Determination of Biochemical Composition of Avena sativa

BANGLADESH RESEARCH PUBLICATIONS JOURNALISSN: 1998-2003, Volume: 4, Issue: 4, Page: 312-319, September - October, 2010DETERMINATION OF BIOCHEMICAL COMPOSITION OF Avenasativa (OAT) AND TO ESTIMATE THE EFFECT OF HIGH FIBER DIET ONHYPERCHOLESTEROLEMIC RATSShumaila Usman 1 , Saima Nazir 1 Sakhawat Ali 1 , Zahida Nasreen *1 and Ammara Najim 2Shumaila Usman, Saima Nazir, Sakhawat Ali, Zahida Nasreen and Ammara Najim. (2010).Determination of Biochemical Composition of Avena sativa (Oat) and to Estimate the Effect of HighFiber Diet on Hypercholesterolemic Rats. Bangladesh Res. Pub. J. 4(4): 312-319. Retrieve fromhttp://www.bdresearchpublications.com/admin/journal/upload/09181/09181.pdfAbstractChemical compositions and nutritive value of oat (Avena sativa) samples(oat grain, hull and oat bran) determined by proximate analysis. Thepercentage value of moisture, ash, protein, fat, fiber, cellulose, lignin andcarbohydrate in oat whole grain, hull and in bran is deliberated. Moreover,evaluation of functional properties of oat, anti-hypercholesterolemic effectof oat bran high fiber diet is studied. The in house prepared oat bran highfiber diet proved effective as it lowered the cholesterol levels significantly inhypercholesterolemic rat.Keywords: Nutritive value, Oat and anti hypercholesterolemic effect.IntroductionOat is one of the most important rabi cereals fodder crop grown in winterthroughout Pakistan both under irrigated and rained conditions. It is a quickgrowing, palatable, succulent and nutritious fodder crop (Nawaz et al 2004). Oatcontains high-unsaturated fatty acid and protein (Wood and Beer, 1998). Theaverage composition of oat is; 15.9% protein, 7.0% fat and 63.2% starch. It grainhas always been an important from of livestock feed. Oat is versatile and grownon many different soil types around the world. It has been shown that oat cantolerate acidic soil with a pH of 4.5, but higher yield require a pH of 5.3 to 5.7(Alam and Adams, 1979).Phenolic compounds with antioxidant activity been identified in oats.Phenolic compounds present in oats, it contributed to functional and nutritionalproperties of the grain (Collins, 1989). It is well known that phenolic acids whichare abundant in whole grains having antioxidant characteristics (Onyeneho andHettiarchchi, 1992).Antioxidants protect the body from degenerative diseases such as cancer(Bailey and Williams, 1993). Phytic acid is the major phosphorus storagecompound of most seeds and cereal grains, it may account for more than 70% ofthe total phosphorus (Rhou and Erdman, 1995).Soluble fiber found in cereals, legumes (including beans peas), some fruits(including apples, pears) and plantain seed husks psyllium (Plantago psyllium). β-glucan is the soluble fiber of cereals, which consist of linear unbranchedpolysaccharides of linked β(1-3)-(1-4)-d-glucopyranose units, that has been shown*Corresponding Author, Email: Zahidanasreen_pcsr@yahoo.com1= Biotechnology and Food Research Centre, PCSIR Labs Complex, Lahore 54600, Pakistan2= Institute of Molecular Biology & Biotechnology, the University of Lahore, Pakistan.

