Free Fatty Acid Profile of an Indigenous Chicken Breed (Basilan) of Mindanao, Philippines

Ryan Vincent C. Suazo1 and Henry Rivero2 MSU-Iligan Institute of Technology, Philippines [email protected], [email protected]

Abstract - The hardiness, adaptability to application into future nutritional and local-agro-climatic conditions, ability to epidemiological studies. utilize farm-byproducts, and resistance to diseases of the Philippine native chickens Keywords - Blood Plasma, Gas Chromatography, have been widely recognized (PCARRD Native Chicken, TLC, Transesterification 2008). Mindanao is home for this indigenous chicken breed, the Basilan 1. INTRODUCTION chicken (also known as Jolojano, Asil Malayo, in the Philippines) that is believed The wild ancestors of the domestic chicken to originate from the Asil (Aseel) group of originated in the South East Asia, where the India and Pakistan, and presently are found Red jungle fowl has the widest distribution of throughout Southeast Asia. These chickens the wild species and to be the antecedent of the are characterized by naked red skin in the native chickens in the Philippines. Basilan is a keel and in the wing. Native or indigenous general term used for the Philippine native chickens such as those belonging to this chicken strain indigenous to and most group from different sources in Mindanao dominant in Mindanao. Native chicken Island have been hypothesized to have Free constitutes a large portion of the total chicken Fatty Acid (FFA) profile variations as population of the Philippines, among them are contributed by the exogenous factors. This the Parawakan of Palawan, Darag of the study focused on the geographic variations Western Visayas, and the Basilan chicken of of fatty acid profile of representative native Mindanao which was the focus of the study. chicken breeds from four identified sources Native chickens are well known for their in Mindanao namely; Agusan, Surigao, adaptability to local agro-climatic conditions, Lanao, and Basilan Island, with two hardiness, ability to utilize farm-by-products representative from each source. Plasma and resistance to diseases. FFA composition, an indicator of FA prominence and dietary intake is linked The natural selection process of native with animal fitness on a short and long term chicken under local environment caused it to basis. Blood lipid and free fatty acid profile develop patterns of behavior that allows them of native chicken breed and the abundance to survive and reproduce even in the most of different fatty acid classes present were marginal environment and minimal determined. In this method, FFAs were management intervention. Some native extracted from the blood plasma using chickens are raised in the backyard of most isopropanol:chloroform solvent system and rural areas in the country. For the past years, directly transesterified for detection using these chickens have been part of the natural gas chromatography-mass spectroscopy setting and provide additional income for (GC-MS). The establishment of this fatty many farmers. Various breeds of native acid profiles in Mindanao indigenous chicken has been studied through time and has chicken breeds are important key for the been commercialized for its potential to the

