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Food Structure

Volume 5 Number 2 Article 9

1986

Structural Characteristics of Eleusine Corocana (Finger ) Using Scanning Electron and Fluorescence Microscopy

C. M. McDonough

L. W. Rooney

C. F. Earp

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Recommended Citation McDonough, C. M.; Rooney, L. W.; and Earp, C. F. (1986) "Structural Characteristics of Eleusine Corocana (Finger Millet) Using Scanning Electron and Fluorescence Microscopy," Food Structure: Vol. 5 : No. 2 , Article 9. Available at: https://digitalcommons.usu.edu/foodmicrostructure/vol5/iss2/9

This Article is brought to you for free and open access by the Western Dairy Center at DigitalCommons@USU. It has been accepted for inclusion in Food Structure by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. FOOD MICROSTRUCTURE, Vol. 5 (1986) , pp. 24 7-256 0730-5419/86$1. 00• . OS SEM, Inc . , AMF O'Hare (Chicago), IL 60666 - 0507 U.S.A.

STRUCTURAL CHARACTERISTICS OF ELEUSINE COROCANA (FINGER MILLET) USING SCANNING ELECTRON AND FLUORESCENCE MICROSCOPY

C.M . McDonough, L. W. Rooney, and C.F. Earp

Cereal Quality Lab, Dept. of Soil & Sciences, Texas A&M University, College Station , Texas 77843-2474

Abstract Introduction

The objective of this study was to docu­ Eleusine corocana {f inger mi I let) i s a ment the microstructure of finger millet wlth sma ll , round-seededo/ain that is widely used scanning electron and fluorescence microsco­ in and some parts of for food pro­ pies. Finger millet is an utricle which is ducts, i.e., , , pudding, cake and por­ spheri ca 1 and about 1 . 5 ITlT1 in diameter. The ridges. The originated in membranous peri carp of finger millet was loose­ and was subsequently introduced into Ind ia by ly associated with the at maturity. sea traders around 3000 B.C. (Hilu et al , 1979; Beneath the loose pericarp wa s a five-layered Hilu and de Wet, 1967; Mehra , 1963 ; Phillips, testa that varied from red to purple in color. 1971). Currently, nine of the eleven culti­ The outer layer was the only testa layer that vated and wild species are found in Africa, autofl uoresced, suggesting the presence of phe­ with the other two species fo und in India. The nolic acids, i . e., . The domesticated species of finger millet - Eleu­ layer was beneath the testa , and was one cell sine corocana ssp. corocana (Mehr a, 1963-1 - layer thick. The starchy had dis­ were invest1gated i"iltFiTSStudy . tinct peripheral, corneous and floury areas. The cultivated Eleusine is an annu­ The ce 11 wa 11 s of the endosperm strongly fl uo­ al that grows up to ~igh, with digitate, resced indicat ing phenolic compo un ds . non- sp ikes that can be arranged in granules were primarily compound, with some sim­ one of three configurations: open (spikes are ple granules in the corneous area . Starch gra­ straight and loose), top-curved (l-2 em of the nu 1e s ize inc rea sed toward the center of the s pike is curved ) , or i n-curved (entire spike is endosperm, wh ile the content decreased. curved) (Hilu and de Wet, 1967; Phillips, 1971 ; The small germ was inset into a shallow depres­ Mehra , 1963). The spike length ranges from sion; a s hort ridge protruded from the utricle 3.5-15 em, depend ing on the location and cli­ ar ound the perimeter of t he germ. Finger mi 1- mate. The grain is globose in shape , l.Z-1.8 let was higher in phenol and tannin content mm in diameter, with a granulated surface tex­ than . Moderate levels of genti­ ture (Hilu and de Wet , 1967). Utricles can sic , cinnamic and coumaric acids , and high appear yellow, whi te, tan, red, brown or vio­ levels of ferulic acid wer e extracted from fin ­ let. Each utricle is characterized by a shal­ ger mi 11 et. low depress ion of the germ and a characteristic protruding ridge around the depression (Hilu and de Wet, 1967; Hilu et al, 1979) . Wh en identifying cultivated finger millet in arc haeologica l and agricultural sites , the Initial paper received February 17, 1986 ridge around the germ depression and the pre­ Manuscript received November 12 , 1986 sence of non-shattering spikes are the two most Direct inquiries to C.M. Mc Donough important determining characteristics (Mehra, Telephone number: 409 845 2925 1963 ; Hilu et al, 1979). Finger millet has a distinctive morphologi­ cal characteristi c that is found a l so in proso and foxtail . The kernel is an utricle and not a t rue caryopsi s (A ng old , 1979) . Th e KEY WORDS: Fluorescence microscopy, finger pericarp is membranous and surround s the en­ ~icrostructure , Eleusine corocana, tire seed, but it i s not fused to the testa scanning e 1ectron microscopy , f1 uorochromes, (Phillips , 1971; Hilu et al, 1979; An9old, rag1. 1979). The peri carp is easily re!Tioved from the utricle by rubbi ng lightly or soaking in water,

