Background. Nutritional qualities of Bitter Melons were examined by chemical analyses viz. 28 minerals, polyphenolics, protein and aminic acids. Analysiss for antioxidants and other 29 biochemical properties every bit good as cookery qualities are besides investigated. Several formulas have 30 been tasted for consumer credence. Along with the possible chemo preventive activities, the 31 popular belief of acrimonious melon bettering glucose tolerance in Type II diabetes and take downing 32 blood cholesterin are being investigated. However, it has non been determined if or which 33 alkaloids, polypeptides, or combination of the chemicals, found in acrimonious melon, are responsible 34 for the good medicative consequence. The functional compounds contained in these nutrients and their 35 medicative effects are needed to thoroughly studied and clinically proven. 36

Methods. Four assortments of acrimonious melon: Indian Green ( IG ) , Indian White ( IW ) , Chinese Green 38 ( CG ) and Chinese White ( CW ) , were used for phytochemical analyses for protein extraction and 39 finding, protein hydrolysis, finding of amino acid, antioxidant and atimutagenic 40 activity. All the analyses were done as per the criterion methods. All values are reported as 41 agencies of three findings. Split secret plan complete randomized design was conducted utilizing JMP 42 5 package bundle ( SAS ) and Tukey HSD process was performed for the significance of 43 differences at the 5 % degree. . 44

Consequences. Melon flesh contained about 93 % wet for all four assortments, whereas the 46 wet content of melon seeds ranged from 53.3 % in Indian Green to 75 % in Indian White. 47 Bitter melon flesh contained 8.4 % to 9.8 % protein, whereas seed contained 27 % to 31 % protein. 48 Glycine was besides higher in acrimonious melon compared to soy proteins. Other aminic acids contents 49

were similar degrees as flesh proteins. . Phenolic contents of seed, SCT, and flesh ranged from 50 4.67-8.02, 4.64-8.94, and 5.36-8.90 mg/CAE dry affair, severally. Phenolic contents of the 51 flesh were significantly higher than those of the SCT and seed and phenolic contents of the seed 52 was the lowest among those of all the tissues. The entire phenolic contents of four assortments were 53 significantly different with the highest was Indian white followed by China white, China Green, 54 and Indian viridity. The antioxidant activities of Indian viridity, Indian white, China green and China 55 white ranged from 79-88, 79-87, 80-86, and 79-87 % suppression, severally. The antioxidant fifty-six activities of the oven-dried samples and the lyophilized samples were 79-88 and 79-86 % 57 suppression, severally. Bitter melon assortments IW and CG showed higher antimutagenic effects 58 against benzo ( a ) pyrene with Salmonella TA98 ( 92-100 % suppression ) and Salmonella TA100 ( 79-59 86 % suppression ) but lower antimutagenic effects against Na azide. 60

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61

Decision. Bitter melon is a good beginning of phenolic compounds. The phenolic extracts 62 showed high suppression consequence to Prevent lipid oxidization. These natural works phenoplasts can be a 63 good antioxidant which may be applied in many nutrient systems to keep the nutrient quality. 64

65

Background 66

Many alien veggies are known for their particular nutritionary and medicative belongingss. Bitter 67 Melon ( Momordica charantia L. ) , an one-year veggie of Cucurbitaceae household, is found to be 68 one of the of import veggies of particular nutritionary and medicative qualities in southern United 69 States. Germplasm lines and land races of Bitter Melon were evaluated since 2000 for their 70 adaptability in Southeast Arkansas. Four adaptable lines/varieties were tested in replicated field 71 tests for productiveness at the University of Arkansas at Pine Bluff Agricultural Research Center. 72

Melons were harvested at their marketable phases get downing in June and stoping in September for 73 output appraisal. Nutritional qualities of Bitter Melons were examined by chemical analyses 74 viz. vitamin, minerals, polyphenolics, protein and aminic acids. Analysiss for antioxidants and 75 other biochemical properties every bit good as cookery qualities are besides investigated. Several formulas 76 have been tasted for consumer credence. Nutrition-related wellness jobs such as 77 high blood pressure, diabetes, ageing, fleshiness, arthritis, cardiovascular disease, are prevailing among 78 deprived rural and urban populations, particularly minorities in the delta part [ 1, 2 ] . Food 79 ingestion wonts, dietetic consumptions, and meal readying methods are believed to lend to 80 these jobs. Human existences are of course adapted to eating nutrients which have a balanced ratio 81 of triglycerides and phospholipids. Based on the overall balance of triglyceride/phospholipid 82 ratio, nutrients from works beginnings have less negative effects on human wellness than nutrients from 83 carnal beginnings [ 3 ] . When these balanced nutrients are consumed, peroxidation jobs of 84 polysaturated fats and lipoids, associated with many of the human illnesss, are unusually 85 minimized [ 4 ] . In add-on to the normally used native vegetable and herbs in the US, many 86 alien herbs and veggies are known for their particular nutritionary and medicative belongingss [ 5 ] . 87 During the past decennary or two, Americans have realized that they could command their ain wellness 88 jobs and cut down hazards of many chronic unwellnesss by altering certain dietetic behaviours [ 6 ] . 89 The popular belief of acrimonious melon to better glucose tolerance in Type II diabetes and lower 90 blood cholesterin are being investigated. It is still to be determined if the chemical components 91 such as certain alkaloids and polypeptides found in acrimonious melons are effectual separately or in 92 combination. Along with the possible chemo preventive activities, the popular belief of acrimonious 93 melon bettering glucose tolerance in Type II diabetes and take downing blood cholesterin are being 94 investigated. However, it has non been determined if or which alkaloids, polypeptides, or 95

