 1-2.9 Mha/year and parasites to new areas (De Silva and Soto, 2009). will not be suitable for agriculture due to land degradation. To achieve the goal of producing enough food for Moreover, at present, as a result of diminishing resources the rapidly expanding world population within the limited of fossil-base fuels, there is a tendency that there will be resources, using genetic improved animals would be a strong competition for land between food crops and promising strategy. Genetic improvement has been proven energy crops (Lambin and Meyfroidt, 2011). to be the most effective approach to increase the yield of *Author for correspondence: fﬁ [email protected] Vol 1. No 2, July-December 2015 Challenges for genetic improvement of livestock and aquatic animals 19

livestock (see Hume et al., 2011). For example, from the and have contributed to the rapid increment of the world 1960’s to 2005, the number of pigs weaned/sow/year has production. However, the contribution to the small holders increased by 50%, while the increment rates have been is less due to many reasons. For instance, farmers could 30% for the number of eggs produced by a laying hen/day, not exploit benefi ts from genetic improvement programs 67% for kilograms of milk/cow/lactation, and the reduction to the maximum potential because a small number of of days until 2 kg of broiler by 60% (reduced from 100 to animals prevents fair or unbiased comparison and selection 40 days) (Hume et al., 2011). Although the genetic for genetic improvement. Moreover, a lack of good improvement programs for aquatic animals are relatively genetic management has caused inbreeding and loss of young compared to those of livestock (Gjerdrem, 2012), genetic variation which eventually results in deterioration the genetically improved strains have tremendously of the breeds (Oltenacu and Broom, 2010). In addition, enhanced world production of some aquatic species, e.g., education background or understanding (selection and Atlantic salmon, Nile tilapia, and Pacifi c white shrimp mating) of the farmers could also cause uncontrolled (Thodesen and Gjedrem, 2006; Gjedrem, 2012). For genetic variation and then slow genetic progress e.g., a case example, the achievement of the GIFT (Genetically of dairy cattle, Holsteinxother breeds multibreed dairy Improved Farmed Tilapia) strain of Nile tilapia population in Central Thailand which showed almost zero (Oreochromis niloticus) was accepted as a major factor for genetic trend (Koonawootrittriron et al., 2009; Sarakul et the expansion of world production (ADB, 2005; Ponzoni al., 2011) et al., 2010), e.g., from 970,756 MT in 2000 to 3,436,526 2.2 Minimizing the adverse impacts of livestock genetic MT in 2013 (FAO, 2015). The Taura syndrome resistant improvement on extinction of local breeds: strain of Pacifi c white shrimp (Litopenaeus vannamei) Although local breeds may be perceived as having low (Gjerdrem, 2012) has triggered an expansion of the culture economic value, they are an invaluable source of useful area of this species around the world and the production traits, for example, tolerance to harsh conditions, tolerance has been increased and reached 3,314,447 MT in 2013 to parasitic and infectious diseases, drought, and poor (FAO, 2015). Interspecifi c hybridization between Thai quality feed. FAO predicted that at present there are 4,500 walking catfish (Clarias macrocephalus) and African major breeds from at least 40 animal species having a sharp-tooth catfi sh (C. gariepinus) (Nukwan et al., 1990) contribution to livestock production. However, it was has caused a rapid increase in Thailand’s annual production predicted that six breeds are dying out every month and 30 by nine fold within 14 years (Na-Nakorn, 2013). percent of the world’s breeds are almost extinct (FAO, 2013b). The scientific evidence was revealed using 2. Challenges for genetic improvement of livestock molecular markers in the Vietnamese Black H’mong pig Despite the enormous past achievements, genetic impro- breed from Ha Giang province that comprised of various vement of livestock in the new era cannot focus on only degrees of genetic admixture from the exotic breeds enhancing production. Rather, the program must be (Landrace and Yorkshire) and it was predicted that after 60 designed to cope with the concerns on various issues, such generations, 100% of Black private alleles will be lost as, enhancing contributions to small-households, sustain- (Berthouly-Salazar et al., 2012). ability of the local breeds, climate change, animal welfare, Despite the risk of losing the local breeds, the lesson and globalization. learnt from genetic improvement of small ruminants in 2.1 Enhancing contributions of the genetic improvement sub-Saharan regions suggested that the cross between local program to small-households: It has been shown that the and exotic breeds is an effective way to improve production genetic improvement programs in the past have benefi ted of livestock (e.g. goat improvement in Africa, Peacock, the intensive commercial farms but had less contribution 2008; Peacock et al., 2011; Mohlatlole et al., 2015). to small-households (Rege et al., 2011). Although the However, the program should be operated on-farm rather intensive commercial system produces a majority of than working solely in a breeding nucleus and that the livestock products (55% pork, 68% eggs, 74% poultry and program should be incorporated into the production system, meat), the small household system of production involves a under close supervision of the experts. large number of smallholders and pastoralists. Thus, it 2.3 Coping with climate change: Livestock plays a role supports livelihood and rural food security. as a contributor to the green-house gases (GHG), while this In general, the characteristic of livestock/poultry sector is impacted by climate change. Genetic improvement genetic improvement in the tropics is crossbreeding and is an effective tool for handling both problems. There are upgrading of local breeds with improved breeds/lines/ scientifi c evidences showing that the emission of greenhouse strains from the temperate region. For example, Holstein gases by livestock could be indirectly reduced by genetic (Black and White cattle; Bos taurus) has been used to improvement, e.g., improvements in growth rate and upgrade crossbreds or native cattle for the improvement of fertility in pigs and dairy cattle in the United Kingdom milk production (Tadesse and Dessie, 2003; Haile et al., resulted in a 0.8% annual reduction in GHG emission while 2009; Koonawootrittriron et al., 2009); Charolais (French improving the feed conversion ratio in broilers and layers creamy cattle; Bos taurus) has been used to cross with reduced the emission of GHG by 1.2 and 1.3 %, respectively Brahman, crossbreds, or native cattle for the improvement (Jones et al., 2008); genetic gain in milk performance has of meat production (Long, 1980). These practices benefi t considerably reduced the environmental impact of diary the intensive commercial livestock production worldwide production in the USA (Capper et al., 2009). 20 Uthairat Na-Nakorn and Skorn Koonawootrittriron R & K

On the opposite corner, it is well understood that animal welfare. Over selection for a single trait may result livestock is impacted by climate change but it is hard to in adverse or unexpected effects on other traits. For example, predict the effect. However, there is a trend that temperature the genetic selection for both rapid growth and high, lean is increasing, e.g. increase of 0.7°C in the past decade meat yield may relate with the appearance of highly (IPCC, 2007), thus environmental adaptability (e.g., heat excitable and diffi cult to handle animals (Grandin, 1994). tolerance) traits will be one of the important desired traits. The increased incidence of the “weaver” condition in Projections suggest that further selection for breeds with Holsteins is possibly related to increased selection for high effective thermoregulatory control will be needed. This milk production; while, broiler chickens have reduced calls for the inclusion of traits associated with cardiopulmonary capacity in relation to their muscle mass environmental adaptability in breeding indices, and more and cannot withstand much physical exertion (Broom, consideration of Genotype-by-Environment interactions 1987; 1993a; Julian, 1993; Julian et al., 1986). (GxE) to identify animals most adapted to specific On the contrary, genetic improvement can promote conditions (Gebryohannes et al., 2015). animal welfare. For example, seven generations of Climate change may result in the spreading selection in laying hens resulted in a strain that shows much of diseases and parasites due to the contact between less feather pecking and cannibalism than a control strain domesticated and wild species that changes their which increased aggressiveness and social dominance distribution because of climate change. This may trigger during adolescence, with no decrease in productivity. the transmission of pathogens or parasites from one species Therefore, this strain does not require beak trimming and to another. For examples, a case of lungworm that is a hence will enjoy a higher level of welfare in cages (Muir common parasite of Dall’s sheep and has never been found and Craig, 1998). in Musk oxen is currently observed in Musk oxen (Kutz et In order to employ genetic improvement programs al., 2004). Moreover, climate change may expand the for this purpose, the traits under selection must be habitats for insects that spread diseases. For example, the precisely identifi ed. These include at least three possible bluetongue disease, which is an insect-borne disease broad categories to consider: behavior, physiology, and infecting ruminants, has spread from Southern to Northern production traits. Europe. Scientists suggested that this may relate with the 2.5 Adaptation to globalization: In the past pathogens rising temperature that may enable the vector to spread to were mostly spread to new areas by the introduction higher latitudes or the rising temperature may enable the of improved breeds and semen for artifi cial insemination. virus to use other species of Culicoides as a vector For example, emergence of porcine reproductive and (Carpenter et al., 2009). respiratory syndrome (PRRS) (Thanawongnuwech et The best adaptation to these problems would be al.,2004a, b) and porcine circovirus–associated diseases through genetic improvement. Therefore, it is urgent to (PCVAD) (Tantilertcharoen et al., 1999) in Asian countries. search for the desired traits, for example, traits that lead to In addition, the movement of livestock can cause the heat tolerance and disease resistance, which potentially emergence of new diseases or re-emergence of fully exist in local breeds. For example, several Latin American controlled diseases. For example, the epidemic of foot and cattle breeds with a very short, sleek hair coat were mouth disease (FMD) in the United Kingdom in 2001 observed to maintain lower rectal temperatures. This is an (Green et al., 2006) and the outbreak of FMD in South East example of a useful genepool for genetic improvement for Asia (Nuntawan Na Ayudhya et al., 2012), were caused by heat tolerance. At present, research in the major “slick hair” the movement of livestock. There is a concern that the gene which is dominant in inheritance and located on uniting of different economic zones may enhance Bovine Chromosome 20 is ongoing (Olson et al., 2003; movement of livestock between countries and eventually Dikmen et al., 2008). However, genetic correlation between may trigger the emergence or re-emergence of diseases, the desired traits and other economic traits should be taken e.g., Wongsathapornchai et al. (2008); Cai (2012). into consideration, e.g., the desired heat tolerance has a Several measures have been established to control these negative correlation with milk yield and reproduction of diseases, and have cost huge amounts of money (e.g. US cattle. Thus heat tolerant dairy cow may not be possible, $ 820 million for global strategy against FMD, FAO, 2012). whereas beef cattle may be improved for heat tolerance Genetic improvement, which shows potential to improve (Prayaga and Henshall, 2005). disease resistance of livestock (Stear et al., 2001), may be 2.4 Enhance animal welfare: Currently the issues on an additional measure to ease the problems. animal welfare are taken more seriously in livestock production. Animal welfare acts were issued in several 3. Challenges for genetic improvement in aquaculture countries to protect animals from inhumane treatments. The culture of aquatic animals, or aquaculture, is following While environments and management are improved to livestock footsteps in that the availability of genetically provide livestock with unstressed conditions throughout improved strains becomes a prerequisite of success, as has their lifespan, genetic improvement is also helpful in this been shown for species like Atlantic salmon, Nile tilapia, regard. and Pacifi c white shrimp. Similar concerns have been In fact, genetic improvement can either compromise raised for aquaculture, however with some diversifi cation. or enhance animal welfare. On the dark side, genetic Due to the different nature of aquatic species and livestock, improvement can have adverse effects that compromise and shorter history of genetic improvement of aquatic Vol 1. No 2, July-December 2015 Challenges for genetic improvement of livestock and aquatic animals 21