247 C.M. McDonough, L.W. Rooney and C.F. Earp and often comes off during harvesting. Hilu et features of the finger millet utricle in al (1979) r eported that dur ing seed develop­ greater detai 1 than has previously been ment, the outer in tegument and part of the peri­ published. carp were resorbed after fertilization, and the inner integument and the rest of the pericarp Materia Is and Methods remained to develop into the thick testa layers and the membranous pericarp. Information on Samp 1es the structure of the testa layer lS not - -T-hree 10 g samples of finger millet available. (E 1eusine corocana ssp. corocana) were Angold (1979) reported that the endosperm obtained from the millet quality trials in had distinct floury and corneous layers ; the Cinzana , , in 1983. These samples were corneous areas fractured along cell wall s, from two different locations. Two of the sam­ while the floury areas fractured through the ples were reddish-brown in color and the third cell. Angold (1979) reported that the starch was reddish-. No visible morphological granules were predominantly smooth and com ­ differences between the , other than pound, Wankhede et al (1979) reported polygonal color,were apparent. Pearl millet (Pennise- granules (8 -15~m), and Paramahans et al (1g80) tem americanum) samples were included in the reported round granules {5-B~m) . Wankhede et chemica I ana lyses for comparison, and were also al (1979) also reported the hilum of the starch obtained from the millet quality trials in granules as faint but visible; they did not Cinzana , Mali, 1983 . mention if subunits within the starch granules Scanning E lee tron Microscopy were visible. The granules were strongly Representative seeds were broken in birefringent under polarized light (Wankhede et half with a dull razor bl ade and mounted on a!, 1979) and gelatinized between 65-85°C aluminum stubs with conductive carbon paint. (Paramahans et a I, 1980). The stubs were coated with a thin layer of The form of protein in the endosperm, gold-palladium, and viewed on a JEOL JSM25 i.e., protein bodies and protein matrix in fin­ scanning electron microscope with an ger millets, was not clearly do cumented accelerating voltage of 12.5 kV. (Muralikreshna et al, 1982; Wa nkhede et al, Fluorescence Microscopy 1979; Tharanathan et al, 1980; Paramahans et Representative kernels were fractured with al, 1980). Wada and Mae da (1980) found protein a razor blade, fixed in a 3% glutaraldehyde­ bodies 1-5 IJ.m in diameter in the scutellum and /phosphate buffer solution (pH=6.8), and then 1-41J.m in the aleurone layer, but they did not passed through an dehydration series : mention the presence of protein bodies in the 70% ethano I , 80% ethano I , 90% ethano I and 95% endosperm. in the form of phytic ethanol for 24 h each. The samples were embed­ acid was found in high levels pr imarily in the ded using the LKB Historesin Embedding Kit scutel lum and secondari ly in the aleurone cells (LKB , Bromma, Sweden). After dehydration, the (Wada and Maeda, 1980; Pore and Magar, 11)79). utricles were left in an infiltration solution Finger millet is not usually decorticated of 50:50 Historesin (no hardener):ethanol for prior to grinding, so phenols in the thick 24 h. Then the utrlcles were left in 100% testa that surrounds the endosperm and germ are Historesin (no hardener) for 24 h, and subse­ included in the . Ramachandra et al quently embedded i n Historesin (with hardener). (1977) found total phenol contents of 0. 8% and They were allowed to dry overnight, and then l.03% in white and brown finger millet, respec ­ sectioned on a rotary microtome at l-2 ~m tively, using the Falin-Denis procedure. They thickness . also reported tannin levels of 0.50% and 0 .61% Stained and unstained slides were prepared in white and brown finger millets, respective­ for viewing by putting a drop of oi 1 on the sur­ ly, using the vanillin -HCl procedure (Maxson face of the slide (with dried sections) and add­ and Rooney, 1972). Hilu et al (1978) reported ing a cover slip . The slides wer e viewed under that the majority of the phenolic compounds in a Zeiss Universal microscope equipped with an finger millet were . Ramachandra et IIIRS epi-illuminating system. Filter combina­ al {1977) found that finger millet samples with tion I was used for autofluorescence (un­ high phenol contents were lower in in vitro stained) and ANS stained slides (exciter filter protein digestibility than similar ITn9errii"il­ 365 nm, barrier filter > 418 nm). Slides let samples that contained fewer phenolic stained with Acid Fuchsin were viewed under fil­ compounds. ter combination III (exciter filter 546 nm, bar­ The use of fl uorochromes and fl uorescence rier filter > 590 nm). Pictures were taken on microscopy in the study of structure has Kodak Ektachrome fi 1m. been we II documented for pear I mi II et (Irving, Bright Field Microscopy 1983; McDonough, 1986), , and Finger m1 II et samp 1es were ground by hand (Fulcher and Wong, l980; Fulcher and Wood, in a mortar t o avoid contamination from other 1983), and (Earp et al 1983a,b; Earp cereal . A small flour sam pl e was 1984) . No studies have been publi shed describ­ placed on a slide wi th glycerol, and viewed ing the use of fluorescence microscopy in the with polarized light to detect birefringence of study of finger millet. starch granules. The objective of this study was to utilize scanning electron and fluorescence microscopy Chemi~~~ ~~~~t~~swere analyzed for nitrogen , to determine the structural and chemical starch and phenol content on a Technicon