combination of the chemicals, found in acrimonious melon, are responsible for the good medicative 96 consequence. 97

98

Introduction 99

100

Certain veggies contain an copiousness of polyphenolics, terpenoids, isoflavones, 101 anthocyanins, aminic acids, minerals, vitamins and other antioxidants that are associated with 102 protection from malignant neoplastic disease, aging, cardiovascular diseases, diabetes, high blood pressure [ 4,7-12 ] . The most 103 popular and utile group of the veggies belong to the household Cucurbitaceae [ 13 ] . One of the 104 members of this household is Acrimonious Melon ( Momordica charantia L. ) , besides known as Karela, balsam 105 pear, or foo gwa, an one-year fruity veggie of the cucurbitaceae household. It is a daintiness to the 106 East and Southeast Asiatic people, particularly the people of India, China, Japan, Taiwan, 107 Bangladesh, Pakistan, Thailand, Malaysia, Indonesia, Philippines, Nepal, Bhutan and Sri Lanka. 108 Bitter melon is besides a popular veggie in West Indies, Brazil, Colombia, Cuba, Mexico, 109 Panama, Peru, and some African and European states. The acrimonious quality, for which the fruit is 110 named, is due to the alkaloid momordicine. The green warty fruits of this works are used as a 111 veggie, rich in vitamin A, vitamin C, and Fe [ 14 ] . Fruits vary in size ( 1.0 ” -9.8 ” long and 1.0-112 5.9 ” broad ) , form ( egg-shaped, unit of ammunition, oblong, nine, etc. ) , fruit colour ( dark viridity to creamy white ) , 113 adulthood ( 45-80 yearss ) , fruit per works ( 6-85 ) and yield per works ( 0.8-12 Ibs ) . The fruits, foliages, 114 and roots of acrimonious melon are traditionally believed to hold medicative value in cut downing blood 115 sugar degrees for diabetic patients [ 15,16 ] . The assorted medicative belongingss of acrimonious melon are 116 good known in eastern Asia [ 17 ] . Sofowora [ 18 ] reported several utilizations of M. charantia in 117 traditional medical specialties in Africa. Pharmacologically, the hypoglycaemic belongingss of the works 118

variety meats were established by Lotlikar et Al. [ 19 ] . The polypeptides from the seeds and fruits of M. 119 charantia were considered as antidiabetic agents [ 20-24 ] . Cucurbitacines isolated from several 120 species of the household Cucurbitaceae showed anticancer effects [ 25-27 ] 7. Lin et Al. [ 28 ] isolated 121 two lectins from seeds of M. charantia, momordin and agglutinin, the momordin suppressing 122 protein synthesis by Ehrlich Ascites cells. Licastro et Al. [ 29 ] announced that two proteins 123 isolated from the seeds of M. charantia inhibited protein synthesis and subsequent Deoxyribonucleic acid 124 synthesis in normal and leukaemic human peripheral blood lymph cells. Spreafico et Al. [ 30 ] 125 reported a protein inhibitor from M. charantia demoing immunomodulatory activity in mice. 126 Although acrimonious melon is used by the cultural people from Asia, Africa and Europe, American 127 consumers have non widely developed a gustatory sensation and grasp for it. The harvest could be 128 introduced in the US as an extra nutrient point possessing particular medicative belongingss. 129 Therefore the functional compounds contained in these nutrients and their medicative effects are 130 needed to thoroughly studied and clinically proven. Research is required to place suited 131 genotypes of this veggie for their production potency and nutritionary and/or nutraceutical 132 values. In these on-going surveies, we have determined that acrimonious melon can be successfully 133 produced in Arkansas and the southern provinces. 134