animals, more challenges will be faced by scientists who et al., 2005; Mahapatra et al., 2008). In fact, there are many work on aquatic animals. There are many limitations physiological traits that relate to growth, disease resistance, towards genetic improvement of aquatic species, for and other desired traits. These physiological traits may have example, lack of well characterized gene-pools, lack of different levels of heritabilities, and thus, would allow effi cient breeding (producing offspring) techniques in some scientists to improve low heritability traits by selecting for species, lack of effi cient tagging techniques, and lack of the traits with high heritabilities, which positively correlate well characterized targeted traits. with the targeted traits. 3.1 Lack of well characterized gene-pools: Aquaculture is young when compare to livestock production and it covers 4. Genetic improvement as a tool for adaptation to a large number of species. Only a few species are future threats domesticated, among which some species have been Aquaculture also faces threats emerging from similar domesticated regardless of genetic principles. As such, the factors as those faced by livestock, for instance, climate well characterized gene-pools are lacking. Genetic change, globalization, and animal welfare, etc. To utilize improvement always started with pooling of genetic a genetic improvement approach to cope with those threats, variation from wild and farm stocks without knowing suitable traits should be characterized and genetic performances (e.g. genetic improvement programs for parameters regarding those traits should be intensively Pacifi c white shrimp, Argue et al., 2002; Rohu, Labeo studied. rohita in India, Mahapatra et al., 2006; Atlantic salmon in 4.1 Adaptation to climate change by genetic improvement Norway, Thodesen and Gjedrem, 2006) with some exceptions in fi sh: The impacts of climate change to aquatic fauna are (e.g., the GIFT Nile tilapia, Eknath and Acosta, 1998, and not conclusive. However, the impact may be similar to that common carp in Hungary, Bakos et al., 2006). Thus a of livestock to some degree, such as the increase in water chance of including populations with undesired traits is temperature (Poff et al. 2002; De Silva and Soto, 2009) possible. may expand culture areas for warm water species to cold 3.2 Lack of effi cient breeding techniques: A lack of climates and may consequently enhance the spreading of effi cient breeding techniques prevented the species from disease and parasites to the cold water areas. In particular, benefi ting from good genetic improvement. Breeding of due to a rising of sea level, invasion of saline water to aquatic animals requires diverse techniques from natural inland water is predicted (Rahel and Olden, 2008; De spawning to artifi cial breeding. The rearing of larvae and Silva and Soto, 2009). Genetic improvement can help in fry also requires knowledge, specifi c to each species in this regards by improving strains to increased salinity some cases. Due to the large number of species involved tolerance. For example, a salt tolerant strain of tilapia was in aquaculture (FAO, 2015), it is understandable that certain developed by interspecifi c hybridization, e.g., hybridization species still do not have effi cient breeding and rearing between the genetically improved strain of Nile tilapia, O. technologies. This can easily lead to unnoticed mis-man- niloticus with a salt-tolerant species, O. mossambicus agement of broodstock. For example, despite the large (Villegas 1990; Garcia-Ulloa et al., 2001) or O. spilurus number of brooders kept by farmers, only a few pairs (Ridha, 2012). contribute to the next generation due to limited success of Climate change potentially enhances eutrophication breeding techniques, and thus it causes a loss of genetic in tropical inland waters and also triggers extreme climate variation and eventually results in inbreeding. Similarly, conditions (De Silva and Soto, 2009). As such, genetic communal spawning practiced in some species, e.g. sea improvement for stress tolerance may lead to fi sh strains bream (Brown et al., 2005) and groupers (Wachirachaikarn that can withstand fl uctuating/extreme culture conditions. et al., 2011), has led to a reduction in the effective population The potential of this purpose is discussed in the later section size. on “Genetic improvement to enhance animal welfare”. 3.3 Lack of efficient tagging techniques: A lack of 4.2 Genetic improvement to cope with transboundary effi cient tagging techniques has caused diffi culty in employing translocation of aquatic animals: Globalization effi cient genetic improvement methods. For example, to obviously enhances intensive translocation of aquatic select based on family performance, single families must animals for commercial culture. Although this practice be separately reared, which requires unaffordable benefi ts aquaculture industries, it also possesses severe infrastructure in most cases. However, with the assistance threats to local biodiversity and also on the aquaculture of of molecular markers and parentage assignment computer native species through disease transfer. For example, the software, families can be communally reared and outbreak of herpes virus in fancy carp (Cyprinus carpio) identifi cation is performed at measurement (e.g. Norris et and other native cyprinid fi shes was caused by the introduction al., 2000; Sekino and Hara, 2007; Karaket et al., 2011; of fancy carp from Japan (Chansu and Tangtrongphairot, Lafarga-de la Cruz). This surely requires a high cost. 2005); the outbreak of Taura syndrome virus in tiger prawn 3.4 Lack of well characterized targeted traits: Genetic in 2004 was a result of the introduction of Pacifi c white improvement of aquatic animals mostly focused on a few shrimp (L. vannamei) which is native to South America traits, such as growth and disease resistance, for which a (Limsuwan and Chucherd, 2007); and the outbreak of Noda majority of selection programs emphasized simple traits, and Extrasmall viruses in giant freshwater prawn such as growth rate, survival rates, or time to death after (Macrobrachium rosenbergii) in Thailand was caused by disease challenging (e.g. Na-Nakorn et al., 1994; Henryon the introduction of conspecifi c exotic stocks for genetic 22 Uthairat Na-Nakorn and Skorn Koonawootrittriron R & K improvement (Chucherd et al., 2007). Measures have been are challenged with similar problems. Thus, they should established to prevent these unwanted circumstances, combine their efforts and exchange experiences so that however, other approaches should also be considered. For genetic improvement of aquatic animals, which has a much example, genetic improvement has high potential to shorter history, can be progressed without repeating the increase resistance to diseases as indicated by moderate to mistakes from livestock. At the same time, assistance from high heritability (Ødegård et al., 2011). However, more people of different areas of expertise, especially in the fi eld parameters that closely relate to resistance must be of physiology and behavioral science, is crucially needed identifi ed and employed in the selection program, e.g., to enhance the precise identifi cation of target traits and data immune related traits. In addition, selection based on collection for genetic improvement programs. molecular genetic information is a new approach that can enhance the effi ciency of the selection program (Ødegård References et al., 2011). ADB. 2005. An Impact Evaluation of the Development of 4.3 Genetic improvement to enhance animal welfare: Genetically Improved Farmed Tilapia and Their Although the issue on animal welfare is not yet strongly Dissemination in Selected Countries. The Asian addressed in aquatic animals, in the near future this issue Development Bank, Manila, Philippines, pp. 137. will surely affect aquaculture, especially the species Argue, B. J., Arce, S. M., Lotz, J. M. and Moss, S. M. involved in international trade. Actually the traits related 2002. Selective breeding of Pacifi c white shrimp to welfare also affect production, for example, stress (Litopenaeus vannamei) for growth and resistance response. Stress in aquaculture occurs from transportation, to Taura Syndrome Virus. Aquaculture 204, 447-460. handling, harvesting, water quality change, crowding, and Ayibotele, N. B. 1992. The World’s Water. Keynote paper, diseases, etc. Even though there was limited information International Conference on Water and the Environ- on selection potential of physiological traits, there was a ment, Dublin, Ireland, 26-31 January 1992, pp. 26. concrete evidence that supported the genetic improvement Bakos, J., Varadi, L., Gorda, S. and Jeney, Z. 2006. Lessons potential of these traits, e.g., signifi cant heritability of from the breeding program on common carp in primary (cortisol) and secondary (glucose, osmolality, and Hungary. In: Ponzoni, R.W., Acosta, B.O. and haematocrit) stress responses (mean heritability, Ponniah A.G. 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Current Topics in the great contribution from livestock genetic improvement Veterinary Medicine and Animal Science 42, programs, their adverse impacts, e.g. less contribution to 101-110. small scale farmers and enhancement of local breed extinction, Brown, R. C., Woolliams, J. A. and McAndrew, B. J. 2005. etc., have been recognized and become challenges for Factors influencing effective population size in scientists in this fi eld. The additional challenges are to commercial populations of gilthead seabream, effi ciently employ genetic improvement to cope with the Sparus auratus. Aquaculture 247, 219-225. problems of animal welfare, emissions of GHG, and Cai, C. 2012.The risk of foot and mouth disease entering adaptation to adverse impacts of globalization. While in China through the movement of animals from Upper the aquaculture sector, genetic improvement was proven Mekong region countries. Ph.D. thesis, Murdoch to enhance global production of some aquatic species. It University. Available on web http://researchre also has high potential to overcome the upcoming pository.murdoch.edu.au/13598/ limitations and threats, particularly climate change, concern Capper, J. L., Cady, R. A. and Bauman, D. E. 2009. The for animal welfare, and globalization. However, due to the environmental impact of dairy production: 1944 large number of species to work with and short history of compared with 2007. Journal of Animal Science 87, development, there are more limitations to tackle on the 2160-2167. aquaculture side. To attain a successful genetic improvement Carpenter, S, Wilson, A. and Mellor, P. S. 2009. Culicoides program, the studies on the following issues are at priori, and the emergence of bluetongue virus in northern characterization of gene pools, development of effi cient Europe. Trends in Microbiology 17(4), 172-178. breeding techniques, development of cost effective tagging Chansu, N. and Tangtrongphairot. C. 2005. Survey Koi techniques, and characterization of target traits for genetic Herpes Virus (KHV) disease of fancy carp improvement. (Cyprinus carpio Linn.) in Bangkok and perimeter It is quite clear that the scientists who have been in March - June 2005. Thai Veterinary Medical working on livestock and aquatic animal genetic improvement Association 56 (3), 13-21. Vol 1. No 2, July-December 2015 Challenges for genetic improvement of livestock and aquatic animals 23

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Clive Burrett1, 3, *, Hathaithip Thassanapak2 and Mongkol Udchachon2, 3 1School of Physical Sciences, University of Tasmania, Sandy Bay, Box 79, Hobart, Tasmania, 7001 Australia 2Applied Palaeontology and Biostratigraphy Research Unit, Department of Biology, Faculty of Science, Mahasarakham University, Kantarawichai, Mahasarakham 44150, Thailand 3Palaeontological Research and Education Centre, Mahasarakham University, Kantarawichai, Mahasarakham 44150, Thailand

(Received 17 October 2015; accepted 18 December 2015)

Abstract - Conodonts from radiolarian cherts in the Loei Terrane of Loei Province, Northeast Thailand, include Palmatolepis triangularis, P. minuta minuta, P. minuta wolskae and polygnathids and indicate a Famennian crepida Zone age. These cherts are about 20 my younger than Givetian reef limestones and are unlikely to have been deposited in a major ocean but rather in a deep marine basin close to a volcanic arc.

Keywords: Conodont, Devonian, palaeoenvironment, tectonics, Indochina Terrane, chert.

1. Introduction cherts were originally thought to have been deposited The Loei Terrane is a complex orogenic belt that constitutes contemporaneously with the Givetian reefs, with the the western margin of the Indochina Terrane and extends limestones later forming synclinal cores (Chairangsee et al., from Laos southwards to Thailand and then swings 1990), it is now known that all reliable radiolarian chert eastwards into Cambodia (Fig.1), (Burrett et al., 2014). dates are Upper Devonian or Tournaisian (Sashida et al., The Loei Terrane contains assemblages of igneous, 1993; Saesaengseerung et al., 2007 a, b; Udchachon et al., metamorphic and sedimentary rocks ranging from Silurian 2011). Mainly because of severe outcrop limitations, to Triassic in age and with deformation in the Palaeozoic structural studies have not been carried out and it is and Triassic (Bunopas, 1982; Khin Zaw et al., 2014). possible that the younger cherts are either faulted against In Loei Province, NE Thailand, Devonian sedimentary or folded with the older limestones (Fig. 3). rocks (Fig. 2) were deposited close to an active volcanic arc that was initiated in the Late Silurian and extended to 2. Samples and results the Triassic (Intasopa and Dunn, 1994; Kamvong et al., We have sampled the radiolarian chert (Fig. 4) using standard 2014; Khositanont et al., 2008; 2013; Panjasawatwong HF treatment techniques. Samples yield abundant Upper et al., 2006; Salam et al., 2014). Givetian patch reefs are Devonian radiolarians, sponge spicules and sample CNA found as north-south trending belts which now outcrop at 4 also contains moderately well preserved, though corroded the summit of hills (Fontaine and Tantiwanit, 1987; and pitted, conodonts. The conodont fauna consists of Fontaine, 1990; Fontaine and Sutheethorn, 2000; Fontaine abundant Palmatolepis and less common Polygnathus. et al., 2005). The Givetian reefs overlie, thin bedded Palmatolepis triangularis (Fig. 5a) ranges from the limestone containing a Givetian conodont fauna which in triangularis to the lower crepida Zone, and P. minuta turn conformably overlies fossiliferous shales, siltstones, wolskae (Fig. 5c) and P. minuta minuta (Fig. 5b, d) range fi ne quartz arenites and minor volcaniclastics (Fig. 3). The through the crepida Zone (Austin et al., 1985; Ziegler, fi ne siliciclastics yield trilobites (a harpetid, a phacopid 1977; Ziegler and Sandberg, 1990). A crepida Zone age is and proetids), abundant brachiopods (orthids and large therefore indicated for radiolarian chert sample CNA 4. rostrate strophomenids), gastropods, bivalves and rare A Polygnathus with a broad elliptical platform is tentaculitids and represent a benthic assemblage that lived in present (Fig. 5 f, g) and is close to P. vialovi Zvereva, 1986, moderately deep shelf seas. Radiolarian cherts are widespread from the Upper Devonian of the Russian Platform (Ziegler in the lowland areas, occur as scattered outcrops and are et al., 2000; Zvereva, 1986). Another Polygnathus (Fig. 5 h, i) strongly and disharmonically folded (Fig. 4). In many is similar to the Upper Devonian P. dengleri Bischoff and localities, cherts are closely associated with mafi c volcanic Ziegler, 1957. Our specimens are closest to those from Alaska rocks and in a few localities with rhyolites. Although, these identifi ed as P. aff. P. dengleri by Savage and Funai (1980).

*Author for correspondence: [email protected] Upper Devonian (Famennian) conodonts from radiolarian cherts, Vol 1. No 2, July-December 2015 Loei Terrane, Loei Province, Northeast Thailand 27

Figure 1. Sketch map showing extent of the Loei Terrane in SE Asia (after Burrett et al., 2014). 28 Clive Burrett et al R & K

Figure 2. Simpliﬁ ed geological map of Loei, NE Thailand showing the distribution of sedimentary sequences and volcanic rocks. The study section is located to the east of Loei. 1–Permo–Triassic volcanic rocks including rhyolite, andesite, tuff, agglomerate; 2–Carboniferous rocks including conglomerate, sandstone, shale, slate, chert and limestone; 3–Permo–Triassic rocks including sandstone, argillaceous limestone, rhyolitic tuff, shale, limestone, and chert; 4–Permo– Triassic granite; 5–Permian rocks including limestone, shale, sandstone; 6–Triassic rocks including sandstone, siltstone, mudstone; 7–Devonian rocks including chert, shale and tuff; 8–Devonian–Carboniferous volcanic rocks including basalt, andesite, tuff; 9–thrust fault; 10–road, 11– border between Lao PDR and Thailand; 12– study locality at Ban Chom Noi, Pak Chom District, Loei Province (after Udchachon et al., 2011).

Figure 3. Generalised stratigraphic column showing postulated fault contact between Givetian shallow marine sedimentary rocks and Upper Devonian conodont-bearing radiolarian chert. Upper Devonian (Famennian) conodonts from radiolarian cherts, Vol 1. No 2, July-December 2015 Loei Terrane, Loei Province, Northeast Thailand 29

Figure 4. Photograph of disharmonically folded Upper Devonian chert at Ban Chom Noi, Pak Chom District, Lat.17° 45’ 49.42” N, Long. 101° 56’ 18.35” E, showing sampling points. Geological hammer as scale.