148 Structural Characteristics of Finger Millet

~: A - Overall view of the endosperm and pericarp. B - Low magnification (4x) view Results and Discussion of finger millet utr i cles showing the membra­ nous pericarp before removal. C - Pericarp, Chemica 1 Ana lyses Finger millet was lower in protein and testa, aleurone and peripheral endosperm starch and higher in polyphenol and tannin layers. 0 - High magnification of the pericarp content than pearl millet (Table 1). Tannins sti 11 attached to the kernel . P - peri carp , T have been rep or ted to negatively af feet the = testa, pe = peripheral endosperm, ce = cor­ protein of flour by binding the protein so that neous endosperm , d =junctions, a = aleurone, it is not biologically available. The relative­ pm = protein matrix , sg = starch granule. ly high tannin content of red finger millet , combined with the low protein content, could significantly decrease the nutritional value of the resulting food product. Autoanalyzer llC system . The Folin-Ciocalteu Gross Morpho 1 ogy method of Kaluza et al (1980) was used to The flnger millet utricles were approxi­ determine the total polyphenol content, and the mately round and averaged 1.5 rrrn in diameter. automated vanillin-HCl method of McDonough et They had a 1000 kernel wei ght of 2.64 g. A al {1983) was used to determine the tannin con­ cross section of the finger mi I let utricle is tent. Crude protein values were obtained via presented in Figure lA. The pericarp was not Kjeldahl digestion, and were analyzed with a fused to the testa and only a portion of it was Technicon Autoanalyzer IIC (Technicon Indus­ retained (Figs . lA-B). A thick, red testa trial Systems, 1976). Total starch content was layer surrounded the entire s urface of the endo­ determined with the hexokinase method sperm. Directly beneath the testa was the (Technicon Instruments Corporation, 1978) . aleurone layer (1 cel l layer thick; Fi g. lC). Finger mi I let samples from each location were The endosperm had peri phera 1, corneous and prepared for high performance liquid chromato­ floury areas similar to those reported for sor ­ graphy (HPLC) analysis us ing the base hydroly­ ghum (i:_ bicolor), corn (.L._ ~) , and sis method of Hahn et al (1983); the samples pearl mi 1~ americanum) (Zeleznak and were analyzed in dup 1 i ca te on a Beckman HPLC Varriano-MarstOO ~ey et al 1983) . system with a 10 ~m C-18 column. Chemical Peri carp analyses, other than the HPLC analysis, were ------rti'e peri carp of finger millet was loosely conducted in triplicate, and reported on a dry associated with the surface of the testa. The weight basis. pericarp was not fused to the testa at any par-