Determination of Biochemical Composition of Avena sativa313to be an important component in cholesterol lowering properties of viscoussoluble fiber (Behall et al 2004).A β-glucan enriched meal thought to increase intestinal viscosity and bileacid binding which in turn may decrease re absorption of bile acids and drive bileacid synthesis from hepatic cholesterol, hence depleting the body ' s cholesterolpool and decrease absorption of intestinal cholesterol (Lia, et al 1995). The abilityof the β-glucan to form highly viscous a solution at low concentrations in thehuman gut is thought to form the basis of it is a health benefits. The primary healthbeneficial component of oat bran is β-glucan; dehulled oats (groats) have a highcontent (3-7%) of linear mixed linkage (1→3) (1→4) - β-D glucan (Wood et al,2003). Oat is effective in lowering the total serum LDL cholesterol due to thepresence of β-glucan in oat (Behall et al 2004).The aim of present work was to find out the effect of oatmeal on plasmalipid profile in rats, fed cholesterol-containing diets. The degree of this positiveinfluence is directly connected to the contents of the bioactive components andthe antioxidant activities of the oatmeal diet.Material and MethodsProcurement and cleaning of raw materialOat whole grain used in this study was purchased from local market. Oatgrain and hull was cleaned manually. After removal of foreign material, weedsand non-grain matters, samples were stored in clean polythene bags until used.Estimation of Moisture, Ash Fat and FiberThe moisture, Ash and fat contents were assessed by using method of(Anonymous, 2005). 3-5g of dried sample was in use in each determination.Estimation of ProteinProtein content of oat and hull sample was estimated by using Kjeldalmethod as reported by AOAC 2005.Estimation of ligninLignin content was estimated by the method of A.S.T.M (1961). 1 gm ofdefatted sample was used for refluxion with 70 mL 1.25% H2SO4 for 2 hours. Refluxsample washed with hot water and then with chloroform. Washed sample treatedwas with 72% H2SO4 for four hours with constant stirring. Sample was filtered (add10 -12 mL distilled water) with whatman filter paper No-1 and ignited it at 550oCfor 4 hours.Estimation of CelluloseCellulose estimated by using the method of Kurschner and Hanak (1930). 1gm of defatted sample was used for refluxion with 15 mL of 80% acetic acid and1.5 mL conc. HNO3 acid for 2 hours. Reflux sample washed with hot water.Washed sample was treated with 72% H2SO4 for four hours with constant stirring.Sample was filtered (add 10 -12 mL distilled water) with whatman filter paper No-1washed with distilled water until it become neutral and finally it washed withalcohol and ignited it at 550oC for 6 hours.Estimation of CarbohydratesThe Carbohydrates were determined by applying following equation.Moister+ Ash+ Fat+ Fiber+ Protein-100 = CHO (Anonymous, 2005)http://www.bdresearchpublications.com/journal/

Usman et al.314I) Development of High Fiber DietThe complete dehulling of oat hull from oat groats carried out. Oat branprepared by grinding of clean oat groat and separating oat bran from oat flourby sieving (Anonymous, 1989).Oat bran was used as fiber source in diet. Aftergrinding of oat bran, fiber was extracted by method of (Anonymous, 2005). Acidhydrolysis of extract was carried out with dilute sulphuric acid. After hydrolysis,fiber was neutralized by NaOH. Sample was washed with the distilled water for thecomplete removal of NaOH.2) Composition of Diet25 g/kg oat bran high fiber diet contained 175g/kg casein protein to fulfilthe protein requirments in animel body. After that 12g/ kg corn oil was added indiet to provide essently fatty acids.Corn strach and sucrose was added in diet assweetness and carbohydrates source. DL- Methionin added in diet for amino acidsource. Vitamin mixture and mineral mixer was added 10g/kg diet and 35g/kgdiet respectively. Oat bran was used as a fiber source and 257g oat bran wasadded in per kg diet (Remi et al 1992). After maually mixing, diet was pelletedwith water and dried in oven and store in air tight bags.Feed intakeFeed intake was determined by the weighing the feed hoppers.Consumption, expressed as grams per rat per day, was obtained by dividing foodintake by the number of rats per cage (Jackson