3. Palaeoecology and palaeoenvironment Inthanon Terrane of northern Thailand (Randin et al., The conodonts belong to the basinal, but not deepest basin, 2006). However, on the basis of chert geochemistry, Palmatolepis-Polygnathus biofacies (Sandberg and Udchachon et al. (2011) conclude that the Loei cherts were Dreesen, 1984; Dreesen et al., 1986; Savoy and Harris, not deposited in a major ocean basin but rather in a basin 1993). Many of the Palmatolepis specimens are small and relatively close to a continental margin. Adjacent and were probably juvenile, which may indicate deposition at interbedded maﬁ c and less common, rhyolitic volcanics a considerable distance from land, as was suggested for and volcaniclastics suggest proximity to a volcanic arc. Upper Devonian conodonts from radiolarian cherts in the 30 Clive Burrett et al R & K

Figure 5. Conodonts from sample CNA 4. All illustrated specimens are stored in the collections of the Palaeontological Research and Education Centre, Mahasarakham University, Mahasarakham, Thailand and given PRC numbers. Scale bar = 20 μm. (a) Palmatolepis triangularis individual ﬁ gured specimen number PRC 250 (total of 38 specimens for this species, including all non-Pa elements from this sample). (b) Palmatolepis minuta minuta. Figured specimen number PRC 251(15 specimens total for this taxon). (c) Palmatolepis minuta wolskae. Figured specimen number PRC 252 (20 specimens total for this taxon). (d) Palmatolepis minuta minuta. Figured specimen number PRC 253. (e) Palmatolepis sp. ‘juvenile’ form. Figured specimen number PRC 254 (63 specimens of unassigned ‘juvenile’ Palmatolepis). (f) Polyg- nathus aff. vialovi Zvereva, 1986. Figured specimen number PRC 255 (6 specimens total for this taxon). (g)Polygnathus aff. vialovi Zvereva, 1986. Figured specimen number PRC 256. (h) Polygnathus aff. dengleri Bischoff and Ziegler, 1957. Figured specimen number PRC 257 (14 specimens total for this taxon). (i) Polygnathus aff. dengleri Bischoff and Ziegler, 1957. Figured specimen number PRC 258. Upper Devonian (Famennian) conodonts from radiolarian cherts, Vol 1. No 2, July-December 2015 Loei Terrane, Loei Province, Northeast Thailand 31

4. Conclusions Vachard, D. 2005. Sedimentary rocks of the Loei The Upper Devonian to Tournaisian age of the Loei Region, Northeast Thailand: stratigraphy, paleontol- radiolarian cherts is confi rmed by the conodonts identifi ed ogy, sedimentology. Bureau of Geological Survey, herein. This shows that the radiolarian cherts were Department of Mineral Resources, Bangkok, pp. 165. deposited neither before nor contemporaneously with Fontaine, H. and Sutheethorn, V. 2000. Devonian and shallow water Givetian reefs but rather represent Carboniferous corals found at Ban Na Klang, Loei widespread deep water conditions about 20 million years province, Northeast Thailand. Journal of the Geo- after the Givetian (Becker et al., 2012). The Famennian logical Society of Thailand 1, 27 – 33. cherts were deposited in a deep water basin but not in a Fontaine, H. and Tantiwanit, W. 1987. Discovery of wide- major ocean basin. spread and very fossiliferous Devonian beds in Northeast Thailand. CCOP Newsletter 12, 25-26. Acknowledgements Intasopa, S. and Dunn, T. 1994. Petrology and Sr-Nd iso- We thank Professors Chongpan Chonglakmani and Nat- topic systems of the basalts and rhyolites, Loei, thawirpj Silatana for their helpful reviews. Thailand. Journal of Southeast Asian Earth Sciences 9, 167-180. References Kamvong, T., Khin Zaw, Meffre, S., Maas, R., Stein, H. Austin, R., Orchard, M. and Stewart, I. 1985. Conodonts and Lai, C. 2014. Adakitic porphyries in the Loei and of the Devonian System from Great Britain. In: Higgins, Truongson Belts, Thailand and Laos: Their genesis A. and Austin, R. (Eds.), A Stratigraphical Index of and implications for geodynamics and metallogeny. Conodonts. Ellis Horwood, Chichester, pp. 93-166. Gondwana Research 26, 115-184. Becker, R. T., Gradstein, E. M., and Hammer, O. 2012. The Khin Zaw, Meffre, S., Lai. C., Burrett, C., Santosh, M., Devonian Period. 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Marc Philippe1, *, Pratueng Jintasakul2, Nareerat Boonchai2 1Université Lyon-1 and UMR5023 of the CNRS, 69200, France 2Northeastern Research Institute of Petriﬁ ed Wood and Mineral Resources (Nakhon Ratchasima Rajabhat University), 184 Moo 7, Suranaree subdistrict, Mueang, Nakhon Ratchasima 30000, Thailand

(Received 19 October 2015; accepted 21 December 2015)

Abstract- Despite its limited stratigraphical value, fossil wood has been revealed to be of interest to characterize the palaeoenvironment evolution during the sedimentation of the Khorat Group. It helps understand the ecosystems in which the rich Thai Dinosaur faunas evolved. The available Khorat group fossil wood record was, however, mostly from the northeastern Thailand (Isan). New data are presented here, from the southwestern Isan provinces of Chaiyaphum and Nakhon Ratchasima. Although 17 new samples were studied, specific diversity was not increased, which suggests that the taxonomical investigation of the Khorat wood record is quite complete. Fossil wood from the Phu Phan Formation is reported for the ﬁ rst time, and assigned to the genus Shimakuroxylon. Despite being sedimentologically similar, the Phra Wihan and Phu Phan formations seem to yield different woods, which suggests that they may have had different palaeoecology or provenance.

Keywords: Fossil wood, Mesozoic, Thailand, palaeobiogeography, taxonomy

1. Introduction the Phu Kradung Formation. The Waritchapum Formation In 2004, a synthesis was published about the fossil woods is mostly individualised in the Phu Phan Range and in of the Mesozoic Khorat Group in north-eastern Thailand seismic data (Mouret et al., 1993), but it is much more (Philippe et al., 2004). This synthesis focused on the diffi cult, if not impossible, to locate accurately in the fi eld stratigraphy and palaeobiogeography. It revealed a medi- or outside this range (V. Suteethorn, personal communica- um-rich record, with four taxa, as well as a wood diversity tion). All wood data published from the Waritchapum decreasing over time, from the Phu Kradung Formation Formation in 2004 has been attributed to the Phu Kradung upward to the Sao Khua Formation. It also evidenced strong Formation. The stratigraphical framework is given in Table biogeographical affi nities with the coeval Indochina (Lao, 1. Wood was collected from six localities (Fig. 1). We tried Cambodia, Vietnam) fossil wood record. The data base for to focus our sampling effort on those formations that are this fi rst synthesis, however, was mostly based on samples little-known from the fossil wood point of view. collected in central Isan, with most of the samples originating Fossil wood specimens MP 1925 to 1930 were from the Upper Phu Kradung Formation in Phu Phan collected as weathered samples on the surface of a Nam Range. Phong Formation outcrop, at Phu Nokkhien, near Nong We collected and investigated more wood samples Buadang, northwest of Chaiyaphum (E776022, N1780406, from the Mesozoic Khorat group in Chaiyaphum and Z47Q). Fossil wood pieces are rounded, pebble to gravel-like, Nakhon Ratchasima Provinces (southern Isan), and tried and associated with other rounded pieces of hard stones to focus on poorly sampled levels. Seventeen samples were like cherts and quartzite. It is not certain if these pebbles studied, including two from the Phu Phan Formation for and gravel resulted from the alteration of a conglomeratic which no wood data has been reported yet. layer within the Nam Phong Formation, or if they are These new results are integrated in a reappraised alterite residues on the surface and from a remote taxonomical framework, as systematical novelties have provenance. They might also have been reworked from been published since the 2004 study (Philippe et al., 2011, underlying formations into the Nam Phong Formation. 2014). These are the fi rst fossil woods documented from the Nam Phong Formation. Unfortunately, however, they were found 2. Material and method to be deeply recrystallized, with no preserved wood The same stratigraphical framework as in 2004 is used here, structure. This recrystallization is probably a sign of some apart from the Waritchapum Formation being merged into reworking.

Author for correspondence: [email protected] 34 Marc Philippe et al R & K

At the Petriﬁ ed Wood Museum in Nakhon Ratchasima Nong Ped, Thayai subdistrict (E785248, N1772021, Z47P). we sampled a Nam Phong Formation fossil wood given by This wood, MP1932, was somewhat better preserved but villagers, which was found alongside the Chi River, in Ban could still only be tentatively identiﬁ ed.

Figure 1. Geological map of Chaiyaphum and Nakhon Ratchasima Provinces, Thailand (Department of Mineral Resources, 2007) with studied petriﬁ ed wood sites. New fossil woods from the Mesozoic Khorat Group of Thailand Vol 1. No 2, July-December 2015 and their palaeoecological implications 35

EXPLANATION In the Phu Kradung Formation, a fossil wood was collected at Khao Noi fossil wood site, near the Lam Sedimentary and Metamorphic rocks Takhong Dam, Pakchong District, Nakhon Ratchasima

Alluvial deposits. Province (E810953, N1605454, Z47P). The specimen (sample MP1924) is a piece of silicifi ed wood, which was Terrace deposits. found isolated in the material from a fi sh pond excavation, Colluvium deposits and weathering deposits. about 100 m NW of a shrine that shelters a large fossil log Mudstone, siltstone and sandstone, with rock salt. identifi ed as Agathoxylon sp. (Songtham et al., 2011). At the Northeastern Petrified Wood Museum in Nakhon Sandstone and siltstone, with lime nodule conglomerate. Ratchasima we sampled fossil woods (samples MP1933 Sandstone and siltstone. and 1934) that were collected at the Ban Pong Din Dam, Siltstone and sandstone; partly calcareous. Pakchong District, Nakhon Ratchasima Province (E787498, N1620835, Z47P). We collected samples from Quartzitic sandstone, sandstone. the base of the southern cliffs of the Phu Khieo Wildlife Siltstone and mudstone, some lime-nodules. Sanctuary, Nang Daet Subdistrict, Nong Bua Daeng Dis- Basal conglomerate, clasts of limestone, sandstone and volcanic; trict, Chaiyaphum Province (E765512, N1799869, Z47P). calcareous mudstone. Fossil wood is abundant there, including large logs and we sampled fi ve specimens: MP 1936-1940 (Fig. 2). Limestone, chert, shale, sandstone and volcanic rocks. In the Phu Phan Formation, at the temple Wat Shale, siltstone, sandstone, chert, schist and hornfels. Tampekah (E762251, N1692708, Z47P), we studied a large Sandstone, conglomerate, siltstone. fossil wood donated by a villager. The wood was not Volcanic sandstone, chert, basalt. sampled as a hand-lens revealed that it is a poorly preserved homoxylous tracheidoxyl. On Phu Kao Hill, Ban Bung

Prue, Thepharak District, Nakhon Ratchasima Province Igneous rocks (E763296, N1695993, Z47P), we studied several wood Olivine basalt, nepheline and basanite pieces among which we selected samples MP1931 and Granodiorite, diorite and monzodiorite. MP1935. All these wood pieces apparently originated from a large fossil log that was broken into numerous parts, Rhyolite, andesite, tuff, volcanic breccia. although it was impossible to be sure that only one log was involved. For anatomical studies, thin sections were prepared transversely, longitudinally radially and tangentially by Source : Dept. of Mineral Resources, Thailand,2007 standard methods (Hass and Rowe, 1999). Figure 1. Geological map of Chaiyaphum and Nakhon Ratchasima Provinces, Thailand (Department of Mineral Resources, 2007) with studied petriﬁ ed wood sites. (Cont.) Table 1. Partial stratigraphy of Mesozoic Khorat Group, Northeastern Thailand (adapted from Philippe et al., 2014). 36 Marc Philippe et al R & K

Figure 2. Sample MP1937 from the base of the southern cliffs of the Phu Khieo Wildlife Sanctuary, Chaiyaphum Province. Note that only the darkest part on the left of the fossil wood was cropping out.

3. Results It cannot help, thus, to decipher the origin of these The results of our xylological investigation are summarized weathered samples on the surface of the Nam Phong in Table 2. The woods from the Nam Phong Formation are Formation outcrops. It is possible that this sample was mostly deeply recrystallized. Their xylological features are actually reworked from older strata within the Nam Phong not preserved, except for the general cross-section pattern. Formation, as quite often happens for silicifi ed wood Only in MP1932 could some radial tracheid pitting and (Philippe et al., 2000). cross-fi elds be observed. They fi tted with those of genus More interestingly, all the specimens from the Phu Agathoxylon Hartig, although not enough pits were Kradung Formation were assigned to the recently described observed to make a certain identifi cation. This wood type Shimakuroxylon japonicum (Shimakura) Philippe, Boura is a very common one in the fossil wood record from the Oh et Pons (Philippe et al., 2014). It is the same taxon that Palaeozoic upward to the Late Cretaceous (Philippe, 2011). was determined from the Phu Phan Formation (Fig. 3).

Table 2. Results of taxonomical investigations. Sample numbers are those in the Philippe’s collection in the Collections paléobotaniques del’Université Lyon-1, Villeurbanne, France.

Sample Formation Identiﬁ cation MP1924 Phu Kradung Homoxylous tracheidoxyl MP1925 Nam Phong Homoxylous tracheidoxyl MP1926 Nam Phong Homoxylous tracheidoxyl MP1927 Nam Phong Homoxylous tracheidoxyl MP1928 Nam Phong Homoxylous tracheidoxyl MP1929 Nam Phong Homoxylous tracheidoxyl MP1930 Nam Phong Homoxylous tracheidoxyl, biseriate rays MP1931 Phu Phan Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons MP1932 Nam Phong cf. Agathoxylon MP1933 Phu Kradung Homoxylous tracheidoxyl MP1934 Phu Kradung Homoxylous tracheidoxyl MP1935 Phu Phan Homoxylous tracheidoxyl MP1936 Phu Kradung Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons MP1937 Phu Kradung Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons MP1938 Phu Kradung Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons MP1939 Phu Kradung Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons MP1940 Phu Kradung Shimakuroxylon japonicum (Shimakura) Philippe, Boura Oh et Pons New fossil woods from the Mesozoic Khorat Group of Thailand Vol 1. No 2, July-December 2015 and their palaeoecological implications 37

4. Discussion and conclusion Despite the 17 newly investigated samples, seven of which were assigned to a fossil taxon, the specifi c diversity has not increase as compared to the 2004 record. This suggests that the taxonomical investigation of the Khorat wood record is quite complete, at least for the Phu Kradung Formation (Table 3). The report of S. japonicum from the Phu Phan For- mation has a double interest. First, it is the fi rst identifi cation of a wood from the Phu Phan Formation, which was not xylologically documented up to now. Second, it shows that after an apparent parenthesis this taxon reappeared in the Khorat Basin by the late Early Cretaceous. Although the fossil wood record of the Phra Wihan and Sao Khua is limited, it does not seem to have included S. japonicum. If this does not result from a taphonomical bias, then the absence of S. japonicum is probably best explained by palaeoecological reasons. While the Phu Kradung Formation has a relatively diverse wood fl ora, the Phra Wihan Formation and Sao Khua Formation records are limited to the genus Agathoxylon. It was hypothesized that this wood diversity impoverishment was due to a palaeoecological shift towards drier conditions. This, in turn, could have resulted from a decrease in the local rainfall, an increase in the evapo-transpiration rate or a decrease in the external water drainage into the Khorat Basin. Whatever the reason, the occurrence of S. japonicum in the Phu Phan Formation points to a return to wetter conditions. Indeed this taxon Figure 3. Typical radial tracheid pitting for Shimakuroxylon was bound to year-round warm and at least seasonally wet japonicum (Shimakura) Philippe, Boura Oh et Pons. Scale bar is climates (Philippe et al., 2014). 150 μm.

Table 3. Wood taxa recognized for the Khorat Group (adapted and completed from Philippe et al., 2004).