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.1 Ryan Vincent C. Suazo and Henry Rivero market. Fatty acid measurement in adipose of blood of the native chickens of the tissue, whole blood, erythrocyte membrane, Philippines can give information needed to serum or plasma, and specific plasma fractions interpret the occurrences of health related serve as biomarkers of exogenously consumed problems such as exposure to infections, fatty acids (Sun Qi et al. 2007 and Baylin et al. intoxication and possible malnutrition that can 2006). Compared to other biomarkers, whole make an accurate diagnosis and effective blood as a biomarker of fatty acid assessment treatments of certain diseases and is acquiescent to the widespread usage in dysfunctions. Also, results of this study can epidemiological studies due to the relative provide data for future studies on physiology, ease of collection, processing and storage. nutrition and pathology of the chickens in Lipid composition and global properties of the general. lipid bilayer play key roles in membranes, often actively modifying the function of 2. METHODOLOGY membrane proteins (Charalambous et al., 2008), and the determination of the fatty acid A. Organism and Sampling profile in plasma and blood samples is used as A total of 44 sexually matured Basilan diagnostic tool in biomedical analysis (David breed chickens were collected from different et al., 2009). localities in Mindanao. Four groups of these chickens comprised the sample populations Fatty Acids (FAs) are chains of specifically from Lanao (del Norte/ del Sur) hydrocarbon groups attached to a carboxyl the (LB) group, Agusan (del Norte/ del Sur) group. They vary in chain length and the (AB) group, Surigao (del Norte/ del Sur) saturation and can be nonessential or essential the (SB) group and the Basilan (BB) served as (unable to be synthesized within the body) the control in this study being the putative pure (Saleh et al. 1998). The FA composition of Basilan breed. Each group comprised of 8 cellular and plasma lipids is major importance females and 3 males, and housed in separate for many biological functions. Conventional cages during the 2-week quarantine which was methods for analyzing the FA composition in located at residential lot in Brgy, Del Carmen. biological samples consist of several The chickens were fed with amaranth mixed analytical steps are cumbersome and time commercial feeds for their food supplement consuming. Therefore, new approaches focus twice a day. on simplifying and reducing sample preparation steps like lipid extraction, lipid B. Preparation of Erythrocytes and Plasma separation, and FAME synthesis. Venous blood of fast indigenous chickens overnight was drawn into tubes This study was focused on identifying any containing EDTA-2Na by wing venipuncture. geographical variations among the four The blood were kept cool in an ice chest after selected sources of indigenous chicken of collection and refrigerated before these were Mindanao which are presumed descendants of processed within 48 hours. Air-dried smears pure Basilan breed; determining the blood were prepared by the method as recommended plasma proteins of the different stages of by Hardie et al. (2002). native chickens (i.e., early laying and late laying); assessing whether age would be a Plasma phospholipid fatty acids were factor for variation on plasma proteins; and obtained from whole blood collected with establishing the baseline reference information EDTA. The blood samples in EDTA-coated on the Mindanao indigenous native chicken tubes were centrifuged at room temperature at that are presumed to belong to the pure about 1,000 rpm for 10 min. and using a Basilan breed of Mindanao, Philippines. The micropipette, the plasma was removed from thorough determination and identification of the packed cells and were transferred into new different and specific plasma protein contents tubes where a 500µl plasma aliquot was set

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.2 Free Fatty Acid Profile of an Indigenous Chicken Breed (Basilan) of Mindanao, Philippines aside for the determination of fatty acids. A gas chromatography instrument expertise was required. Erythrocyte membrane fatty C. Determination of Lipid Composition of acids were extracted from pelleted Erythrocytes erythrocytes with chloroform:methanol (2:1, Extraction of lipids from erythrocytes was v/v), evaporated to dryness and were extracted essentially performed according to the method with hexane. Detection and quantitation were reported by Mawatari et al. (2007). Concisely, performed using gas chromatography. Fatty 500 µl of packed erythrocytes were hemolyzed acid methyl ester from 12:0 through 24:1n9 with an equal amount of 10mM phosphate were detected using a 50 min gradient from buffer (pH 7.4). Four milliliter of methanol 190°C to 240°C. were added to the lysate followed after 40 min by 4 ml chloroform. After an 3. RESULTS AND DISCUSSION additional 30 min, the extract was centrifuged and the residue were re-extracted with 4 ml of A. Plasma Fatty Acids of Indigenous Chicken methanol/ chloroform (1:1, v/v). Methanol and Breeds of Mindanao chloroform used in the lipid extractions The plasma fatty acid profiles of the contains butylhydroxytoluene (50mg/l). indigenous chickens from different Pooled extracts were washed with 10ml of provenances in Mindanao are useful 50mM KCl to make a biphasic mixture. One information in comparing the lipid milliliter of the lower phase of the lipid extract composition and the geographic differences in was dried under N2 gas. the fatty acids that define the local native chickens. D. Fatty Acid Detection by FT-IR Plasma lipids were extracted using the B. The Quality of the Hexane Extract method of Folch et al. (1957). The fatty acids The overall consistencies of the hexane in the processed plasma were esterified to extracts from the blood of the indigenous methyl esters using 14% boron trifluoride in chickens of Mindanao that were used for FTIR methanol and followed by extraction with analysis and later for GC-MS are shown in hexane. The esterified fatty acids were kept in Fig 1. tightly sealed tubes for detection of the fatty acids by Fourier Transformed Infrared (FTIR) analyzer at the Chemistry Department of MSU-Iligan Institute of Technology. The detected fatty acids in samples were repacked for Gas Chromatography-Mass Spectrophotometry (GC-MS) analysis at the Natural Sciences Research Institute of the