249 C. M. McDonough, L.W. Rooney and C.F. Earp

Table 1 1 Chemical Analyses of Finger and Pearl Millets

Protein Starch Total Polyphenol Tannin 1000 Kerne I Wt (%) (%) (%) (%) (gms)

Finger Millet 8.28-8.56 70.6-75.2 0. 55-0.59 0.17 -0 .32 2.64

Pearl Mi 1\et 22 10.20-14.40 76 .7-86.1 0 .19-0. 33 0 .0 6 .8-14.3

Values repres en t means of three replicates per sample , dry weight basis . 2 Nx6.25.

Table 2 When viewed in cross section, junctions between the mounds in the outer testa layers were visi­ ~henolic Acid Analyses 1 of F1nger and Pearl Millets ble (Fig. lC); these probably correspond to the "interlocking" sections seen from the surface 2 3 Pheno 11 c Finger Millet Pearl Millet (Fig. 2A). Acids Location 1 Location Range The testa contained five distinct layers (Fig. 3A , 8) . The first layer was 1.5 11m thick (Fig. 3A), and autofluoresced blue, indi­ Ferulic 405 370 624.7-786.3 cating the presence of ferulic acid or lignin Coumari c 67 46 211.4-346.6 (Fulcher et al., 1972). Beneath this was the Gentisic 53 70 79.0-114.2 second and thickest layer which contained the Cinnamic 35 35 271.4-415.1 mounds already described (Fig. 2). The layer Catfeic 15 17.7 11.3- 37.5 was from 5.5-17. 5 1Jm thick and appeared to be Vanillic 15 6. 5- 26 .1 striated . This large layer had darker pigmenta­ Protaca techui c 14 33 3.8- 22 .7 tion than the lower layers, and thus coul d con­ p-OH Benzoic 9 15 .8- 26.0 tain different phenol i c compounds than the Syri ngi c 7 10.5- 23.7 others. The third and fourth layers were Sinapic 4 15.4- 27 .7 approximately the same thickness (1.4-2.1 ~m) , Total Unknowns 149 147 646.5-892.8 although the striated patterns appeared to be Tot a 1 Acids 173 126 1896-2118 different. The third layer had distinct wave formations throughout, wh ile the fourth layer Values expressed as ~J,g phenolic acidS/IJTI was predominantly straight, with some isolated samp l e , dry weight basis. wave patterns (Fig. 38). Both layers appeared ~Seed from locations in Mali, West A~rica. to be close to the same shade in color. The Values are the averages of two repl1cates each fifth layer was 1 IJffi wide and was distinctly of a bronze, slate, and tan var iety . different in color from the previous layers as These va 1 ues are included only for comparison . seen in Figure 36 . When viewed with autofluor­ escence, the top (#1) layer was the only one ticular place. The peri carp was a fragile, mem­ that fluoresced . The other layers were visible branous layer that was easily removed by rub­ but were illuminated only by fluorescence from bing or washing prior to use. Figure lB shows other structures; they did not fluoresce them­ the utricles with varying amounts of the peri­ selves. carp covering the dark testa which is the pro­ The testa appeared to contribute the bulk tective covering of finger millet. Figures lA of the polyphenol and tannin compounds to the and lC show the details of a portion of the flour . However, no hand dissection studies pericarp that remained on the kernel through­ were performed to conf irm this . The sam ples in out sample preparation. Figure 10 shows the this study contained 0.57 mg/100 mg total poly­ edge of the pericarp. There were several phenols (Folin - Ciocalteu test) and a maximum of layers of tissue visible, but there were no 0.32 mg/lOOmg equivalents (vanillin/­ cell contents observed. Collapsed cells in the HCl test; Table 1). Phenolic acids of finger outer layers of the pericarp are visible in millet co nsisted of high levels of ferulic acid Fig. 1C -O. and only moderate levels of gentisic, cinnamic Testa Layers and coumar ic acids (Table 2). Finger millet The extern a 1 appearance of the finger contained lower quantities of phenolic acids millet testa was quite striking and different than pearl millet. Tannins are measured by the from other . The first layer was com ­ vanillin method and also react positively in posed of sections of tissue that "interlocked" the total polyphenol determination. The tan ­ like the pieces of a jigsaw puzzle (Fig. 2A). nins were not determined by the HPLC methodolo­ Each section was composed of 2-4 dimpled gy. Therefore, there is a difference between mounds. There were open spaces underneath some the phenols determined by HPLC versus the color­ of the mounds that contained granules of imetric methods . The outermost testa layer, unknown composition (Fig . 2B); it is unknown if and the a leurone and endosperm cel l walls were these open spaces were true characteri sties of the only areas that exhibited blue autofluor­ the layer, or if they were artifacts (Fig. 2B) . escence, which indicated that ferulic aci d was