Phu Kradung Phra Wihan Sao Khua Phu Phan Formation Formation Formation Formation Agathoxylon saravanensis +++ Brachyoxylon orientale + Brachyoxylon serrae + Shimakuroxylon japonicum ++

It is speculated that changes in the sedimentary facies only driver in the sedimentary changes. Changes in the between the various formations of the Khorat Group reﬂ ect sediment provenance, palaeoecological differences induced mostly regional climatic variations, and then the wood by changes in the accommodation space or subsidence as record of the thick white-sandstone dominated Phu Phan well as more subtle changes in the biosphere may also have Formation should be similar to that of the Phra Wihan played a signiﬁ cant role. Formation. Indeed there is a striking parallel between the abrupt vertical transition from the Phu Kradung Formation Acknowledgments mixed siliclastic red-beds to the Phra Wihan massive white V. Suteethorn and C. Mouret are warmly acknowledged sandstones and the similar transition from the Sao Khua for so generously sharing their knowledge of the Khorat Formation red beds to the white sandstones of the Phu Phan Group. Thanks to Suthep Thipchote, a teacher from Nang- Formation (Booth and Sattayarak, 2011). The discrepancy daet Wangchomphu Wittaya School, Chaiyaphum Province between the Phra Wihan Formation fossil wood record, for guiding us the localities in 2011. Kraisri Tenpuksee, which is limited to Agathoxylon saravanensis, and the Phu Nakhon Ratchasima Rajabhat University, kindly prepared Phan Formation fossil wood record, which is limited to S. the location map. japonicum, suggests that climatic variations were not the 38 Marc Philippe et al R & K

References Booth, J. and Sattayarak, N. 2011. Subsurface Carbonifer- ous –Cretaceous geology of north-eastern Thailand. In: Ridd, M. F., Barber, A. J. and Crow, M. J. (Eds), The geology of Thailand. Geological Society Lon- don, pp. 185-222. Department of Mineral Resources. 2007. Geological map of Thailand, Scale 1:2,500,000. Ministry of Natural Resources and the Environment of Thailand. Hass, H. and Rowe, N. P. 1999. Thin section and wafering. In: Jones, T.P. and Rowe N. P. (Eds.), Fossil Plants and Spores: Modern Techniques. Geological Society London, pp. 76-81. Mouret, C, Heggemann, H, Gouadain, J. and Krisadasima, S. 1993. Geological history of the siliciclastic Mesozoic strata of the Khorat Group in the Phu Phan Range area, northeastern Thailand. In: Thanasup- thipitak T. (Ed.). Proceedings of the International Symposium on the biostratigraphy of mainland Southeast Asia: Facies and palaeontology. Chiang Mai University, pp. 23-49. Philippe, M. 2011. How many species of Araucarioxylon? Comptes Rendus Palevol 10, 201-208. Philippe, M., Szakmany, G., Gulyas-Kis, C. and Jozsa, S. 2000. An Upper Carboniferous-Early Permian silicifi ed wood in the Miocene conglomerate, from the Western Mecsek Mts. (Southern Hungary). Neues Jahrbuch für Geologie und Paläontologie, Monatshefte 2000 (4), 193-204. Philippe, M., Suteethorn, V., Lutat, P., Buffetaut, E., Cavin, L., Cuny, G. and Barale, G. 2004. Stratigraphical and palaeobiogeographical signifi cance of fossil wood from the Mesozoic Khorat Group of Thailand. Geological Magazine 141 (3), 319-328. Philippe, M., Suteethorn, V. and Buffetaut, E. 2011. Une nouvelle espèce du genre Brachyoxylon dans le Crétacé inférieur d’Asie du Sud-Est. Géodiversitas 33 (1), 25-32. Philippe, M., Boura A., Oh, C. and Pons, D. 2014. Shimakuroxylon a new homoxylous Mesozoic wood genus from Asia, with palaeogeographical and palaeoecological implications. Rev. Palaeobotany Palynology 204, 18-26. Songtham, W., Grote, P. J. and Chitnarin, A. 2011. Field excursion guide book for Route 2. World Conference on Palaeontology and Stratigraphy, Nakhon Ratcha- sima, 27 November – 2 December, pp. 17. Wang, Y., Zhang, W., Zheng, S., Jintasakul, P., Grote, P. J. and Boonchai, N. 2006. Recent advances in the study of Mesozoic–Cenozoic petrifi ed wood from Thailand. Progress in Natural Science 16, 501–506. Vol. 1 No. 2 page 39-45 2015 DOI:10.14456/randk.2015.12 RESEARCH & KNOWLEDGE Research Article Development of intake manifold for Student Formula CMU F-914 using theoretical and simulated designs

Niti Kammuang-lue* and Worawit Borikhut Department of Mechanical Engineering, Faculty of Engineering, Chiang Mai University, Chiang Mai, Thailand

(Received 19 November 2015; accepted 25 December 2015)

Abstract - The objectives of this study were to design, develop and construct an intake manifold for the Student Formula CMU F-914 race car, or the F-914, by using theoretical analysis together with simulation software and experimental verifi cation of the design results. Then to compare the pressure drop in the intake manifold of the F-914, which was designed in this study, and the intake manifold of the previous race car, the F-813. The designed intake manifold consisted of an ABS restrictor with the narrowest passage located at 25% of the restrictor’s length measured from the inlet, a carbon fi ber surge tank with internal volume of 1,558 cm3, aluminum elliptical profi le bellmouths, and aluminum runners with a length of 102 mm. The experimental verifi cation showed that after the pressure drop across the throttle valve was considered in the simulation, the pressure drop from the simulation was almost the same as the one from the experiment for both intake manifolds. Moreover, it was found that the intake manifold of the F-914 had lower pressure drops than those of the F-813 by 43%, 94%, 66%, 69%, and 29% at engine speeds of 6,000, 7,000, 8,000, 9,000, and 10,000 rpm, respectively. Therefore, the theoretical and simulated designs presented in this study had acceptable reliability and they could be practically used for designing the intake manifolds.

Keywords: Intake manifold, restrictor, bellmouth, surge tank, runner, Student Formula

1. Introduction economy event, measuring the fuel consumption of the The Society of Automotive Engineers Thailand (TSAE) race car during the endurance event. The total score for the annually hosts a national racing competition for students, dynamic events is 675. Then (2) is three static events which is called the TSAE Auto Challenge Student consisting of (i) engineering design, (ii) cost report, and Formula (Society of Automotive Engineers Thailand, (iii) business presentation, for which the total score is 325. 2015), to promote the students’ abilities in design, research, In the past, the CMU Auto Club, a student activity club in development, and construction of various parts and the Department of Mechanical Engineering, Faculty of equipment installed in high performance race cars, and to Engineering, Chiang Mai University, has participated in give opportunities for the students to use their knowledge the competition every year. In the previous season, obtained in the classroom in the field of automotive 2014-2015, the CMU Auto Club sent a race car to join the engineering to build their own practical race car. The competition with the name of “Student Formula CMU competition and also all the race cars are restricted by the F-914,” as shown in Figure 1. SAE Student Formula rules (2014 SAE Student Formula Rules, 2013), which are globally accepted. The competition, which has a total score of 1,000, consists of two-major events as follows: (1) dynamic events consisting of ﬁ ve tests as (i) acceleration event, measuring the acceleration of the race car in a straight line; (ii) skid-pad event, testing for dynamic stability of the car; (iii) autocross, evaluating the car’s maneuverability and handling qualities on a tight course; (iv) endurance, evaluating the durability, reliability, strength, and overall performance of the race car and the Figure 1. Student Formula CMU F-914. endurance and consciousness of the driver; and (v) fuel

*Corresponding Author’s E-mail: [email protected] 40 Niti Kammuang-lue and Worawit Borikhut R & K

The primary design concepts of the Student limited time-consumption and convenience. However, the Formula CMU F-914 (referred to as “F-914”) were reliability of the design result is not certain. The result lightweight components and agile movement developments should be verifi ed before being applied to practical uses. and modifi cations on the predecessor race car —“the It can be seen from the pros and cons of each design Student Formula CMU F-813” (F-813). The F-813 joined methodology that the design following the theoretical the competition in the previous season 2013-2014, and was calculation simultaneously with software simulation is an focused on the downsizing of the car, such as, a decrease outstanding design scheme because accurate results can be in all dimensions of the car, a decrease in the size of the easily obtained while the design procedure could be suspension parts, wheels, and tires, and a design for a new fi nished in a short time. For this reason, the theoretical and transmission system along with a gear ratio corresponding simulating approaches were chosen for the design scheme to the smaller wheels and tires. However, the engine for the intake manifold in this study. Moreover, additional installed in the F-914 was the same as the one installed in experiments were conducted to verify the reliability of the F-813, due to its power and torque suiting the size and the design result, especially the one obtained from the weight of the car and the characteristic of the competition. simulation. Although the engine was identical, due to a decrease in the size and weight of a number of parts, especially in the suspension and transmission systems, the intake manifold, (1) Air filter which was optimally designed and developed for the engine installed on the F-813, could not be used with the engine (2) Throttle valve installed in the F-914 without modifi cation. Not doing this might cause a decrease in the engine’s performance and a (4) Surge tank disagreement between the engine’s output and the changed (3) Restrictor car’s components. In light of this reason, the design, development, and construction of the intake manifold used (5) Bellmouths with the engine on the F-914 were undertaken. An intake manifold is a group of tubes or parts that is used to deliver atmospheric air to the cylinders. It generally consists of six main parts: (1) air fi lter, (2) throttle valve, (6) Runners (3) restrictor, (4) surge tank, (5) bellmouths, and (6) runners, as shown in Figure 2. Since the installed engine was a Figure 2. General components of intake manifold. normally aspirating engine (NA-engine) — an engine without air charging devices, the performance of the engine Determining the needs of the suitable intake manifold depends on the engine’s volumetric efficiency. The for the F-914 became a signifi cant part of this study, of effi ciency subsequently depends on the mass of the air fed which the objectives were to design, develop, and construct into the engine. In general, when the engine operates at a the intake manifold for the F-914 using theoretical analysis certain speed and load, the pistons will suck the air with a together with simulation and experimental verifi cation on certain vacuum pressure. Therefore, if the intake manifold the design results. Then to compare the pressure drop in can be designed to have a suitable air passage, such as the intake manifold of the F-914 to the predecessor F-813. highly smooth wall surface, very low fl ow resistance, The usefulness of this study is that a reliable, convenient, straighter tube, and minimum number of a sudden changes and rapid design scheme for the intake manifold was in cross-sectional area of the air passage, etc., the pressure obtained. This scheme could be developed further to be drop in the air fl owing into the intake manifold will be used in the design of the intake manifolds installed in minimal. This leads to a higher mass of the air fl owing into industrial-scale produced cars. the engine that increases the engine’s performance. The methodology to design the intake manifold to have the 2. Design methodology lowest pressure drop as possible can be classifi ed in the The target of the design of the intake manifold in this study methods as follows: (i) theoretical analysis (Harrison, 2004; was that the pressure drop in the air fl ow in the intake Wang, 2011); the design result is the most accurate, but the manifold of the F-914 was lower than that of the F-813 existing theories do not cover some criterions on some parts within the engine speed range of 7,000-9,000 rpm, which of the intake manifold, especially on the parts with more was the speed range most frequently used in the competition. complex shapes. (ii) Empirical correlation; this method is The scopes of the design were as follows: (i) the intake easy and convenient although the design result is acceptably manifold was designed for use with a 4-cylinder, 4-stroke precise only when the design criteria are identical with the gasoline engine (Suzuki, GSX-600R, model year 2007, condition in the correlation establishment. (iii) Numerical 599-cm3 displacement volume), which was installed on the method (Jemni, 2011; Sakowitz, 2014); the design result F-914. (ii) The intake manifold was restricted by the 2014 is reliable while the method is complicated, and the designers SAE Student Formula rules (2014 SAE Student Formula must be experts in mathematical problem solving. Then Rules, 2013). (iii) The diameter of the restrictor located (iv) computer-aided simulation (Solid Works Flow Simulation, behind the throttle valve did not exceed 20 mm. Then (iv) 2015); this method is now widely used according to its no part of the intake manifold could be exposed throughout Development of intake manifold for Student Formula CMU F-914 Vol 1. No 2, July-December 2015 using theoretical and simulated designs 41 an outline of the frame and to the driver’s cockpit. Details from 30 to 80 mm, the optimum location decreased from of the design can be described as follows. 50% to 20%, and in turn, moved towards the inlet of the restrictor. According to the results, a restrictor with an 2.1 Design of restrictor outlet diameter of 60 mm was chosen to be constructed An air restrictor is a device used to limit the width of an since this size matched with the diameter of the bellmouths air channel, which does not exceed 20 mm. It is always (described in next topic) and the best location of the nar- installed between the throttle valve and the air surge tank rowest passage was at 25% of the total length. on every race car. This is according to the competition’s regulations (2014 SAE Student Formula Rules, 2013). An 2.2 Design of bellmouths objective of this rule is to reduce the engine’s power to a A bellmouth, or a bell-shape nozzle, is a device used to level that suits amateur drivers. The inlet diameter of the reduce a cross-sectional area of a fl uid fl ow’s channel restrictor was initially defi ned as being equal to 38 mm, between a bigger surge tank and a smaller runner. The which was the same as the diameter of the throttle valve installation of bellmouths diminishes the turbulent air fl ow installed on the inlet of the restrictor. The length of the at the inlet of the runners and consequently reduces the restrictor was initially defi ned as 125 mm since this length friction from the air fl ow. Since the outlet of the bellmouth did not lead the intake manifold to be exposed out of the has to be directly connected to a runner with the same frame, as restricted by the rules. The theory of fl uid fl ow diameter, the size of the runner should be obtained prior to could be used for the design of the restrictor. The principle designing the bellmouth. Generally, the cross-sectional area of gradual contraction and expansion loss showed that the of the suitable runner should be the same as that of the range of the suitable taper angle should be within 10o-40o engine’s intake port to maintain the intake air fl ow’s velocity (Fox, 2010). However, the principle did not inform the as a constant. The intake port shape of the engine was most suitable location of the narrowest cross-sectional area almost oval with a cross-sectional area of 940.78 mm2. The of the fl ow passage in the restrictor related to the total equivalent internal diameter of each runner could be found length of the restrictor. As when the location of the from: narrowest passage is changed, the ratio between the inlet and outlet taper angles of the restrictor and the pressure drop were consequently changed. Moreover, the outlet diameter of the restrictor was another parameter that affected the pressure drop across the restrictor. As some conditions of the restrictor could not be designed using the An aluminum tube was chosen to be the raw material of theory of fl uid fl ow, a simulation of the air fl ow in various the runner because of its smooth surface and low density shapes of the restrictor must be conducted using the fl ow (Incropera, 2002), and as 34.6-mm internal-diameter simulation module in the SolidWorks software (Sakowitz, aluminum tubing is not in general markets, the closest 2014). The results from the simulation can be exemplifi ed internal diameter of the tube was alternatively selected as as shown in Figure 3. It can be noted from Figure 3 that 36.0 mm. In light of this reason, the outlet diameter of the the location of the narrowest passage was defi ned as the bellmouth was selected to be 36 mm. length percentage measured from the inlet related to the Previous experimental study found that a bellmouth length of the restrictor. with an elliptical internal surface profi le, as expressed in Equation (2), allowed the intake air to fl ow through with a higher mass fl ow rate than that of the airfoil profi le and simple radius (Blair, 2006).