University of Philippines in Diliman, Quezon City. Prior to subjecting the analytical Fig 1. Initial Hexane Extracts (Unfiltered) procedures for GC-MS, scanning FTIR of the Blood Plasma of the Indigenous Chickens analysis was done according to the methods by from Different Provenances in Mindanao. Sjahfirdi et al. (2011). The FTIR scanning process was conducted with wavenumber C. Fatty Acid Functional Group Detection by ranging from 400-4000cmˉ¹ with resolution of FT-IR Spectroscopy 4cmˉ¹. The scanning results were percentage This part of the research specifically absorbance on specific wavenumber for every identified the functional groups (FGs) of amide functional group in each sample. interest for putative fatty acids (FAs) in the plasma of the Mindanao indigenous chicken E. Gas Chromatographic Analysis of Plasma breeds from different provenances. Different Fatty Acids FGs have specific assignments for

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.3 Ryan Vincent C. Suazo and Henry Rivero wavenumber (cm-1) for the identification and Mass Spectroscopy instrument. Spectral detection of the molecular motions as analysis of FTIR showing different types of summarized in Table 1. functional groups of the protein associated with blood lipids. Transmittance indicating TABLE 1 high concentrations in the blood plasma -1 SUMMARY OF THE WAVENUMBER (CM ) samples. ASSIGNMENTS OF THE MOLECULAR FUNCTIONAL GROUPS AND MOTIONS IN ANY GIVEN COMPOUND E. Fatty Acid Scanning by GCMS Gas Chromatography is a more convenient Functional Molecular Motion Wavenumber and precise method for qualitative and Group (cm-1) quantitative analysis of fatty acid methyl ester Amines N-H stretch 3417 (FAME), and comparative chromatographic Amines N-H stretch 3416 Alkanes C-H Aliphatic stretch 2961 analysis of changes in the concentrations of Alkanes C-H Aliphatic stretch 2925 FAME. When GC is combined with MS, a Alkanes C-H Aliphatic stretch 2853 powerful analytical tool is created. A Mercaptans S-H stretch 2359 Mercaptans S-H stretch 2341 researcher can take an organic solution, inject Anhydrides C=O stretch 1743 it into the instrument, separate the individual Alkenes C=C stretch 1659 components, and identify each of them. (isolated) Furthermore, the researcher can determine the Alkenes C=C stretch 1652 (isolated) quantities (concentrations) for each of the Alkenes C=C stretch 1651 components. (isolated)

Amines N-H bend 1558  Agusan Stock Alkanes CH2 bend 1466 Alkanes CH2 bend 1457 The IR spectral analysis of Agusan stock in Alkenes C-H in-plane bend 1416 Fig 2. showed signals at wavelength ranges Alkyl C-F stretch 1380 characteristics of the functional groups that are Halides Alkyl C-F stretch 1379 commonly found in fatty acids. Halides Alkyl C-F stretch 1378 Halides

Source: Fourier Transform Infrared Spectroscopic Analysis of Protein Secondary Structures, (Kong and Yu 2007).

D. FT-IR Profiling of Detectable Compounds FTIR has been used as a universal instrument in analyzing varieties of samples to identify the functional groups of any given chemical compounds such as carbohydrate and esters, as well as inter-atomic chemical bonds. FTIR has high accuracy level in the process in identifying the aforementioned. It is Fig 2. IR Spectroscopic Detection of Fatty Acid also safe, fast and sensitive (Smith, 1979) and (Methyl Ester) in the Plasma of Indigenous Chickens has been used in research to analyze chemical from Agusan Provenance. components of gallstone (Kleiner et al. 2002), blood samples of renal failure patients (Devi GCMS chromatogram of the esterified fatty et al. 2009), protein plasma in blood (Petibois acid in the plasma of the indigenous chicken of et al. 2001), and breast cancer diagnosis Agusan provenance in Fig 3. showed (Dimitrova et al. 2009). Functional groups measured peaks at different retention time. were detected by FTIR at different levels of The peaks in the chromatogram represents the signals and specified by Gas Chromatography- signal created when a compound elutes from the GC column into the detector. Each of the

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.4 Free Fatty Acid Profile of an Indigenous Chicken Breed (Basilan) of Mindanao, Philippines peaks represents an individual compound that was separated from a sample mixture.