250 Structural Characteristics of Finger Millet

~igure 2: A ~ Top view of the "interlock­ Figure 3: Cross section of the testa . A - log", mound-llke structures found in the testa. The five testa layers in re la tion to the aleu­ B - Cross section of the testa showing a mound rone . 8 - Four of the five testa layers, with an open space located beneath it. M = showing wave formations and contour striations . mound, T = testa, sp = space, a = aleurone, pe 1-5 =testa layers , w = wav e formation, A= peri phera 1 endosperm . aleurone cell , a= aleurone cell wall.

present in those areas. Ferulic acid has been Starchy Endosperm shown previously to be associated with blue The starchy end osperm comprised most of autofluorescence (Fulcher et al , 1972) . The the weight of the finger millet utricle . testa is strongly fused to the aleurone layer Figures 4A-O and SA-O are micrographs and cannot be easily removed . illustrating the three different starchy endo­ A1 eurone Layer sperm areas {peripheral, corneous, and floury). The a leur one layer was one cell layer This arrangement was similar to that found in thick, surround ed the entire endosperm, and was pearl millet , sorghum, and corn {Rooney et al similar to those seen in corn, sorghum and 1983) . When stained with Acid Fuchsin, the red pearl millet (Fig. 46 ). The cells were small fluorescence decreased in intensity from the (18 x 7.6 ~m) and were packed with aleurone exterior to the inter'ior of the kernel, indicat­ bodies which ranged from 0.9-2.2 J..lffi in diame­ ing that the protein content decreased . ANS ter, similar to values reported by Wada and stained section showed similar results. The Maeda (1980). The cell walls showed intense com pound starch granules increased in size from autofluorescence, which suggested that they con­ the exterior to the interior of the endosperm, tained phenolic acids , probably ferulic acid similar to sorghum (Earp 1984) and pearl millet (Fulcher et al, 1972). Starch granules were (McDonough 1986) . not present. In Fig. 38, the aleurone cell The peripheral endosperm layer wa s uneven wa l l has pulled away from the fifth testa layer in width, ranging from 1-3 cell layers thick. and is not visible. The smallest cells in the endosperm were found

25 1 C.M. McDonough, L.W. Rooney and C.F. Earp

Figure 4: A- Three discrete layers of the starchy endosperm. B - Aleurone cell. C - areas, and short and thick in others. Packed in Peripheral e ndosperm showing s tarch granules be tween the compound gran u 1es were sma 11 er s i m­ and protein bodies. 0 - Closeup at the starch ple granules. The starch granules were 3 . 0- granules in the peripheral endosperm showing 19.0 ~-tm in diameter . There were occasional, the separations between subunits of the starch thin patches of protein matrix present in the granules, protein bodies and protein matrix. corneous endosperm cells (Fig. SC). The starch T =testa, A= aleurone layer, PE =peripheral granules did not have i ndentation s , suggesting endosperm~ CE = corneous endosperm, FE = floury that the corneous endosperm was not as com ­ endosperm, PM = protein matrix, SG = starch pacted as the peripheral area. granule, PB =protein bodies, Y =subuni ts of ln contrast to the highly organized corne­ starch granule, cw =cell wall, i =pits. ous layers, the floury endosperm was a mass of broken cell walls and starch granules with in this layer and were very angular in shape . little semblance of organization. Figure SA The cell contents were tightly packed with a shows the abrupt transition into the floury large number of protein bodies embedded in a endosperm area. The starch granules were muc h thick protein matrix (Fig . 4C,D). The protein smaller in size in the floury area. However, bodies averaged 2 J.Jm in diameter. The starch further inspection revealed that the small gra ­ granules present had many indentations from the nules originated in compound granules that pr o­ protein bodies (Fig . 4C), and most were com­ bably fell apart during cutting and preparation pound (fig . 40). The compound starch granules of the samples. Intact compound starch gra ­ were 8.0-16 . 5 ~m in diameter. An9old (1979) nules ranged from 11-21 ~-tm in diameter, while also reported the presence of compound gra­ the granule fragments averaged 4 ~m. Only a nules. S001e simp l e granules were present in few protein bodies were present and little, if the peripheral endosperm cells. any, protein matrix was seen. The protein was The corneous endosperm comprised the bulk sporadically dispersed on the surface of the of the starchy endosperm. The individual endo­ starch granules . sperm cells broke along cell walls and retained Compound and simple starch granules show a prismatic shape (figs 5A , B) , and contained strong birefringence when viewed with polarized both compound and simple starch granules. The light (Fig. 50). Two compound granules are endosperm cells were long and narrow in some visible, as well as several simple granules of VarlOUS SlZeS.