et al 1994).The average feed intake was 13g/day/rat.Functional Properties of oat bran High Fiber DietHigh fiber oat bran diet was used for the determination of functionalproperties of oats by the determination of antihypercholesterolemic effect of highfiber oat bran diet on hypercholesterolemic rat.Experimental Design and Induction of Hypercholesterolemia30 albino rats of either sex weighing between 200 to 300gm were selectedfor the experiment. The rat was divided into 3 groups each group consist of 10 rat.Table 1. Animal groups, their diet and treatmentGroups Animal conditions Treatment1 Normal (Control) Chick starter diet2 Hypercholesterolemic Chick starter diet3 Hypercholesterolemic 25% oat bran high fiber diet.Induction of HypercholesterolemiaThe rat was made Hypercholesterolemic by oral administration of 1% ofcholesterol powder (1gm/kg) for 10 days as prescribed by (Reeves et al 1993).Then, the hypercholesterolemic condition was confirmed by using respectivediagnostic kit at the 11 th day of experiment. After confermation ofhypercholesterolemic condition of rats, the day at which rats were treated with25% oat bran high fiber diet was considered as first day of experiment.Blood collection and analysis1ml blood was collected from coccygeal vein of albino rats. Collectedblood was centrifuged at 3000 rpm for 10 minutes and serum was seprated. Theenzymatic kit was used to assess serum levels using spectrophotometer.http://www.bdresearchpublications.com/journal/

Determination of Biochemical Composition of Avena sativa315Examination of serum cholesterol1973).The cholesterol level was determined by using method of (Richmond,Estimation of cholesterol by enzymatic kit.12 test tube were used, two of them labeled as blank/standard. Remaining10 tubes were labelled as 1,2,3……10 for each sample of rat,s serum from eachhypercholesterolemic group. 1000µl reagent was taken in all the tubes by micropipette. 10µl of stanard solution from kit was added to the tube labeled asstandard and 10µl of serum sample was taken in tubes labeled as 1,2,3……12.Contents of all the tubes were mixed well and then incubated at 37oC for 10minutes. Absorbance of standard and sample was measured against the blank atwavelenth of 546 nm. Similar procedure was used the rest of the samples.Statistical AnalysisFor biochemical properties, mean deviation and starndad deviation wascalculated while for funtional properties ANOVA was applied at 0.05 level ofsignificance (p

Usman et al.316Table 4. Multiple ComparisonsDependentVariableCholesterol level at1 st dayGroupG1Control+chick starter dietG1Control+chick starter dietG2Hypercholesterolemic+chick starter dietG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietGroupG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietG1Control+chick starterdietG3Hypercholesterolemic+Oat bran high fiber dietG1Hypercholesterolemic+chick starter dietMeanDifferenceStd.ErrorSignificant level-41.8000* 2.16282 .000*-42.6000* 2.16282 .000*41.8000* 2.16282 .000*-.8000 2.16282 .714**42.6000* 2.16282 .000*G3Hypercholesterolemic+Oat bran high fiber dietG2Hypercholesterolemic+Oat bran high fiber diet.8000 2.16282 .714**Cholesterol level at14 dayG1Control+chick starter dietG1Control+chick starter dietG2Hypercholesterolemic+chick starter dietG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietG1Control+chick starterdietG3Hypercholesterolemic+Oat bran high fiber dietG1Hypercholesterolemic+chick starter diet-41.2000* 2.15664 .000*-28.0000* 2.15664 .000*41.2000* 2.15664 .000*13.2000* 2.15664 .000*28.0000* 2.15664 .000*G3Hypercholesterolemic+Oat bran high fiber dietG2Hypercholesterolemic+Oat bran high fiber diet-13.2000* 2.15664 .000*Cholestrol level at28 dayG1Control+chick starter dietG1Control+chick starter dietG2Hypercholesterolemic+chick starter dietG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietG2Hypercholesterolemic+chick starter dietG3Hypercholesterolemic+Oat bran high fiber dietG1Control+chick starterdietG3Hypercholesterolemic+Oat bran high fiber dietG1Hypercholesterolemic+chick starter diet-39.5000* 2.23391.000*-6.5000* 2.23391 0.07*39.5000* 2.23391 .000*33.0000* 2.23391 .000*6.5000* 2.23391 0.07*G3Hypercholesterolemic+Oat bran high fiber dietG2Hypercholesterolemic+Oat bran high fiber diet-33.0000* 2.23391 .000**Significant level as p< 0.05**Highly signifacanthttp://www.bdresearchpublications.com/journal/