Fig 4. IR Spectroscopic Detection of Fatty Acids in Plasma of Indigenous Chickens from Lanao Provenance.

GCMS chromatogram of the esterified fatty acid in the plasma of the indigenous chicken of Fig 3. GCMS Chromatogram Lanao provenance in Fig 5. showed measured (A, Full View; B, Zoom View of 27-27.7 min) of the Esterified Fatty Acid in the Plasma peaks at different retention time. Gas of the Indigenous Chicken of Agusan Provenance. chromatogram showed detected peaks containing different compounds with The fatty acids detected by GCMS from the corresponding molecular formula and blood plasma of the indigenous chicken of molecular weight at different retention time. Agusan provenance, summarized in Table 2, Each of the peaks represents an individual shows that of the eight fatty acids detected, compound that was separated from a sample seven are saturated fatty acids (SFA) and one mixture. is a monounsaturated fatty acid (MUFA). There was no polyunsaturated fatty acid (PUFA) detected.

TABLE 2 DETECTED FATTY ACIDS IN THE BLOOD PLASMA OF AGUSAN STOCK CHICKEN

Fig 5. GCMS Chromatogram (A, Full View; B, Zoom View of 23-29.3 min) of the Esterified Fatty Acid in the Plasma of the Indigenous Chicken of Lanao Provenance.

The fatty acids detected by GCMS from the Total No. of Fatty Acids Detected = 8 blood plasma of the indigenous chicken of Lanao provenance, summarized in Table 3,  Lanao Stock shows that from the seventeen fatty acids The IR spectral analysis of Lanao stock in detected, five are saturated fatty acids (SFA), Fig 4. showed signals at wavelength ranges six are monounsaturated fatty acid (MUFA), characteristics of the functional groups that are and six polyunsaturated fatty acid (PUFA). As commonly found in fatty acids. summarized in Table 4, The detected ones are steroids in the blood plasma of Lanao Stock.

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.5 Ryan Vincent C. Suazo and Henry Rivero

TABLE 3 DETECTED FATTY ACIDS IN THE BLOOD PLASMA OF LANAO INDIGENOUS CHICKEN

Fatty Acid MW Chemical Formula Na me Designation Myristic acid 14:0 242 C13H27COOCH3 Palmitic acid 16:0 270 C15H31COOCH3 Margaric acid 17:0 284 C16H33COOCH3 Stearic acid 18:0 298 C17H35COOCH3 Arachidic acid 20:0 326 C19H39COOCH3

7-hexadecenoic 16:1n9 268 C15H29COOCH3 acid Fig 6. IR Spectroscopic Detection of Fatty Acids 16:3n6 264 C15H25COOCH3 in Plasma of Indigenous Chickens 17:3n6 278 C16H27COOCH3 from Surigao Provenance. 18:1n7 296 C17H33COOCH3

18:1n8 296 C17H33COOCH3 Oleic acid 18:1n9 (cis) 296 C17H33COOCH3 Esterified fatty acid in the plasma of the Elaidic acid 18:1n9 (trans) 296 C17H33COOCH3 indigenous chicken of Surigao provenance in Fig 7. was specified by the GCMS. Gas 18:1n11 296 C17H33COOCH3 18:2n2 294 C17H31COOCH3 chromatogram showed detected peaks Linoleic acid 18:2n6 294 C17H31COOCH3 containing different compounds with 20:4n3 318 C19H31COOCH3 Arachidonic 20:4n6 332 C19H31COOCH2CH3 corresponding molecular formula and acid molecular weight at different retention time.