252 Structural Characteristics of Fi nger Hillet

~~Jufj o~~Y :ndo! ~~ ~~ f ~~~e~~~w~e~ ~~~n~~~~e~~~o - sperm cells with the cell wall partially Germ The germ wa s located in a depression sur­ removed. C - Closeup of the large compound starch granules in the corneous endosperm. rounded by a characteristi c ridge (Figure 6A) SCJile of the smaller starch granules also appear which extended completely around the circumfe­ to have fissures. 0 - Birefringence of rence of the germ. The hi Tum was located imme ­ compound and simple starch granules . C£ = diately adjacent to the germ in a separate but corneous endosperm, F£ = floury endosperm, CW = somewhat shal low er depression. The style was cell wall membrane, SG =starch granule, arrow located on the opposite side of the utricle = protein matrix, C =compound granule, S = from the germ, but does not appear in any of simple granule. the figures. The scutellum cells had a smooth round appearance and were 25.0-35.0 11m in dia­ meter (Fig. 68) . The scutellum was separated from the floury endosperm by the scutellar epi­ thelium; the cells were approximately 1g 11m seed with only a thin testa . The testa layer wide. The protein bodies in the scutellum and of finger millet is unique among the cereal scutellar epithelium were visible as small grains in its structure; it contained five spheres beneath the cell wa lls (Fig . 68). The layers . The surface is covered by a thin layer size of the protein bodies ranged from 1.5-6.0 that autofl uoresces. With SEM , the testa sur ­ 11m in diameter, simi Jar but somewhat higher face has a series of mounds that interlock with than values reported by Wada and Maeda (1980) . each other. The inner layers of the testa contain tannins while the outer layer contains phenolic compounds which autofluoresce . The ce 11 wa 11 s of the endosperm under go strong The structura l characteristics of finger autof 1uorescence. The starchy endosperm millet are quite different from those of pearl contains mostly compound starch granules, but a mil l et (Table 3). The finger millet utricle few simple granules are present, especially in has a thin membranous peri carp composed of seve­ the peripheral area. The testa of finger ral layers of collapsed cells which provides millet adheres strongly to the aleurone layer , little protection from the environment. In con­ and the flour is generally produced by grinding trast, pearl mi 11 et has a peri carp fused to the the .