%Lres Since a is the length of the runner, which was the best value obtained from the part study (Kammuang-lue, L res 2012), and was 23 mm. b is the difference in the radius between the inlet and outlet of the bellmouth. As a width z of the fl ange between the runners and the surge tank was initially defi ned as 70 mm, the internal diameter of the x y inlet of the bellmouths was 62 mm. In the case of the outlet, the internal diameter was restricted by the size of Figure 3. Simulation of air fl ow in restrictor. the runners, which was 36 mm. Therefore, b could be found from (62 – 36) / 2 = 13 mm. When a and b were It can be seen from the simulation that when the outlet substituted into Equation (2), the elliptical equation diameter of the restrictor increased from 30 to 80 mm, the describing the profi le of the bellmouth in this study equaled pressure drop across the restrictor continuously decreased. to obtain as Equation (3). The shape of the designed In addition, it was found that the optimum location of the bellmouth is shown in Figure 4. narrowest passage of the restrictor strongly depended on the outlet diameter. When the outlet diameter increased 42 Niti Kammuang-lue and Worawit Borikhut R & K

of sound in air, which is 350 m/s. At and Lt are the cross-sectional area and the length of the tuned pipe — the portion of the intake manifold between the throttle valve

and the inlet of the surge tank, respectively. VR is the resonating volume or the internal volume of the whole intake manifold. Since the cross-sectional area of the

restrictor was not uniform, At in the previous equation was substituted by an average cross-section area, which could be found from:

x When Equation (4) was substituted with Equation (5) and with all the above deﬁ ned constants, the resonating volume y was achieved as:

Figure 4. Designed bellmouth.

2.3 Design of surge tank An air surge tank is an empty chamber used to reserve suffi cient intake air for combustion. The suitable surge tank must have enough capacity to continuously distribute intake air to each cylinder. The volume of a surge tank can be determined using the Helmholtz resonator model The resonating volume was the internal volume of the air (Heisler, 1995), expressed as: fl ow channel from the throttle valve to the intake valve’s face, which consisted of the volume of the surge tank, all the bellmouths, and the runners. The volume of the surge tank can be found using Equation (7). The length of each runner was initially defi ned to be 102 mm, which was the longest length for the intake manifold that could fi t in the Where f is the resonant frequency of the effective engine R race car. speed, which was defi ned as 7,000 / 60 Hz. C is the velocity

2.4 Simulation of intake manifold 1. It can be seen that the intake manifold model No. 1 It can be seen from the above design that the exact shape caused the lowest pressure drop. Therefore, this optimized and dimensions of the restrictor, bellmouths, and runners shape of the surge tank and intake manifold was chosen to were completely obtained, except for the surge tank of be constructed and installed on the F-914 for further which only the suitable volume was determined, but the experiments. It should be noted that although an error in suitable shape was still in a black box. From this point, the pressure drop obtained from the simulation could be eight possible shapes of the surge tank were created in this observed in each of the modeled intake manifolds, the study with the same internal volume of about 1,558 cm3. magnitudes of the errors were nearly the same and were Then, each shape was modeled in the simulation software less than 2%. Therefore, the errors could be reasonably and connected to the restrictor, bellmouths, and runners to neglected from consideration when selecting the optimized be the modeled intake manifold. Consequently, the air fl ow shape of the surge tank and intake manifold. in each of the modeled intake manifolds was simulated to fi nd the pressure drop and streamline. The pressure at the 3. Experimental setup and procedure outlet of the runners was initially set to 50 kPa, which was The optimized intake manifold for the F-914 was con- an average pressure due to the suction of the engine at an structed to have the same shape and dimensions as in the engine speed about 7,000 rpm. The atmospheric pressure design and simulation results. It consisted of the restrictor, of the surrounding air before entering the restrictor was which was extruded by a 3D printer with the raw material defi ned as 101.325 kPa. The pressure drop along each of acrylonitrile butadiene styrene (ABS), which is a modeled intake manifold was recorded as shown in Table thermoplastic. The surge tank was shaped from carbon Development of intake manifold for Student Formula CMU F-914 Vol 1. No 2, July-December 2015 using theoretical and simulated designs 43

fi ber reinforced with aluminum plate for strength. The to the fl ange made of an aluminum plate machined to have bellmouths were machined from an aluminum block by a four holes with the same shape as the intake port of the CNC machine. The profi le of the internal diameter of the engine. All components were attached together to form the bellmouth was the same as in the elliptical equation, from complete intake manifold. High-temperature sealant was above. The runners were made of an aluminum tube. One applied to all the fi ttings to prevent any leakage. Finally, end was welded to the outlet of the bellmouth and the the intake manifold was installed onto the engine as shown bottom plate of the surge tank and another end was welded in Figure 5.

Table 1. Modeled intake manifold and pressure drop.

Modeled intake Pressure Modeled intake Pressure Modeled intake Pressure manifold drop (Pa) manifold drop (Pa) manifold drop (Pa) No. 1 No. 4 No. 7

48,546 48,650 48,760

No. 2 No. 5 No. 8

48,622 48,701 48,569

No. 3 No. 6 F-813

48,630 48,760 50,534

the inlet of the restrictor and the outlet of each runner to measure the local pressure. The investigation was divided into two parts. (i) Verifi cation of the pressure drop obtained from the simulation, in which the experimental procedure was started after the engine was hot started and accelerated until the local pressure at the runner was 50 kPa. Then, the difference in head of the water inside the manometer was measured and used to determine the pressure drop with Equation (8). The intake manifold of the F-813 was substituted and tested with an identical procedure to fi nd the pressure drop. Finally, the pressure drop of each intake manifold obtained from the experiment was compared to the one obtained from the simulation to verify the reliability of the simulation. (ii) Comparison of the pressure drop in the intake manifolds between the F-914 and the F-813, in which the experimental procedure was the same as in the fi rst part except that the Figure 5. Complete intake manifold for the F-914. engine was accelerated until the speed was equal to 6,000, 7,000, 8,000, 9,000, and 10,000 rpm without any restriction The experimental investigation of the pressure drop in the local pressure at the runner. The pressure drop at a across the intake manifold was conducted by installing a certain engine speed was then measured and calculated. manometer into the inlet of the restrictor and the outlet of The pressure drops obtained from the F-914 and the F-813 each runner. Moreover, vacuum gauges were installed into intake manifolds were fi nally compared. 44 Niti Kammuang-lue and Worawit Borikhut R & K

if the pressure drop across the throttle valve was taken into account and the simulation was conducted again, the 4. Results and Discussion pressure drop from the simulation would be closer to the 4.1 Veriﬁ cation of pressure drop obtained from simulation one from the experiment for both intake manifolds. The It could be seen from the pressure drop obtained from the percentage difference between the pressure drops obtained experiment and simulation, as shown in Table 2, that the from the experiment and simulation could be reduced to pressure drop from the simulation differed from the one 7.17% and 13.73% for the intake manifolds of the F-914 from the experiment for both intake manifolds. This was and F-813, respectively. From this it can be concluded that because the pressure drop across the throttle valve was not the design results obtained from the simulation had acceptable considered in the simulation. On the contrary to the actual reliability and the method could possibly be used as the experiment, the throttle valve must be installed onto the scheme for designing the intake manifold. intake manifold for controlling the engine speed. Therefore,

Table 2. Pressure drop obtained from experiment and simulation for both intake manifolds.

Intake manifold Pressure drop F-914 F-813 (i) Experiment (Pa) 2,241 2,435 (ii) Simulation, without throttle valve (Pa) 48,546 49,571 (ii) – (i) Difference (%) 2,066.51 1,936.18 (iii) Simulation,with throttle valve (Pa) 2,080 2,100 (iii) – (i) Difference (%) 7.17 13.73

4.2 Comparison of pressure drop in intake manifold an ABS restrictor with the narrowest passage located at between F-914 and F-813 25% of the restrictor’s length measured from the inlet, a It could be seen from the comparison of the pressure drop carbon fi ber surge tank with an internal volume of 1,558 in both intake manifolds that the intake manifold of the cm3, aluminum elliptical profi le bellmouths, and aluminum F-914 had a lower pressure drop than the intake manifold runners with the length of 102 mm. The experimental of the F-813 of 43%, 94%, 66%, 69%, and 29% at the verifi cation showed that the pressure drop from the simulation engine speeds of 6,000, 7,000, 8,000, 9,000, and 10,000 obviously differed from the one from the experiment for rpm, respectively, as shown in Figure 6. Moreover, it could both intake manifolds since the pressure drop across the be seen that when the engine speed was 7,000 rpm, the throttle valve was not included in the simulation; however, intake manifold of the F-914 had the lowest pressure drop after the pressure drop across the throttle valve was of 87.5 Pa, which was due to this being the speed used in considered in the simulation, the pressure drop from the the design following the Helmholtz resonator model. It can simulation was almost the same as the one from the be concluded that the design and development of the intake experiment. Moreover, it was found that the intake manifold manifold for the F-914 could signifi cantly increase the of the F-914 had a lower pressure drop than that of the performance of the air delivered to the engine installed on F-813 at every engine speed. The practical outcome from the Student Formula CMU F-914. Although, there was no the design and development of the intake manifold was a evaluation of the power and torque obtained from the lower elapsed time in the acceleration event of 0.69 s, or engine with each intake manifold, due to limited instruments, 13.69%. the competition results in the acceleration event could be F-914 F-813 directly deduced for the engine’s performance. It was found 4,000 that the Student Formula CMU F-914 took 4.35 s to cover 3,500 the 75-m drag strip, while the Student Formula CMU F-813 3,000 took a longer time of 5.04 s. This can practically verify that the design and development of the intake manifold in this 2,500 study increased the overall engine’s performance and 2,000 subsequently decreased the time in the acceleration event 1,500 by 0.69 s, or 13.69%. 1,000 Pressure drop (Pa) Pressure 500 5. Conclusions 0 The intake manifold for the Student Formula CMU F-914 6,000 7,000 8,000 9,000 10,000 has been thoroughly designed, developed, and constructed Engine speed (rpm) in this study using theoretical analysis together with simulation software and experimental verifi cation of the Figure 6. Pressure drop in both intake manifolds. design results. The designed intake manifold consisted of Development of intake manifold for Student Formula CMU F-914 Vol 1. No 2, July-December 2015 using theoretical and simulated designs 45

6. Nomenclature Wang, J. 2011. Theory of ﬂ ow distribution in manifolds. 2 Ain,p Cross-sectional area of an intake port, mm Chemical Engineering Journal 168 (3), 1331-1345. 2 Ar Cross-sectional area of a runner, mm Society of Automotive Engineers Thailand. 2015. 2014 2 At Cross-sectional area of a tuned pipe, m SAE Student Formula Rules, Society of Automotive

Dr Inside diameter of a runner, mm Engineers, USA. http://www.tsae.or.th. C Velocity of sound, m/s Solid Works Flow Simulation. 2015. Dassault Systèmes fR Resonant frequency, Hz SolidWorks Corperation, USA.http://www.solidworks. Δh Head difference, m com/sw/products/simulation/ﬂ ow-simulation.htm.

Lres Length of a restrictor, m

Lt Length of a tuned pipe, m ΔP Pressure drop, Pa 3 Vb Bellmouth volume, m 3 VR Resonating volume, m 3 Vr Runner volume, m 3 Vs Surge tank volume, m 3 Vt Tuned pipe volume, m x Length of a bellmouth, mm y Different radius between inlet and outlet of a bellmouth, mm ρ Density of water in manometer, kg/m3

Acknowledgements This study was supported by the Department of Mechanical Engineering, Faculty of Engineering, Chiang Mai University, MD Racing, and all of the CMU Auto Club’s sponsors and members. The authors would like to express their sincere appreciation for all of the support provided.

References Blair, G. P. and Cahoon, W. M. 2006. Special investigation: Design of an intake bellmouth. Race Engine Technology 17, 34-41. Fox, R. W., Pritchard, P. J. and McDonald, A. T. 2010. Introduction to fl uid mechanics. 7th ed., John Wiley & Sons, USA, pp. 745. Harrison, M. F., De Soto, I. and Rubio Unzueta, P. L. A. 2004. Linear acoustic model for multi-cylinder IC engine intake manifolds including the effects of the intake throttle. Journal of Sound and Vibration 278 (4-5), 975-1011. Heisler, H. 1995. Advanced engine technology. Edward Arnold Publisher, UK, pp. 800. Incropera, F. P. and DeWitt, D. P. 2002. Introduction to heat transfer. 4th ed., John Wiley & Sons, USA. Jemni, M. A., Kantchev, G. and Abid, M. S. 2011. Infl uence of intake manifold design on in-cylinder fl ow and engine performances in a bus diesel engine converted to LPG gas fueled, using CFD analyses and experimental investigations 36 (5), 2701-2715. Kammuang-lue, N. and Chattanjai, T. 2012. Design of an intake manifold for the Student Formula CMU F-611. Proceedings of the Fourth International Conference on Science, Technology and Innovation for Sustainable Well-Being (STISWB IV), Thailand, pp. 112-120. Sakowitz, A., Mihaescu, M., and Fuchs, L. 2014. Flow decomposition methods applied to the fl ow in an IC engine manifold. Applied Thermal Engineering 65 (1-2), 57-65. Vol. 1 No. 2 page 46-51 2015 DOI:10.14456/randk.2015.11 RESEARCH & KNOWLEDGE Research Article Predictive suspension control system by inclinometer and accelerometer

Jarut Kunanoppadol* SARD, Department of Mechanical Engineering, Faculty of Engineering and Industrial Technology, Silpakorn University, Thailand

(Received 19 November 2015; accepted 25 December 2015)

Abstract - This paper presents a conceptual idea of a predictive suspension control system using two simple sensors, which are a bi-axial inclinometer and a bi-axial accelerometer, for measuring four input signals. The inclinometer is for measuring road parameters that consist of a slope angle and an incline angle, while the accelerometer is for measuring dynamic parameters that consist of an accelerating G-force and a turning G-force. A suspension control unit (SCU) is for calculating the four output signals using the concept of vehicle weight distribution. The percentage of the vehicle weight distributed to each wheel are the output signals for controlling the adjustable suspension parts, such as spring and shock absorber on each wheel independently. This article relates to the ﬁ rst phase of developing a conceptual idea. However, the prototyping and experimental phases will be conducted in the next phase.