Determination of Biochemical Composition of Avena sativa317Results and DiscussionOat is distinct among the cereals due to its multifunctional characteristicsand nutrtional profile. Oat provides a potential source of low cost protein withgood nutrional assessment. The use of oat in human diet has also been motivatedby the quality of its fiber components which act as hypoglycemic andhypochloesterolemic agents. Phenolic compounds present in oats maycontribute to functional and nutritional properties of the grain. It is well known thatphenolic acids which are abundant in whole grains have antioxidantcharacteristics.Oat bran has been demonstrated to provide many human healthbenefits such as serum cholesterol lowering effect (Butt et al 2008).Bearing in mind the importance of oat , the current work was planned todetrmine the biochemical composition of oat and to estimate the effect of highfiber diet on hyperchloesterolemic rat. The results of proximate analysis of oatsample (oat whole grain, oat hull and oat bran) and antihypercholeaterolemiceffect of oat bran high fiber diet are given in Table # 2 and 3.Oat has long beenrecognized as a good source of protein and fiber (Peterson, 1992). Oat brancontained the highest amount of total fat as ompared to oat whole grain andoat hull as shown in Table 2. Our finding is in accordance with Grausgruber (2004)study.The percentage of protein in oat bran is higher (15.23%) as compared tothe oat whole grain and oat hull, at the same time, fiber content in oat bran washigher (14.13%)as compared to the oat whole grain. Proten and fiber content inoat whole grain was 13.53% and 12.47% respectively as given in table 2. Similarresult were reported by the Grausgruber (2004). Protein content in oat hull was7.3% while the fiber content was 31.97% as given in table 2.These results are inagreement with the findings of Melicia and Grossmann (2006).Most of the minerals in oat are associated with the bran (Peterson, 1992)The percentage of ash content in oat hull was higher as compared to the oatwhole grain and bran.Ash content in oat whole grain was 3.57% and in oat hullwas 7.3%. These results correlate with the consequence of Melicia and Grossmann(2006) as listed in table 2. Production of oat bran for human food usage hasincreased due to its antihypercholesterolemic.Fibrous feeds have not been used for the non ruminants due to theirdocumented reduction of diet digestibility, especially in young and growinganimals. Nonetheless some type of fible and fiber sources don’t exert suchnegative effects on nutrition digestibility but can have protective effect on guthealth and reduce cardiovascular disease risks (Keogh et al 2003).These protective and physiological effects are dependent on complexmixture of stucture, chemical and physiological properties of dietary fiber (waterholding capacity, viscosity gel formation and bile acid binding). Oat bran, animportant source of water soluble fiber, has been shown to reduce serumcholesterol. Oat bran high fiber diet is effective in lowering the plasma cholesterollevel in hypercholesterolemic rats and cholesterol level decreased significantly asshown as in Table No. 4 which relatively correlates with the results of (Dierzuk,2006).ConclusionIn this study, we were able to show the following facts, the contents of totaldietary fibers and the contents of the protein fractions, antioxidant compounds,and the antioxidant activity in oatmeal is higher than the control diets withoutoatmeal. Oatmeal diet significantly hindered the rise of plasma lipids in rats givenhttp://www.bdresearchpublications.com/journal/