253 C.M. McDonough, L.W. Rooney and C.F . Earp

Earp CF, Doherty CA, Fulcher RG, Rooney LW. (1983b ) 8-glucans in the caryops is of So rghum bicolor (L.) Moench. Food Microstr~ 183-188. Earp CF. (I984) Microscopy of the mature and developing caryops~ s of (L.) Moench. Ph. D. 01ssertat1on. Texas A&M University, College Station, Texas. Fulcher RG, O'Brien TP, Lee JW. (lg72) Stu- dies on the aleurone layer. I. Conventional and fluorescence microscopy of the cell wall with emphasis on phenol- complexes in wheat. Aust. J. Biol. Sci. 25, 23-24. Fulcher RG, Wong SI. (lg80) Inside cereals- A fluorescence microchemical view. Page 1 in: Cereals for Food s and Beverages. Inglett GE, Munck L, (eds.) Academic Press, New York, NY. Fulcher RG, Wood PJ. (1983) Identification of cerea 1 by fluorescent mi crosco­ py. Page 111 in: New Frontiers in Food Micro­ structure. Bechtel 08, (ed.) AACC, St. Paul, MN. Hahn DH, Faubion JM, Rooney LW. (1g83) Sor­ ghum pheno lic acids, their HPLC separation and their relationship to fungal resistance. Cereal Chern., 60, 255 -25g. Hilu KW, de wet JMJ. (1g67) of Eleusine corocana, Econ. Bot., 30, f9g-208. ~~- - Hilu KW, de Wet JMJ, Harlan JR. (lg7g) Archaeobotanical studies of Eleusine corocana ssp. corocana ( fi ngermTll"et). Am. J. Bot., ~30 -333. Hilu KW, de wet JMJ, Seigler D. (1g78) Flavo­ noid patterns and systemics in Eleusine. Biochem. Syst. Ecol., 5, 247 - 24g,---- Irving DW. {1983) -Anatomy and histochemistry of Echi nocloa turnerama {Channe 1 mi 11 et) s~Cerea 1 Chern. 60, 155-160. ~~~e~~ a: ~h;r~~~t!~~~!~:n~~s~:~m g~~~~rf ~ce. -Kaluza WZ, McGrath RM, Roberts TC, SchOder T =testa, R =ridge, E =embryonic axis, S = HS. (1980) Separation a~ phenols ?f Sorghum scutellum, CE = corneous endosperm, FE = floury bicolor {L.) Moench gra1n. J. Ag n c. Food endosperm, SE = scutellar epithelium. Chern., 28, 11g1.11g6. MaxsonEo, Rooney LW. (1972) Two methods for tannin ana~ysi~ for Sorghul bicolor (L.) Moench gra1n. Crop Sci., 2, 253-256. McDonough CM. (lg86.) Structure of the mature pearl millet ( americanum) Appreciation is expressed to Mr s . Sheryl caryopsis. M. S. Thesis, Texas A&M University, Beavers for her assistance with the chemical College Station, Texas. analyses and to Julie Poe for the HPLC phenol McDonough CM, Beavers S, Rooney LW. (1g83). analyses. This research was partially sup ­ Factors affecting the polyphenol content in ported by the INTSORMIL Title XII Sorghum and cereals. Cereal World, 28, 559. Millet Research Program, which is supported in Mehra KL. (1963) DifferentiaTion of the culti­ part by Grant AI D/DSAN/SI I /G-014g from the vated and wild Eleusine species. Phyton, Agency for International Development, 20(2), 18g-1g8. ~~- Washington, D. C. 20253. -Mura 1i kreshna G, Paramahans SV, Tharanathan RN. (lg82) Carbohydrate make-up of minor mi 1- lets. Starke, 34, 3g7-401. Pararnahans sv-:-wankhede DB, Tharanathan RN . Angold RE. (lg7g) Cereals and bakery {1980) Studies on varagu starch. Starke, products, In: Food Microscopy, Vaughan RG, 32, 1og-112. (ed.), Academic Press, London , 75-138. -Phi 11 ips SM. (lg71) A survey of the Earp CF, Daher ty CA, Rooney LW. (lgB3a) Eleusine Gaertn. (Graminae) in Africa. Kew Fluorescence microscopy of the pericarp, iliJTl:;-21(2), 251-270. a 1eurone layer, and endosperm ce 11 wa 11 s of PoreMS, Magar NG. (1979) composi- three sorghum . Cereal Chern . 60, tion of hybrid varieties of finger millet. 408-412. - Ind. J. Agric. Sci., ~(7), 526-531.

254 Structural Characteristics of Finger Millet

Table 3 Structural Characteristics of Finger and Pearl Millet

Characteri sties Finger Millet Pearl Millet

Type of Seed utricle caryops 1 s Peri carp unattached attached Seedcoat --nllckness 10 . 8-24. 2~m # layers 5 Continuous yes A1 eurone ~yers I I Cell Widt h 7 .6~m 5.0-15.0~m Cell Leng th 18.0~m 16.0-30.0~m Starch Granu les simple 2 simple, compound ~;~: : Peripheral 8.0-16.5~m 6.43~m Corneous 3.0 - 19.0~m 7 .40~m Floury ll.0-21.0~m 7. 60~m Protein Bodies Size 1.9-2.0~m 0.6-0. 7~m Location peripheral, corneous , peri ph era 1, corneous, very few in floury floury areas Protein Matrix loca t1on peri ph era 1, corneous peripheral, corneous, none in floury floury areas Germ --Size 270 x 980~m 620 x 1420~m Endosperm/Germ Rat io3 11: I 2. 5 :I

1Data taken from Mc0onough~1986: Range of starch granule_ sHes tnc lude both simp le and compound granules. 3 Surf ace area approx i rna t 1 on .