Total No. of Fatty Acids Detected = 17

TABLE 4 DETECTED STEROIDS IN THE BLOOD PLASMA OF LANAO INDIGENOUS CHICKEN

Fig 7. GCMS Chromatogram (A, Full View; B, Zoom View of 29-31.5 min) of the Esterified Fatty Acid in the Plasma of the Indigenous Chicken of Surigao Provenance.

The fatty acids detected by GCMS from the blood plasma of the indigenous chicken of  Surigao Stock Surigao provenance, summarized in Table 5, The IR spectral analysis of Surigao stock shows that of the twelve fatty acids detected, in Fig 6. showed signals at wavelength ranges five are saturated fatty acids (SFA), four are characteristics of the functional groups that are monounsaturated fatty acid (MUFA), and three commonly found in fatty acids. are polyunsaturated fatty acid (PUFA).

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.6 Free Fatty Acid Profile of an Indigenous Chicken Breed (Basilan) of Mindanao, Philippines

TABLE 5 DETECTED FATTY ACIDS IN THE BLOOD PLASMA OF SURIGAO INDIGENOUS CHICKEN

FATTY ACIDS Designation MW Chemical Formula Myristic acid 14:0 242 C13H27COOCH3 Palmitic acid 16:0 270 C15H31COOCH3 Margaric acid 17:0 284 C16H33COOCH3 Stearic acid 18:0 298 C17H35COOCH3 Arachidic acid 20:0 326 C19H39COOCH3 Hexadecadienoic 16:2n4 266 C15H27COOCH3 acid

18:1n6 296 C17H33COOCH3 Vaccenic acid 18:1n7 296 C17H33COOCH3 Fig 9. GCMS Chromatogram 18:1n8 296 C17H33COOCH3 (A, Full View; B, Zoom View of 26-31.7 min) Oleic acid 18:1n9 (cis) 296 C17H33COOCH3 of the Esterified Fatty Acid in the Plasma Linoleic acid 18:2n6 294 C17H31COOCH3 of the Indigenous Chicken of Basilan Provenance (Control Group, Presumed Pure Basilan Breed Eicosadienoic acid 20:2n6 322 C19H35COOCH3 from the Mainland Basilan).

Total No. of Fatty Acids Detected = 12 The fatty acids detected by GCMS from the blood plasma of the indigenous chicken of  Basilan Stock Basilan provenance, summarized in Table 6, The IR spectral analysis of Basilan stock in shows that of the sixteen fatty acids detected, Fig 8. showed signals at wavelength ranges five are saturated fatty acids (SFA), four are characteristics of the functional groups that are monounsaturated fatty acid (MUFA), and commonly found in fatty acids. seven polyunsaturated fatty acid (PUFA). Detected steroids in the blood plasma of Basilan indigenous chicken was shown in Table 7.

Fig 8. IR Spectroscopic Detection of Fatty Acids in Plasma of Indigenous Chickens from Basilan Provenance.

GCMS chromatogram of the esterified fatty acid in the plasma of indigenous chicken of Basilan provenance in Fig 9. showed different detected compounds and lipids of interest with corresponding molecular formula and molecular weight at different retention time.

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.7 Ryan Vincent C. Suazo and Henry Rivero

TABLE 6 detectable groups of fatty acids from the blood DETECTED FATTY ACIDS plasma of indigenous chicken. Agusan IN THE BLOOD PLASMA OF BASILAN INDIGENOUS CHICKEN indigenous chickens were the sole chicken stock that exhibited the presence of Undecanoic acid (C11:0), Lauric acid (C12:0) and Heptacosanoic acid (C27:0) which are saturated fatty acids. Another saturated fatty acid detected was the Tridecanoic acid (C13:0) which was presented only in Agusan and Basilan stocks. Palmitic acid (C16:0) was also a saturated fatty acid detected in Lanao and Surigao indigenous chickens. Arachidic acid (C20:0) was the most common saturated fatty acid present from the indigenous chicken among the three provenances, Lanao, Surigao, and Basilan stocks. PUFAs acids like n3 and n6 were also detected in all samples. Basilan indigenous chicken was the sole chicken stock that showed the presence of Eicosapentaenoic acid (EPA, C20:5n3) which could be converted into a group of biologically-important substances

known as eicosanoids (Montgomery et al. Total No. of Fatty Acids Detected = 16 1990) and Linolenic acid (C18:3n3).