Usman et al.318added cholesterol. The present study demonstrated that intake of oat bran highfiber diet is effective in lowering plasma cholesterol levels in hypercholesterolemicrats.ReferencesA.S.T.M. Method, (1961). American Society for Testing Materials and standardmethod of lignin in wood. 1106-56T pages 848. 71-79.Alam, S.M and Adams. WA (1979). Effect of soil pH on the growth and mineralcontent of oats. Pakistan Journal Science and Indstrial Research, 22:147-151.Anderson, J. W. Storey, L and Sieling, B. (1984). Hypercholestrolemic effects of oatbran or bean intake for Hypercholestrolemic men. J Am Cli Nutr, 40: 1146.Anonymous. (1989). American Association of Cereal Chemists (AACC).Committeeadopts oat bran def. Cereal Foods World, 34: 1033.Anonymous. (2005). Association of official Analytcal Chemists (AOAC): Officialmethod of analysis 18 th Ed. Washington D.C.Bailey, G.S and Williams. D.E. (1993). Potential mechanisms for food relatedcarcinogens and anticarcenogens. Food Technology, 47:105-118.Behall, K.M., Scholfield, D.J and Hallfrish, J. (2004).Diets containing barleysignificantly reduces lipids in mildly hypercholesterolemic men and women.American Journal Clinical Nutrtion, 80:1185-1193.Butt, M.S., Nadeem, M.T., Iqbal, M.K and Rabia, S. (2008). Oat unique among thecereals. European Journal of Nutrtion, 47: 68-79.Collins, F.W. (1989). Oat phenolics: Avenanthramides, novel substituted Ncinnamoylanthranilate alkaloids from oat groat and hulls, Journal ofAgricultural and Food Chemistry, 37: 60-66.Dierzuk, F. (2006). The effect of cereal by product dietary fiber on the serum lipidprofile and short chain fatty acid concentration in caecal digesta of rats.Journal of Animal Food Science, 1:57-60.Grausgruber, H., Judith, S., Regine, S and Peter, R. (2004). Variability in chemicalcomposition and biological active consitituents of cereals, GenaticVariation for Plant Breeding, 23-26.Jackson, K.A., Suter, D.A.I and Torring, D.L. (1994). Oat bran, barley and maltedbarley lower plasma cholesterol relative to wheat bran but differ in theireffect on cholesterol in rat feed diet with and without cholesterol. Journal.Nutrition, 1678-1684.Keogh, G.F., Cooper, G.J.S and Mulvey, T.B. (2003). Randomized controlledcrossover study of the effect of a highly beta glucan enriched barley oncardiovascular disease risk factors in mildly hypercholesterolemic men.American. Journal of Clinical Society of Nutrition,78:711-718.Kurschner, K and Hanak, A. (1930). Determination of cellulose: Z. utersuch.Lebensm, 59: 448-485.Lia, A., Hallmans, A.S., Sandberge, B., Aman, P. and Andersson. (1995). Oat betaglucan increases bile acid excretion and a fiber rich barley fractionincreases cholesterol excretion in ileostomy subjects. American Journal ofClinical. Nutrition, 62: 1245-1251.http://www.bdresearchpublications.com/journal/

Determination of Biochemical Composition of Avena sativa319Melicia, C.G and Grossmann, M.V.E. (2006). Oats hulls treated with alkalinehydrogen peroxide associated with extrusion as fiber source in cookies.Journal of Cienc. Tecnol. Aliment.Caminas, 26 (1),123-126.Nawaz, N., Razzaq, A., Ali, Z and Yousuf. (2004). Performance of different oat(Avena sativa L.) varieties unnder the agro-climatic conditions ofBahawalpur- Pakistan Journal of Agricultural and Biology, 1560-8530 : 824-626.Onyeneho, S.N and Hettiarachchy, N.S. (1992) Antioxidant activity of durumwheat bran. Journal Agricultural Food Chemistry, 49: 1496-1500.Peterson, D.M. (1992). Compositon and nutritional characteristics of oat grain andproducts. Marshall HG and ME Sorells (ed). Oat science and technology.American Technical Society Agricultural Crop Science of America,Madison, WI, USA, 265-292.Reeves, P.G., Nielsen and Fahey, G.C. (1993). AlN-93 Purified diet for laboratoryrodent: finally report of the American Institute of Nutrition AD Hoe writingCommittee on reformulation of the AIN-76 rodent diet. Journal Nutrition, 23:1939-1951.Remi, D.S., Dirk, F and Annick, V. (1992). Cholesterol lowering effects and utilizationof protein lipid, fiber and energy in rats fed unprocessed and baked oatbran. Journal. Nutrition. 22: 1318-1324.Rhou, J.R and Erdman, J.V (1995). Phytic acid in health and disease. CRC CriticalReviews in Food Science and Nutrition, 35:495-508.Richmond, W. (1973). Cholesterol enzymatic, lipid, colourimetric test- CHOD/PAPmethod Clinical Chemistry, 19: 1350.Steel, R.G.D and Torrie, J.H. (1982). Principles and procedures of statisties 2 ndedition, McGraw, Hill Book, New york (USA).Wood, P., Weisz, M.U., Beer, C.W, Newman and Newman R.K. (2003). Structure of(1→3) (1→4) – β-D-glucan in waxy and non waxy barley. Cereal Chemistry,80: 329-332.Wood, P.J and Beer. (1998). "Functional oat product" in G Mazza, Functional Food,Biochemical and processing aspects, Technomic Publishing Lancaster,Phennsylvania. Ppl-37. ISBN: 1-56676-487-4.http://www.bdresearchpublications.com/journal/