Ramachandra G, Virupaksha TK, Wankhede DB, Shehnaz A, Raghavendra Rao MR. Shadaksharaswamy M. (1977) Relationship between {1979) Preparation and physico-chemical proper­ tannin levels and in vitro digestibility in ties of starches and their fractions from fin­ finger millet (EleUSine corocana Gaertn.). ger millet (Eleusine corocana) and foxtail J. Agric. Food ~5TST;l101-1104. millet (Setar:laiTiTl~arke~ 31, Rooney LW, Faubion JM, Earp CF. (1983) 153-159.------Scanning electron microscopy of cereal grains . Zeleznak K, Varriano-Marston E. (1982) Pearl Page 201 in: New Frontiers in Food millet (Penni setum ameri canum [l.] Leeke) Microstructure. Bechtel DB, {ed.) AACC, St. and grain sorghum (Sorghum bicolor [l.] Paul, MN. Moench) ultrastructure. Am. J. Bot., 69 , Technicon Industrial Systems. {1976) 1306-1313. - Individual/simultaneous determination of nitro­ gen and/or phosphorus in BD acid digestions. Discussion with Reviewers Method #334-74A/A. Tarrytown, NY. Techni con Instruments Corporation. (1978) A.W. MacGregor: In the SEM micrographs, how Glucose hexokinase method #SF4-0046 - FA8. can one distinguish small starch granules from Tarrytown , NY. protein bodies? How do the authors know that Tharana than RN, Paramahans SV, Wankhede DB . the small bodies in Fig. 4C are protein bodies? {1980) Amylolytic suscepti bility of native Are the small bodies on the en dosperm si de of groundnut and ragi star ch granules as viewed by the sc ute llar epithet i urn/endosperm junction scann ing electron microscopy. Stiirke, 32, (Fig. 6B} s tarch granules or protein bodies? 158-161. - Authors: We used f1 uorescence mi eros copy to Wada T, Maeda E. (1980) A cytological study help identify the various round bodies found on the phosphorus accumulating tissues in the throughout the endosperm. Fluorochromes speci­ Gramineous seeds . . Jour. Crop Sci., fic for protein helped us locate protein bodies ~(3 ) ' 173-181. in the endosperm and distinguish them from starch granules. Protein bodies are usually

255 C.M. McDonough, L.W. Rooney, and C.F. Earp

round, not angular as seen in the photographs. In Figs. 4C and 68, the small bodies wer e identified as protein by staining a simi lar embedded section of the kernel with protein­ pas it i ve f 1uorochromes.

A.W . MacGregor: How consistent was the testa layering shown in Fig. 3? We re these layers found in all testa sections examined? Figs . 26 and 3A show two different testa sections at simi Jar magnifications, but layers 3 and 4 are not readily apparent in Fig. 28. Authors: The five layer testa was observed ~finger millet utricles examined, includ­ ing the utricle from wh ich Fig. 26 was taken. The spaces were open areas inside the testa layers. They were observed in all utricles. We do not believe that they were artifacts. The spaces distor ted the layers, thus masking their appearance. All layers of the testa were vi sible in areas without spaces.

A.W. MacGregor: Would the authors care to speculate about the formation of the floury area in the endosperm? Is there any possibility that cell wall and protein degrading enzym es have been secreted from the embryo and have degraded this portion of the endosperm? Authors: The grain samples used were not deter1orated. The characteristics of the floury en do sperm s hown in the micrographs were consistent in all utricles.

A. W. MacGregor: In the discussion on testa layers, the authors mention the presence of granules in the open spaces underneath the mounds of the layer . Are these granules vi sible in Fig. 26 in the area marked sp? Is H possible that these are protein bod ies from underlying aleurone cells? Authors: The granules are present in the 1nter1or portions of the space in Fig . 26 . The composition of the granules is unknown, but they were observed i n all of the s paces seen in the samples .

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