TABLE 7 A number also of omega-6 fatty acids have DECTED STEROIDS also been found, which include the Linoleic IN THE BLOOD PLASMA acid (C18:2n6) present in all indigenous OF BASILAN INDIGENOUS CHICKEN chicken except Agusan stock; and the STEROID NAME MW Chemical Formula Arachidonic acid (C20:4n6) found only in .gamma.Sitosterol; Sitgmast-5-en- 414 C29H50O 3-ol, (3.beta,24S)-; Stigmast-5-en- Lanao and Basilan chicken stock. Both Lanao 3.beta.ol (24S)-; Clionasterol; and Basilan chicken had Elaidic acid Fucosterol, .beta-dihydro- (C18:1n9) trans, which was a monounsaturated Cholesterol; Cholest-5-en-3-ol 386 C27H46O (3.beta)-; Cholesterol; Cholest-5- type (MUFA). A type of omega-7 fatty acid, en- 3.beta.-ol; Cholesterin; Vaccenic acid (C18:1n7) was also detected Cholesterol base H

Cholestane-3, 5-diol, 5-acetate, 446 C29H50O3 from both indigenous chickens of Surigao and (3.beta., 5.alpha) Basilan, which naturally occurs as trans-fatty Cholest-5-ene, 3-bromo-, (3.beta)-; 448 C27H45Br Cholest-5-ene, 3.beta.-bromo-; acid in ruminants. The Basilan indigenous Choleesteryl bromide; 3.beta.- chicken established different types of Bromocholest-5-ene monounsaturated fatty acids including the Cholesterol chloroformate; 448 C28H45ClO2 Cholesteryl chloroformate; Hexadecadienoic (C16:2n4), Hexadecatrienoic Cholesteryloxycarbonyl chloride (C16:3n6), and Heptadecatrienoic (C17:3n6)

acids. The presence of C18:1n8 a type of F. Fatty Acid Profile of Indigenous Chicken omega-8 was detected in the blood plasma of Breed of Mindanao Lanao, Surigao, and Basilan stocks. Natural The overall profile of free fatty acids of the steroids were also found to be in the blood indigenous chicken of Mindanao is plasma of Lanao and Basilan indigenous summarized in Table 8 and the profile of each chicken which could have valuable benefits in provenance was compared according to the human nutrition.

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.8 Free Fatty Acid Profile of an Indigenous Chicken Breed (Basilan) of Mindanao, Philippines

TABLE 8 from Lanao provenance, 16 fatty acids from FATTY ACID PROFILE Basilan, 12 fatty acids from Surigao, and a OF INDIGENOUS CHICKEN BREED (BASILAN) OF MINDANAO total of 8 fatty acids from Agusan. Therefore, fatty acid profiles of the four representative

PROVENANCES provenances can be established by the FATTY ACID Agusan Lanao Surigao Basilan scanning method of GC-MS.

SFA 11:0 14:0 14:0 13:0 ACKNOWLEDEGMENT 12:0 16:0 16:0 14:0 13:0 17:0 17:0 17:0 The authors would like to express their 14:0 18:0 18:0 18:0 17:0 20:0 20:0 20:0 deepest gratitude and appreciation to the 18:0 people who offered their valuable time and 27:0

immense knowledge to make this study a MUFA 18:1n9 16:1n9 18:1n6 18:1n7 success. This work was patriotically funded. 18:1n7 18:1n7 18:1n8 18:1n8 18:1n8 18:1n9 18:1n9 18:1n9 (cis) REFERENCES (cis) (cis) 18:1n9 18:1n9 (trans) (trans) (Arranged in the order of citation in the 18:1n11 same fashion as the case of Footnotes.) - PUFA 16:3n6 16:2n4 16:2n4 [1] Baylin A, Campos, H. “The use of fatty

17:3n6 18:2n6 16:3n6 acid biomarkers to reflect dietary 18:2n2 20:2n6 17:3n6 18:2n6 18:2n6 intake”. Curr Opin Lipidol 2006, 17:

20:4n3 18:2n7 22-27. 20:4n6 20:4n6 20:5n3 [2] Charalambous, K., Miller, D., and

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[3] Devi, T.S.R., Gunasekaran, S., Hudson, 4. CONCLUSION J.W., and Joybell, I.S.A. “Analysis on Fatty acid profiles of indigenous chickens renal failure patients blood samples: from identified provenances in Mindanao characterization and efficacy study”. established different fatty acid classes. The Indian Journal of Science and existence of saturated, monounsaturated Technology, 2(2), 46-50. (MUFA) and polyunsaturated fatty acid [4] Dimitrova, M., Ivanova, D., (PUFA) specified the potential sources of fatty Karamancheva, I., Milev, A., and acids in these indigenous chicken. Common Dobrev, I. “Application of FTIR- saturated fatty acids identified by GCMS spectroscopy for diagnosis of breast among the blood of the indigenous chicken cancer tumors”. Journal of the from four provenances include: Myristic acid University of Chemical Technology and (14:0), Margaric acid (17:0), and Stearic acid Metallurgy, 44(3), 297-300. (18:0), and Oleic Acid (18:1n9). Oleic Acid [5] Folch, J., Lees, M., Sloane Stanley, GH. was identified to be the most common “A simple method for the isolation and monounsaturated fatty acid among four purification of total lipids from animal provenances. Linoleic acid (18:2n6), a tissue”. J Biol Chem; 226: 497-509. polyunsaturated fatty acid, was identified from [6] Hardie, D. et al. “From Pixels to the blood of indigenous chicken of Picograms: A beginner’s guide to Surigao, Lanao, and Basilan provenances, Genome Quantification”. Feulgen Image but not of Agusan. Gas Chromatography-Mass Analysis Dinsitometry, Volume Spectroscopy identified a total of 17 fatty 50(6):735-749. acids from the blood of indigenous chicken

The Fourth International Congress on Interdisciplinary Research and Development, 30 - 31 May 2014, Thailand 8.9 Ryan Vincent C. Suazo and Henry Rivero

[7] Kleiner, O., Ramesh, J., Huleihel, M., Cohen, B., Kantarovich, K., Levi, C., Polyak, B., Marks, R.S., Mordehai, J., Cohen, Z., and Mordechai, S. “A comparative study of gallstones from children and adults using FTIR spectroscopy and fluorescence microscopy”. BMC Gastroenterology, 2(3), 1-14. [8] Kong, J., and Yu, S. “Fourier transform infrared spectroscopic analysis of protein secondary structures”. Acta Biochimica et Biophysica Sinica, 39(8), 549-559. [9] Mawatari, S., Okuma, Y., and Fujino, T. “Anal Biochem”. 370, No. 1, 54-59. [10] Petibois, C., Cazorla, G., Cassaigne, A., and De´le´ris, G. “Plasma protein contents determined by fourier- transform infrared spectrometry”. Clinical Chemistry 47(4), 730-738. [11] Saleh, J., Summers, LKM., Cianflone, K., Fielding, BA., Sniderman, AD., and Frayn, KN. “Coordinated release of acylation stimulating protein (ASP) and triacyglycerol clearance by human adipose tissue in vivo in the postprandial period”. J Lipid Res; 39:884-91. [12] Sjahfirdi, L., Aziz, S.N., Maheshwari, H., Astuti, P., Suyatna, F.D., and Nasikin, M. “Estrus period determination of female rats (Rattus norvegicus) by Fourier Transform Infrared (FTIR) through identification of reproductive hormones metabolites in urine samples”. International Journal of Basic and Applied Sciences 11(3), 158- 163. [13] Smith, A.L. “Applied infrared spectroscopy: fundamentals, techniques, and analytical problem-solving”. A WileyInterscience Publication, John Wiley and Sons, New York, Chichester, Brisbane, and Toronto. [14] Sun Qi, Ma J., Campos, H., Hankinson, SE., and Hu, FB. “Comparison between plasma and erythrocyte fatty acid content as biomarkers of fatty acid intake in US women”. Am J Clin Nutr, 86:74-81.

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