Keywords: Predictive suspension control, inclinometer, accelerometer

1. Introduction (Poussot-Vassal et al., 2012). The sequence of the feedback The vehicle weight distributed to each wheel is infl uenced control system is shown in Table 1. The vehicle weight by two factors: a static factor and a dynamic factor. In the distributed to each wheel (front/rear and right/left) is static mode, the vehicle weight distribution depends on the infl uenced by the static and dynamic effects. On each vehicle parameters and the road parameters. The vehicle wheel, the sensors are installed to measure the force or the parameters consist of the vehicle dimensions and the defection independently, and their signals are used as four position of the center of gravity (CG); while, the road input signals. The control unit uses these input signals for parameters consists of the slope angle and the incline angle the calculation, and then sends the feedback signals to (Gillespie, 2014; Janzar, 2009; Popp et al., 2010; Rajamani, control the adjustable suspension parts on each wheel 2005). The position of the CG is identified by three separately. components in the x-, y-, and z-axis. In a dynamic mode, This article aims to develop a new conceptual idea the vehicle weight distribution is affected by dynamic to pre-control an adjustable suspension using a predictive effects consisting of the acceleration effect and the turning signal instead of a feedback signal. As shown in Table 1, effect. The vehicle suspension is designed for support a bi-axial inclinometer and a bi-axial accelerometer were under all conditions (static and dynamic modes); however, used for detecting the static and the dynamic conditions, standard suspension parts cannot be adjusted. Recently, respectively. An inclinometer is for measuring the slope there have been many studies that have developed the and the incline angle, while an accelerometer is for adjustability of suspension parts, including patents measuring the accelerating and the turning g-force. These (Fukumura et al., 1990; Naganathan and Thirupathi, 1995; sensors can be included in a suspension control unit (SCU). Henry et al., 1995; Armstrong, 2010) and research The signals from both sensors are used as the input signals (Gonzalez Rodriguez et al., 2011; Lee et al., 2012; Milecki for the SCU for the calculation. The predictive signals from and Hauke, 2012; Donoso et al., 2013). Not only the the control unit are sent to each wheel to pre-control the suspension parts, but the control systems have been adjustable suspension parts. The advantages of this system continuously developed (Poussot-Vassal et al., 2008; are the cost of these sensors, the convenience of their Dugard et al., 2012; Poussot-Vassal et al., 2012; Koch et installation, and the possibility to combine these sensors al., 2010). Traditionally, a feedback signal has been used in the SCU to avoid an installation problem. as an input signal for the control unit to control the adjustable This article is divided into fi ve sections. The concept suspension. The feedback signal is normally measured by of the vehicle weight distribution is overviewed in the next force sensors (load cell) or defection sensors installed at section, and the vehicle parameter identifications are the suspension parts on each wheel. The problems are that presented in the third section. A case study is provided in these sensors are expensive and their installation is diffi cult. the fourth section, and then the article is concluded in the They have to be installed on moving parts, such as a shock last section. Moreover, the source code for the analysis is absorber (Koch et al., 2010), a spring, or an un-sprung part also provided in an appendix. *Author for correspondence: [email protected] Vol 1. No 2, July-December 2015 Predictive suspension control system by inclinometer and accelerometer 47

Table 1. Feedback control and predictive control.

Feedback control system Predictive control system 1. Static effect (slope angle and incline angle) and 1. Inclinometer and accelerometer installed in cabin area dynamic effects (accelerating effect and turning effect). for measuring the input signals. 2. Vehicle weight distributed to suspension part on each 2. SCU installed in cabin area for calculation. wheel (front/rear, right/left). 3. Predictive signals for pre-control of the adjustable 3. Force or defection sensor installed at suspension part suspension system. for measuring the input signals. 4. SCU installed in cabin area for calculation. 5. Feedback signals for post-control of the adjustable suspension part.

2. Vehicle weight distribution becomes zero. It means that the wheel does not touch the The vehicle weight distribution on each of the four wheels road surface. Moreover, the weight fraction can be (front/rear and right/left) is infl uenced by four factors, negative. The meaning is that there is an external force which are a front wheel factor, a right wheel factor, an acting upward at that wheel to turn over the vehicle. The accelerating factor, and a turning factor (see Table 2.). The weight distributed to the wheel also affects a friction force weight distributed to each of the four wheels is considered based on the friction coeffi cient between the tire and the in terms of a weight fraction or a percentage of the total road. This friction force infl uences the accelerating ability, weight (the value in the curly brackets ‘{}’). A critical point the braking ability, and the turning ability. happens when the weight fraction of one wheel (or more)

Table 2. Vehicle weight distribution on each wheel.

Left wheel Right wheel Front wheel

Rear wheel

2.1 Front wheel factor horizontal distance from a middle line to the CG and the The front wheel factor ( ) is the fraction of the vehicle wheelbase. It is positive if the CG is on the right side of weight distributed to both front wheels (right and left) the vehicle (driver side for Thailand). The wheel track depending on the position of the vehicle’s center of fraction is the ratio of the distance between the centerline gravity (CG) in the x- and y-axis and the slope angle ( ) of the right to the left wheels and the wheelbase. If there , as shown in Equation 1. The position of the CG along the is a different between the front track and the rear track, an length of the vehicle is defined in the CGx fraction average value is typically used. The incline angle is positive ( ), which is the ratio of the horizontal distance between in a case where the incline is upward from the right wheel the CG from the rear axis and the wheelbase (see to the left wheel. It can be measured by an inclinometer. Fig.1.).The position of the CG along the height of the vehicle is defi ned in the CGy fraction ( ) , which is (2) the ratio of the vertical distance of the CG from the road and the wheelbase. The slope angle is specifi ed as positive for an upward slope, while it is specifi ed as negative for a 2.3 Accelerating factor downward slope. It is measured by an inclinometer. The acceleration factor (f_a) is the effect of the accelerating G-force (G_a) on the front wheels and the rear wheels, as (1) shown in Equation 3. The G-force is measured and compared to the acceleration of gravity. The accelerating 2.2 Right wheel factor G-force is positive for an acceleration (increasing the The right wheel factor ( ) is the fraction of the vehicle speed), while it is negative for a deceleration (decreasing weight distributed to both right wheels (front and rear) the speed). It can be measured by an accelerometer. depending on the position of the CG, the wheel track frac- Theoretically, the effect of the accelerating G-force is to tion ( ), and the incline angle ( ) , as shown in Equation increase the weight distributed at the rear wheels, while it 2. The position of the CG along the width of the vehicle is decreases the weight distributed at the front wheels defi ned in the CGz fraction ( ), which is the ratio of the (Gillespie, 2014; Popp et al., 2010; Garrett et al., 2001). 48 Jarut Kunanoppadol R & K

Figure 1. Vehicle distance fraction.

(3) (6)

2.4 Turning factor Secondly, the scales for measuring the weight at the The turning factor ( ) is the effect of the turning G-force rear wheels (right and left) are lifted up equally, and then ( ) on the right wheels and the left wheels, as shown in the weight distributed to the front wheels is re-measured Equation 4. For this paper, the turning G-force is deﬁ ned ( ). The CGy fraction ( ) can be determined from the as being positive for left-turning, while it is deﬁ ned as wheel radius ( ), the wheelbase ( ), and the lift ( ), as being negative for right-turning. Theoretically, when shown in Equations 7 and 8. turning left, the effect of the turning G-force increases the weight distributed to the right wheels, while it decreases (7) the weight distributed to the left wheels.

(8) (4) These are the processes to identifying all GC position 3. Vehicle parameter identifi cation fractions. The only parameter that affects the CG position The application of this concept must be initiated by is the wheel radius. In the case that the wheels are changed identifying the vehicle parameters that are all the CG (tire, rim, or both), the measurement must be redone. fractions in x-, y-, and z-axis and the wheel track fraction. Another vehicle parameter needed for the analysis is the There are two steps to determine the CG fractions that must wheel track fraction, which can be simply calculated from be done in a laboratory or a workshop. Firstly, the vehicle the wheel track and the wheelbase. These values can be weight at each wheel is measured on fl at ground. The CGx from the vehicle specifi cation or measurement. fraction ( ) is calculated from the weight distributed to the front wheels (right and left) ( ) and the total weight 4. Case study (the summary of four wheels) ( ), as shown in Equation This article provides a case study that illustrates the 5. The CGz fraction ( ) is calculated from the weight application of the concept. The information of the case is distributed to the right wheels (front and rear) ( ), the shown in Table 3. In this case, the slope angle and the total weight ( ), and the wheel track fraction (ratio of the incline angle were specified as zero (non-slope and wheel track and the wheelbase) ( ), as shown in Equation 6. non-incline); however, in real-time use, they are measured by a bi-axial inclinometer (one axial for the slope angle (5) and another axial for the incline angle). Table 3. Case study.

Vehicle parameters

CGx fraction 0.40 CGz fraction 0.05

CGy fraction 0.20 Wheel track fraction 0.60 Road parameter Slope angle 0 degree Incline angle 0 degree Vol 1. No 2, July-December 2015 Predictive suspension control system by inclinometer and accelerometer 49

The calculation processes can be simply done via respectively. The summary of the weight distributed to the spreadsheet software, such as Microsoft Excel, for front wheels is found to be 36.00% of the total weight. one-position analysis, as shown in Figure 2. For multiple- There would be 22.40% and 41.60% of the vehicle weight position analysis to cover a range of accelerating G-forces distributed to the rear-left wheel and the rear-right wheel, and turning G-forces, it is more appropriate to calculate respectively. The summary of the weight distributed to the via mathematical software, such as SCILAB. The results rear wheels is found to be 64.00%. The summary of the are shown in Figure 3. The source code for SCILAB is also weight distributed to the left wheels and the right wheels provided in the appendix. would be 35.00% and 65.00%, respectively. The vehicle Based on the vehicle parameters and the road weight distribution for other values of the accelerating parameters in Table 3, for the case where the accelerating G-force and the turning G-force can be determine from the G-force is 0.20 and the turning G-force is 0.20, the weight graph in Figure 3. In real-time use, these G-forces are distributed to each wheel can be determined. There would measured by a bi-axial accelerometer (one axial for the be 12.60% and 23.40% of the vehicle weight distributed accelerating G-force and another axial for the turning to the front-left wheel and the front-right wheel, G-force).

Figure 2. Analysis via Microsoft Excel.

Figure 3: Vehicle weight distribution (via SCILAB) 50 Jarut Kunanoppadol R & K

The vehicle weight distributed to each wheel can be used Both sensors can be installed separately or included with as the input signals to the SCU for the calculation, then the the SCU. Four input signal are sent to the SCU for the SCU will send the predictive signal to control the adjustable calculation, then the four output signals are used to control suspension part. Figure 4 presents a diagram of the predictive the adjustable suspension parts at each wheel. The advantages control system. The bi-axial inclinometer is for measuring of this system are using a predictive signal to pre-adjust the slope and the incline angles, and the bi-axial accelerometer the suspension, using simple and inexpensive sensors, and is for measuring the accelerating and a turning G-force. easy installation.

Figure 4. Predictive suspension control diagram.

5. Conclusion recovery system. US20100006362 A1, 14-Jan-2010. The conceptual idea of a predictive suspension control Donoso, A., Chacón, J. M., González Rodríguez, A. and system was developed for controlling adjustable suspension Ureña, F. 2013. On an adjustable-stiffness spring parts. This system uses two simple sensors, which are a composed of two antagonistic pairs of nonlinear leaf bi-axial inclinometer and a bi-axial accelerometer, for springs working in post-buckling. Mech. Mach. measuring the road parameters and the dynamic parameters, Theory 63, 1–7. respectively. The road parameter consists of the slop angle Dugard, L., Sename, O., Aubouet, S. and Talon, B. 2012. and the incline angle, while the dynamic parameters consist Full vertical car observer design methodology for of the accelerating G-force and the turning G-force. These suspension control applications. Control Eng. Pract. sensors can be included with the SCU to avoid the installation 20 (9), 832–845. problem, installed separately in the cabin area, or by using Fukumura, T., Shinbori, T., and Ezure, N. 1990. Car a sensor in a mobile phone. By using the vehicle weight suspension system. US4921224 A, 01-May-1990. distribution concept, the weight distributed to each wheel Garrett, T. K., Steeds, W. and Newton, N. 2001. Motor can be determined and used as the output signals for vehicle, 13th ed. Warrendale, PA: Society of Automotive pre-controlling the adjustable suspension parts, such as Engineers Inc. spring and shock absorber. Gillespie, T. D. 2014. Fundamentals of vehicle dynamics. Warrendale, PA: Society of Automotive Engineers Acknowledgements Inc. I would like to thank the Department of Mechanical Engi- González Rodríguez, A., Chacón, J. M., Donoso, A. and neering, Faculty of Engineering and Industrial Technology, González Rodríguez, A. G. 2011. Design of an ad- Silpakorn University that has supported all the research in justable-stiffness spring: Mathematical modeling and my laboratory (Silpakorn Automotive Research & Devel- simulation, fabrication and experimental validation. opment: SARD) Mech. Mach. Theory 46 (12), 1970–1979. Henry, R. R., Applebee, M. A. and Murty, B. V. 1995. Full References car semi-active suspension control based on quarter Armstrong L. D. 2010. Vehicle suspension kinetic energy car control. US5475596 A, 12-Dec-1995. Vol 1. No 2, July-December 2015 Predictive suspension control system by inclinometer and accelerometer 51

Jazar, R. N. 2009. Vehicle dynamics: Theory and application, pi/180)//right wheel factor 3rd ed. Springer, New York. accfactor=gacc(i)*xcgy//accelerating factor Koch, G., Fritsch, G. and Lohmann, B. 2010. Potential of turnfactor=gturn(j)*xcgy/xt//turning factor low bandwidth active suspension control with frontleft(j,i)=(frontfactor-accfactor)*(1-rightfactor-turnfac- continuously variable damper. Control Eng. Pract. tor)*100//front-left wheel(percentage) 18 (11), 1251–1262. frontright(j,i)=(frontfactor-accfactor)*(rightfactor+turnfa Lee, J. H., Yi, B.-J. and Lee, J. Y. 2012. Adjustable spring ctor)*100//front-right wheel(percentage) mechanisms inspired by human musculoskeletal rearleft(j,i)=(1-frontfactor+accfactor)*(1-rightfactor-turn- structure. Mech. Mach. Theory 54, 76–98. factor)*100//rear-left wheel(percentage) Milecki, A. and Hauke, A. 2012. Application of magnetor- rearright(j,i)=(1-frontfactor+accfactor)*(rightfactor+turnf heological fl uid in industrial shock absorbers. Mech. actor)*100//rear-right wheel(percentage) Syst. Signal Process. 28, 528–541. end Naganathan, G. and Thirupathi, S. R. 1995. Active suspension end systems and components using piezoelectric sensing //Ploting the results and actuation devices. US5390949 A, 21-Feb-1995. scf(0) Popp, K., Schiehlen, W., Kröger, M. and Panning, L. 2010. subplot(221)//front-left wheel Ground vehicle dynamics. Springer, Berlin. contour(gturn,gacc,frontleft,5) Poussot-Vassal, C., Sename, O., Dugard, L., Gáspár, P., xtitle(‘front-left wheel’,’ ‘,’accelerating g-force’) Szabó, Z. and Bokor, J. 2008. A new semi-active subplot(222)//front-right wheel suspension control strategy through LPV technique. contour(gturn,gacc,frontright,5) Control Eng. Pract. 16 (12), 1519–1534. xtitle(‘front-right wheel’,’ ‘,’ ‘) Poussot-Vassal, C., Spelta, C., Sename, O., Savaresi, S. M. subplot(223)//rear-left wheel and Dugard, L. 2012. Survey and performance contour(gturn,gacc,rearleft,5) evaluation on some automotive semi-active suspension xtitle(‘rear-left wheel’,’turning g-force’,’accelerating g- control methods: A comparative study on a single- force’) corner mode. Annu. Rev. Control 36 (1), 148–160. subplot(224)//rear-right wheel Rajamani, R. 2005. Vehicle dynamics and control. Springer, contour(gturn,gacc,rearright,5) New York. xtitle(‘rear-right wheel’,’turning g-force’,’ ‘) scf(1)//front-left wheel contour(gturn,gacc,frontleft,5) Appendix: SCILAB source code scf(2)//front-right wheel clc; clear contour(gturn,gacc,frontright,5) // vehicle weight distribution project by JK scf(3)//rear-left wheel //vehicle parameters contour(gturn,gacc,rearleft,5) xcgx=0.40//cgx fraction scf(4)//rear-right wheel xcgy=0.20//cgy fraction contour(gturn,gacc,rearright,5) xcgz=0.05//cgz fraction xt=0.60//wheel track fration //road parameters slope=0//slope angle(degree) incline=0//incline angle(degree) //calculation gmin=0//minimum g-force(g) gmax=0.5//maximum g-force(g) interval=0.1//calculating interval(g) msize=(gmax-gmin)/interval*2+1//matrix size frontright=zeros(msize,msize)//front-right wheel matrix frontleft=zeros(msize,msize)//front-left wheelmmatrix rearright=zeros(msize,msize)//rear-right wheel matrix rearleft=zeros(msize,msize)//rear-left wheel matrix gacc=zeros(msize) gturn=zeros(msize) for i=1:msize gacc(i)=-gmax+(i-1)*interval//accelerating g-force for j=1:msize gturn(j)=-gmax+(j-1)*interval//turning g-force frontfactor=xcgx*cos(slope*%pi/180)-xcgy*sin(slope*% pi/180)//front wheel factor rightfactor=(1/2+xcgz/ xt)*cos(incline*%pi/180)+xcgyxt*sin(incline*% Vol. 1 No. 2 page 52-56 2015 DOI:10.14456/randk.2015.10 RESEARCH & KNOWLEDGE Research Article Power plant effi ciency improvement by optimizing main infl uential control parameters

Polachate Tosomboon and Thosapon Katejanekarn* Building Energy Systems Laboratory, Department of Mechanical Engineering, Faculty of Engineering and Industrial Technology, Silpakorn University, Nakhon Pathom 73000, Thailand.

(Received 19 November 2015; accepted 25 December 2015)

Abstract - This study was an attempt to increase power plant effi ciency by adjusting the process control parameters that resulted from a numerical analysis of the power plant effi ciency and its relevant operating parameters. The optimum values for the four main control parameters that would make the power plant run at the maximum effi ciency were evaluated for two regular load levels of 245 and 300 MW. The optimum values from the model were validated against the plant simulator due to the fact that it was too risky to adjust the real power plant. The results showed that if the plant was operated at the optimum combinations of the main control parameters, the effi ciency of the plant at the load levels of 245 and 300 MW would be 0.3859 and 0.3935, respectively. From the operating data during the last two years, the plant effi ciency at these two load levels would be improved by 0.0178 and 0.0149, which is equivalent to natural gas cost savings of 752,704 Baht/day (about 22,138 US$/day) and 609,862 Baht/day (about 17,937 US$/day), respectively.

Keywords: Power plant efﬁ ciency, optimization, control parameters, numerical analysis

1. Introduction and changing the angles of the chimney walls and slopes In Thailand, electrical energy consumption has increased of the collectors resulting in a 0.53% higher effi ciency by approximately 4.3% annually (Energy Policy and (Ghorbani et al., 2015). Planning Offi ce, 2014), which in turn causes the country Adjusting the process control parameters is another to build more power plants to meet the demand. More than group of methods that can improve effi ciency with much 80% of energy sources to produce electricity in Thailand less, or even no, investment, for example, increasing the are fossil fuels. Natural gas (NG) shares 67% of the energy steam inlet temperature from 508 to 568oC and keeping the mix (Electricity Generating Authority of Thailand, 2015a), pressure constant at 124.61 bar can cause a 3.79% higher of which around 60% comes from the gulf of Thailand and effi ciency (Geete and Khandwawala, 2013), tuning the around 40% has to be imported (Electricity Generating pre-dried lignite-fired power system causes a 1.51% Authority of Thailand, 2015b). higher effi ciency (Han et al., 2014), and modifying the Electrical power plants are one of the most intense combustion control systems causes a 1.69% higher fossil fuel consumers. With a slight improvement in their effi ciency (Mikulandric et al., 2013). effi ciency, a signifi cant amount of savings can be obtained. Power plant effi ciency analysis may be performed in For example, a 735-MW power plant fueled by natural gas two ways. One is performing a theoretical thermodynamic currently operates at an efficiency of 35.12%. If the analysis, which is accurate but could be quite complicated. effi ciency is improved by only 1%, the fuel cost would be The other is analyzing it numerically, which is more rapid reduced by as much as 542,450 Baht per day or 198 million and simple but less accurate. Baht (about 5.8 million US$) per year (based on the cost In this study, a thermal power plant was analyzed of natural gas of 300 Baht or 8.8 US$ per MMBtu). numerically to improve its effi ciency by adjusting the There are several methods to improve the effi ciency process control parameters. Four main infl uential control of power plants. Some methods require a high investment parameters were optimized to achieve combinations that to improve, replace, or install equipment, for example, yield the highest effi ciency at two regular operating loads upgrading the boiler and installing an air separation unit of 245 MW and 300 MW. The results were then validated resulting in a 3.3-4.0% higher effi ciency (Kotowicz and against the plant simulator, due to the risk involved with Balicki, 2014), installing a low-pressure economizer an actual test with the real plant. resulting in a 0.46% higher effi ciency (Wang et al., 2014),

*Author for correspondence: [email protected] Power plant efﬁ ciency improvement by optimizing main inﬂ uential Vol 1. No 2, July-December 2015 control parameters 53

2. Methodology adjusted by the operators in the control room, four control A 735-MW thermal power plant was studied in this work. parameters (reheat steam temperature, condenser vacuum, It is operated 24 hours a day and fueled mainly by natural fi nal feed water temperature, and excess oxygen) were gas. The secondary fuel is heavy oil. There are 12 main selected as the most infl uential ones that would be opti- control parameters, as shown in Table 1. By considering mized in this study. which ones infl uence the effi ciency the most and can be

Table 1. Main control parameters of power plant.

Item Description Unit 1 Main steam pressure bar 2 Main steam temperature °C 3 Reheat steam pressure bar 4 Reheat steam temperature °C 5 Final feed water temperature °C 6 Feed water fl ow t/h 7 Superheat spray water fl ow rate t/h 8 Reheat spray water fl ow rate t/h 9 Condenser vacuum bar 10 Stack temperature °C 11 Excess oxygen % 12 Circulating water temperature °C

A multiple least-squares regression model was results from the plant simulator, due to the fact that it was formed to evaluate the power plant effi ciency in terms of too risky to perform a test with the real plant. If the the four main infl uential control parameters, as shown in difference between the results from the regression model Eq. (1). and the plant simulator did not exceed the propagated error 2 3 4 η = a0 + a1A + a2A + a3A + a4A + … of the plant effi ciency, the optimum values of the four 2 3 4 + a5B + a6B + a7B + a8B + … parameters were considered to be valid. 2 3 4 + a9C + a10C + a11C + a12C + … 2 3 4 + a13D + a14D + a15D + a16D + … (1) 3. Results and Discussion where, η = power plant effi ciency, 3.1 Reliability of plant simulator A, B, C, D, … = main infl uential control Since it was not possible to adjust the main four control parameters, parameters in the real power plant, the plant simulator was used instead. Table 2 shows the comparison between a0, a1, a2, a3, … = coeffi cients. By applying a statistical analysis to the operating the operating parameters retrieved from the real power data of the power plant from the last two years, only the plant and the ones obtained from the plant simulator at the terms that have signifi cant effects were maintained in the regular load level of 245 MW. It can be seen that the real model. values and the values from the plant simulator were very To fi nd the optimum values of the four main parameters close to each other. The highest difference was 1.73%, that give the highest plant efficiency, the concept of which was smaller than the average uncertainty (2%) of univariate search was implemented. The fi rst parameter the measuring devices in the power plant. The comparisons was varied while the remaining three parameters were kept were done on many sets of operating data and the fi ndings constant until the maximum effi ciency was reached, then were always similar. Therefore, it can be concluded that the value of the fi rst parameter was fi xed at this point. the plant simulator is reliable and can represent the real After that, the next parameter was varied while the other power plant. three were fixed until the maximum efficiency was achieved. This process was repeated until the values of all 3.2 Power plant effi ciency models four parameters did not change anymore. Then, the fi nal Starting from the full form of the model according to Eq. values of the four parameters were considered to be the (1) and then applying a statistical analysis, only the terms optimum values. that have signifi cant effects were kept. The power plant The search for the optimum values of the four main effi ciency models for the regular load levels of 245 and parameters was carried out at two regular load levels of the 300 MW were obtained as shown in Eqs. (2) and (3), power plant, which were 245 and 300 MW. The results respectively. from the regression model were then compared with the 54 Polachate Tosomboon and Thosapon Katejanekarn R & K

Table 2. Comparison between main control parameters from actual data and plant simulator (sample data set at load level of 245 MW).

Simulator Difference Item Description Unit Actual data data (%) 1 Electrical power load MW 245.10 245.00 0.04 2 Main steam pressure bar 116.38 116.33 0.04 3 Main steam temperature °C 508.17 508.00 0.03 4 Reheat steam pressure bar 15.85 15.77 0.51 5 Reheat steam temperature °C 505.49 504.90 0.12 6 Final feed water temperature °C 227.41 227.00 0.18 7 Feed water fl ow rate t/h 787.32 779.00 1.06 8 Superheat spray water fl ow rate t/h 19.66 20.00 -1.73 9 Reheat spray water fl ow rate t/h 0.00 0.00 0.00 10 Condenser vacuum bar 0.4666 0.4669 0.07 11 Stack temperature °C 80.69 81.54 -1.05 12 Excess oxygen % 5.83 5.89 -1.03 13 Circulating water temperature °C 26.29 26.36 -0.27

2 η = 0.0275XR + 125.1040XC - 13.6364XC - The power plant effi ciency as calculated from the model 2 3 0.0009XF + 0.0115XO - 1.1744XO + 76.2771 (2) shown in Eq. (2) was 0.3859, while the effi ciency from the plant simulator was 0.3867. The difference between the 2 two values was 0.0008, which was less than the η = -0.0323XR + 0.3420XC + 0.2220XC + 2 2 propagated uncertainty of the power plant efficiency 0.0023XF + 1.7405XO + 2.0392XO + 38.1421 (3) (absolute uncertainty = 0.0011, relative uncertainty = o 0.33%). Therefore, this suggests that the optimum values where, XR = reheat steam temperature ( C), of the main control parameters obtained from the model XC = condenser vacuum (bar), o were valid. XF = fi nal feed water temperature ( C), The average value of the plant effi ciency at the load XO = excess oxygen (%). level of 245 MW during the last two years was approximately 0.3689. If the plant could operate at the optimum 3.3 Optimum values of main control parameters effi ciency obtained from the model, then the plant effi ciency The optimum values of the four main infl uential control could be increased by 0.0178, which is equivalent to a parameters at the regular load level of 245 MW obtained reduced heat rate of 450.20 kJ/kWh or a natural gas cost from the univariate search concept are shown in Table 3. saving of 752,704 Baht/day (about 22,138 US$/day).

Table 3. Optimum values of main control parameters at 245 MW load level.

Item Description Unit Optimum Values 1 Reheat steam temperature °C 508.22 2 Condenser vacuum bar 0.0246 3 Final feed water temperature °C 225.85 4 Excess oxygen % 4.66 5 Average effi ciency during the last 2 years - 0.3689 6 Optimum effi ciency from the model - 0.3867 7 Difference - 0.0178 8 Absolute error - 0.0012 9 Relative error % 0.36 Power plant effi ciency improvement by optimizing main infl uential Vol 1. No 2, July-December 2015 control parameters 55

The optimum values of the four main infl uential previous section, it is still worth determining how much it control parameters at the regular load level of 300 MW would help to reduce the operating cost of the plant at this obtained from the univariate search concept are shown in combination of the parameters. Table 4. The power plant effi ciency calculated from the The average value of the plant effi ciency at the load model shown in Eq. (3) was 0.3935, while the effi ciency level of 300 MW during the last two years was approxi- from the plant simulator was 0.3910. The difference mately 0.3761. If the plant could operate at the optimum between the two values was 0.0025, which was higher than efficiency obtained from the model, then the plant the propagated uncertainty of the power plant effi ciency efficiency could be increased by 0.0149, which is (absolute uncertainty = 0.0011, relative uncertainty = equivalent to a reduced heat rate of 364.76 kJ/kWh or a 0.33%). Even though more data were needed to make the natural gas cost saving of 609,862 Baht/day (about 17,937 optimum values of the main control parameters obtained US$/day). from the model valid, according to the criteria stated in the

Table 4. Optimum values of main control parameters at 300 MW load level.

Item Description Unit Optimum Values 1 Reheat steam temperature °C 529.58 2 Condenser vacuum bar 0.0432 3 Final feed water temperature °C 238.85 4 Excess oxygen % 5.54 5 Average efﬁ ciency during 2 years - 0.3761 6 Optimum efﬁ ciency - 0.3910 7 Difference - 0.0149 8 Absolute error - 0.0011 9 Relative error % 0.30

4. Conclusion References This study aimed to increase the power plant effi ciency by Geete, A. and Khandwawala, A. I. 2013. Thermodynamic adjusting the process control parameters using the results analysis of 120 MW thermal power plant with from a numerical analysis of the power plant effi ciency and combined effect of constant inlet pressure (124.61 its relevant operating parameters. The optimum values of Bar) and different inlet temperatures. Case Studies the four main control parameters that would make the in Thermal Engineering 1(1), 17–25. power plant run at the maximum effi ciency were evaluated Ghorbani, B., Ghashami, M. and Ashjaee, M. 2015. for two regular load levels of 245 and 300 MW. The Electricity production with low grade heat in thermal optimum values from the model were validated against the power plants by design improvement of a hybrid dry plant simulator due to the fact that it was too risky to adjust cooling tower and a solar chimney concept. Energy the real power plant. The results showed that if the plant Conversion and Management 94, 1–11. was operated at the optimum combination of the main Han, X. Q., Liu, M., Wang, J., Yan, J., Liu, J. and Xiao, F. control parameters, the effi ciency of the plant at the load 2014. Simulation study on lignite-fi red power system levels of 245 and 300 MW would be 0.3859 and 0.3935, integrated with fl ue gas drying and waste heat recovery respectively. From the operating data during the last two – performances under variable power loads coupled years, the plant effi ciency at these two load levels would with off-design parameters. Energy 76, 406–418. be improved by 0.0178 and 0.0149, which would be Kotowicz, J. and Balicki, A. 2014. Enhancing the overall equivalent to natural gas cost savings of 752,704 Baht/day effi ciency of a lignite-fi red oxyfuel power plant with (about 22,138 US$/day) and 609,862 Baht/day (about CFB boiler and membrane-based air separation unit. 17,937 US$/day), respectively. Energy Conversion and Management 80, 20–31. Mikulandrić, R., Loncar, D., Cvetinovi, D. and Spiridon, Acknowledgements G. 2013. Improvement of existing coal fired The authors would like to thank the power plant in thermal power plants performance by control Ratchburi, Thailand for the data used in this study. The systems modifi cations. Energy 57, 55–65. authors would also like to thank the Department of Wang, C. J., He, B., Yan, L., Pei, X. and Chen, S. 2014. Mechanical Engineering, Faculty of Engineering and Thermodynamic analysis of a low-pressure Industrial Technology, Silpakorn University for the support economizer based waste heat recovery system for a to complete this work. coal-fi red power plant. Energy 65, 80–90. 56 Polachate Tosomboon and Thosapon Katejanekarn R & K

Energy Policy and Planning Office, 2014. Electrical consumption and production of Thailand. Ministry of Energy, Thailand. http://www.eppo.go.th, access on 16/04/2015. Electricity Generating Authority of Thailand. 2015a. Fuel consumption for electrical generating. http://www. egat.co.th, access on 20/04/2015. Electricity Generating Authority of Thailand. 2015b. Natural gas price and electricity charge. http://www. egat.co.th, access on 3/05/2015. Vol. 1 No. 2 page 57-62 2015 DOI:10.14456/randk.2015.9 RESEARCH & KNOWLEDGE Research Article

Heat transfer characteristics of thermo-syphon: Case study of modifying inclination angle of evaporator and condenser sections

Kittipon Khangam and Narong Srihajong Department of Mechanical Engineering, Faculty of Engineering, Rajamangala University of Technology Isan Khon Kaen Campus, Thailand, 40000

(Received 19 November 2015; accepted 25 December 2015)

Abstract - Experiments were carried out to study the heat transfer characteristics of a thermosyphon when modifying the evaporator and condenser sections to be oriented at 0, 60 and 90 degrees with respect to the 70 degree inclination of the adiabatic section (Z shaped). The thermosyphon was made of copper tube with an outer diameter of 15.87 mm and 300, 450 and 600 mm total lengths. The evaporator, adiabatic and condenser sections were equal in length. R-134a, ethanol and distilled water were employed as the working fl uids with a 50% fi lling ratio by evaporator volume. The evaporator section was heated by hot water at 60, 70 and 80 oC, whereas 20 oC cool water was supplied to the condenser section. The test was conducted at various inclination angles, i.e., 0, 20, 40, 60, 70, 80, 90, 110, 130, 150, 170 and 180 degrees. The results show that the heat fl ux signifi cantly depends on the working temperature. R-134a had the best performance followed by ethanol and distilled water. The heat fl ux decreased with an increase in the total length. When increasing the orientation angles of the evaporator and condenser sections, the heat fl ux rose due to the assistance of gravity. The maximum heat fl ux was 26.85 kW/m2 with the lowest thermal resistance of 0.065 oC/W. In addition, the optimal tested orientation angle was in the range of 40-110 degrees with respect to the horizontal plane.

Keywords: Thermosyphon, heat transfer characteristic, modify, inclination angle

1. Introduction A thermosyphon is regarding as a highly effective A thermosyphon is a wickless heat pipe, which is also heat transfer device, which can be found in many applications, known as a gravity-assisted heat pipe. It contains a small for instance heat exchanger, electronic cooling or power amount of working fl uid for heat transfer (Shabgard et al., generation, etc. The heat transfer performance of the 2014). In addition, for basic principle (see Fig. 1), heat is thermosyphon is affected by many factors, such as, section supplied to the evaporator where the working fl uid pools length, working fl uid, fi lling ratio, inclination angle and inside, and is changed into vapor. Then, the temperature heat input. These factors have been studied by numerous and pressure cause the vapor to fl ow toward the condenser. investigations that aimed to enhance its performance (Jiang The vapor adjacent to the condenser releases its latent heat et al., 2014). and condenses into liquid, after which it returns to the Previous experiments found that a thermosyphon evaporator owing to gravity (Fadhl et al., 2013). works well when oriented at 60-90 degrees. Researchers have attempted to improve the performance by modifying the inclination angle of the evaporator and condenser sections to 0, 60 and 90 degrees with respect to the 70 degree inclination of the adiabatic section, as illustrated in Figure 2.

2. Experimental and procedure The tested thermosyphon was made of copper tube with a 14.06 mm inner diameter and 15.87 mm outer diameter. The length of the evaporator, adiabatic and condenser sections were equal, and varied at 100, 150 and 300 mm. The evaporator and condenser sections were modiﬁ ed to be oriented at 0, 60 and 90 degrees with regard to the 70 degree inclination of the adiabatic section (see Fig. 2.). Figure 1. Schematic of thermosiphon. *Author for correspondence: [email protected] 58 Kittipon Khangam and Narong Srihajong R & K

Distilled water, ethanol and R-134a were used with a ﬁ lling 60, 70, 80, 90, 110, 130, 150, 170 and 180 degrees. The ratio of 50% of the evaporator volume. Experiments were entire surface of the tested thermosyphon was wrapped performed at various inclination angles, namely 0, 20, 40, with insulating material to prevent heat loss.

Figure 2. Thermosyphon with modiﬁ ed evaporator and condenser sections oriented at 0, 60 and 90 degrees vs. conventional thermosyphon.

The test rig is shown in Figure 3 and consists of a thermosyphon, a hot bath, a cool bath, fl ow meter and data acquisition. The evaporator section was heated by hot water at 60, 70 and 80 oC from the hot bath (Thermo Fisher Scientific EX-35 with ±0.01oC), while the condenser section was cooled with circulated water at 20 oC from the cold bath (EYELA CA-112CE cold bath with ±2oC). The mass fl ow rate of the water jackets of both the evaporator and condenser were controlled by a fl oating rotameter at the rate of 0.4 l/min. There were four thermo- couples of type-K installed at the inlet-outlet for both the evaporator and condenser, as can be seen in Figure 4, and temperature probes were installed on three points on each of the outer surfaces (3x3 = 9 points). All installed thermocouple points were monitored and recorded by a data logger (Yogokawa MV1000 with ±0.7oC accuracy). Figure 4. Installed thermocouple point.

The experiment begins by supplying the hot water and cool water at the desired temperatures to the water jackets. The ﬂ ow rate was controlled to be 0.4 l/min. The temperature was recorded with the data logger for 10 minutes or until the temperature was stable. Calculate the heat transfer rate at the condenser using the following (On-ai et al., 2013): (1)

Where, is the heat transfer rate (W), is

the mass ﬂ ow rate of the cooling water (kg/s), is the speciﬁ c heat of the cooling water (J/kg. oC) and is the temperature difference of the inlet and outlet of the condenser (oC). Figure 3. Schematic diagram of test rig. Heat transfer characteristics of thermo-syphon: Case study of modifying Vol 1. No 2, July-December 2015 inclination angle of evaporator and condenser sections 59

The heat transfer rate from the evaporator section is examined using the same approach, and is given by: (5)

(2) Where, is the temperature difference between the average wall temperature of the evaporator The rate of heat transfer per unit area or heat ﬂ ux and condenser sections. . is the heat transfer (W) in the (W/m2) is express as (Jiao et al., 2008): evaporator section that can be calculated by similar methods as . (3) Above all, the thermal performance can be deﬁ ned as follows (Cengel, 2002):

(4) (6)

3. Results and Discussion Where, and are the heat transfer areas of 3.1 Effect of working temperature 0.00518, 0.00767 and 0.0106 m2 at the section lengths of The experimental results showing the effect of the working 100, 150 and 200 mm ( and ), respectively. temperature on the heat fl ux are in Figure 5. It can be seen that when the working temperature at the evaporator section The thermal resistance could be evaluated using the increases, the heat fl ux increases as well, due to the direct following equation (Park et al., 2002): effect of the heat causing the fl uid to boil. At 80 oC for the working temperature, the highest heat fl ux at each tested orientation was achieved.

Figure 5. Effect of working temperature on heat ﬂ ux at various tested angles (R-134a). 3.2 Effect of working ﬂ uid

Figure 6. Effect of working ﬂ uid on heat ﬂ ux at various tested angles (working temp. = 80°C). 60 Kittipon Khangam and Narong Srihajong R & K

Figure 6 presents the heat fl ux with different working length of the theremosyphon on the heat fl ux. The length fluids tested at various orientation angles. The figure of the thernosyphon was varied at 300mm, 450mm and indicated that R-134a is an excellent medium when 600mm. The results are shown in Figure 7, and they are compared with ethanol and distilled water. diffi cult to classify owing to the tested conditions only covering small differences in the aspect ratio. However, 3.3 Effect of total length when considering all the tested orientations, the length of The experiments also investigated the effect of the total 300 mm obtained the highest heat fl ux.

Figure 7. Effect of evaporator length on heat ﬂ ux at various tested angles (working temp. = 80oC).

3.4 Effect of inclination angle of evaporator and condenser

Figure 8. Effect of inclination angle of evaporator and condenser on heat ﬂ ux at various tested angles (R-134a).

Experiments were set up to examine the heat fl ux the case of the 90 degree orientation, lower thermal when modifying the evaporator and condenser section resistance values were obtained than at any other tested orientations to 0, 60 and 90 degrees with respect to the 70 orientation. The lowest thermal resistance from this degree inclination of the adiabatic section. From Figure. 8 experiment was 0.065°C/W it can be concluded that the 90 degree orientation was the best due to gravity promoting the enhancement of the heat 3.5 Effect of tested angle on thermal performance fl ux. However, in the case of the 0 degree orientation, the Figure. 10 displays the effect of the tested orientation on graph shows more fluctuation owing to the tested the thermal performance. When increasing the tested angle orientations of 0-60 degrees giving inappropriate positions from 0 to 40 degrees, the thermal performance sharply of the evaporator section, as can be seen in Figure. 8. rises, followed by a slight increase (60 to 110 degrees) and afterward it decreases gradually from 130 to 180 degrees. 3.5 Effect of thermal resistance It can be concluded that the optimal tested orientation When considering the thermal resistance being affected by angle is in the range of 40 to 110 degrees. the modifi ed section, as mentioned in 3.4, it is clear that in Heat transfer characteristics of thermo-syphon: Case study of modifying Vol 1. No 2, July-December 2015 inclination angle of evaporator and condenser sections 61

Figure 9. Effect of inclination angle of evaporator and condenser sections on thermal resistance at various tested angles (R-134a).

Figure 10. Effect of tested angle on efﬁ ciency at various working temperatures (R-134a).

4. Conclusion Acknowledgements The experimental results can be concluded as follows: The authors would like to express their appreciation to 4.1 The heat flux depends on the operating Rajamangala University of Technology Isan and the temperature. Research & Development Institute, Faculty of Engineering, 4.2 The best working fl uid that can achieve a high Rajamangala University of Technology Isan, Khon Kaen heat fl ux is R-134a, followed by ethanol and Campus for providing fi nancial support for attending The distilled water consequently. 7th International Conference on Science, Technology and 4.3 The heat fl ux decreased with an increase in the Innovation for Sustainable Well-Being 2015 (STISWB- evaporator length, and 100mm was found to be the best. VII). 4.4 In the case of modifi ed evaporator and condenser sections at 0, 60 and 90 degree orientations, the heat References fl ux increased as the orientation angle increased, and 90 Cengel, Y. A. 2002. Heat transfer a practical approach. 2nd degrees was the best (max. 26.85 kW/m2, min. 0.065 oC/W ed., McGraw-Hill, New York. of thermal resistance). Furthermore, when compared with Fadhl, B., Wrobel, L. C. and Jouhara, H. 2013. Numerical a conventional thermosyphon, it shows that the conven- modeling of the temperature distribution in a tional thermosyphon achieved a higher heat fl ux. two-phase closed thermosiphon. Applied Thermal 4.5 The optimal tested orientation angle was in the Engineering 60, 122-131. range of 40-110 degrees. Jiang, F., Chen, W. J., Liu, C. Z., Shi, J. T. and Li, X. L. 2014. Heat transfer enhancement in a three-phase closed thermosiphon. Applied Thermal Engineering 65, 495-501. 62 Kittipon Khangam and Narong Srihajong R & K

Jiao, B., Qiu, L. M., Zhang, X. B. and Zhang, Y. 2008. Investigation on the effect of fi lling ratio on the steady state heat transfer performance of a vertical two-phase closed thermosiphon. Applied Thermal Engineering 28, 1417-1426. On-ai, K., Kummuang-lue, N., Terdtoon, P. and Sakul- changsatjatai, P. 2013. Effect of working fl uid types on thermal performance of vertical closed-loop pulsating heat pipe. The 5th International Conference on Science, Technology and Innovation for Sustainable Well-Being, Luang Prabang, Lao PDR. Park, Y. J., Kang, H. K. and Kim, C. J. 2002. Heat transfer characteristics of a two phase closed thermosyphon to fi lling charge ratio. International Journal of Heat and Mass Transfer 45, 4655-4661. Shabgard, H., Xiao, B., Faghri, A., Gupta, R. and Weissman, W. 2014. Thermal characteristics of a closed thermosyphon under various fi lling conditions. Interna- tional Journal of Heat and Mass Transfer 70, 91-102.