135

Methods 136

137

Four selected assortments of acrimonious melon were tested in replicated field tests in 2003-2004 on 138 Calloway silt loam dirt at the UAPB Agricultural Research Center in Pine Bluff. Tests were 139 conducted in randomised complete block designs with four reproductions. Plants were grown on 140 unsloped treillages with 10 ft. between rows and 12 ft. within rows. Two four-week old seedlings 141

were transplanted in each hole. N, P, and K fertilisers were applied pre-plant utilizing 200 lbs per 142 acre of 13-13-13. Irrigation was provided by trickle irrigation system. Weedss were controlled by 143 covering the beds with black plastic. Roundup ( Glyphosate ) was used in commanding weeds 144 around the beds. Melons were harvested twice a hebdomad get downing in mid-June and stoping mid-145 September. Fruit samples were stored in certain plastic bags in ice thoraxs for phytochemical 146 analysis. 147

148

Bitter Melon Sample Preparation. The acrimonious melons were washed with deionized H2O, 149 drained at ambient temperature and cut in half lengthwise. The seeds were removed and flesh 150 was thinly sliced and collected individually from the seeds. Flesh was divided into two halves. One 151 half was freeze dried and the other half was oven dried. Seeds and flesh was oven dried at 80 OC 152 for 3 yearss. Dried flesh and seeds were so land to flour and passed through a 60-mesh U.S.A. 153 criterion proving screen ( W.S. Tyler, Incorporated, Mentor, OH ) . 154

155

Protein Extraction and Determination. Bitter melon protein isolates were prepared following a 156 standard process for soy protein isolation [ 31 ] . Ten gms of acrimonious melon flour were extracted 157 with 100 milliliters of H2O at pH 8.0 by stirring at ambient temperature for 2 h. The suspension was 158 so centrifuged at 1500 g for 30 min at 10 OC to divide the supernatant from the solid stage. 159 The supernatant was so adjusted to pH 4.5, held at 4 OC for 2 H and centrifuged once more at 1500 g 160 for 30 min at 10 OC. The ensuing precipitate was dissolved in 20 milliliters deionized H2O, adjusted 161 to pH 8.0 and lyophilized. Soy proteins were isolated utilizing the same process. Protein content 162 of acrimonious melon flour and its protein isolates were determined utilizing a Kjeldahl Unit ( Kjeltec 163 Analyzer Unit, Foss Tecator AB, Hoganas, Sweden ) . Approximately 50 milligrams of each sample was 164

digested with concentrated sulphuric acid in the presence of 1 Kjeltab Unit with 0.1 N HCl. Kjet-165 Sorb was used as the receiving system during titration. Protein contents were calculated utilizing a factor of 166 6.25. 167

168

Protein Hydrolysis. Protein hydrolysis was done utilizing a protocol described by Eveleigh and 169 Winter [ 32 ] . Ten milligram of proteins were dispersed in 5 milliliter of 6 N HCl. Two hundred microliters of 170 the suspension were taken into a vacuity hydrolysis tubing ( Pierce Chemical Co. , Rockford, IL ) . 171 The tubing was connected to a vacuity pump for 5 min. The tubing was so sealed and placed in a 172 Reacti Heating Module ( Pierce Chemical Company ) at 150 OC overnight ( 12 H ) . After dialysis, 173 the tubing was cooled to room temperature before the aliquot was transferred to a 2.5 milliliter amber 174 phial. The hydrolyzate was so evaporated to dryness under a watercourse of N at 60 OC. To the 175 dried amino acids was so added 0.5 mL sample thining buffer ( Beckman Coulter Inc. , 176 Fullerton, CA ) and the aliquot was filtered through a 0.2 Aµm filter prior to amino acerb analysis. 177

178

Amino Acid Composition and Analysis. Amino acerb composing was analyzed utilizing an 179 Amino Acid Analyzer 126 Beckman HPLC system with a station column derivatization reactor 180 ( Beckman Instruments, Inc. Palo Alto, CA ) . Bitter melon flour and protein isolates were 181 hydrolyzed with 6 M HCl to bring forth aminic acids and so analyzed with the Amino Acid 182 Analyzer. Chromatographic equipment consisted of a Beckman liquid chromatograph theoretical account 126 183 HPLC equipped with a System Gold Nouveau package ( Beckman Instruments, Inc, Palo Alto, 184 CA ) was used. The optical density of the wastewater was monitored at 570 nanometer. The nomadic stage 185 consisted of Na buffer Na-E ( pH 3.3 ) , Na-F ( pH 4.3 ) and Na-D ( pH 6.3 ) ( Beckman Coulter 186 Inc. , Fullerton, CA ) . A Hi-performance regeneration solution ( Na-R ) ( Beckman Coulter Inc. , 187

Fullerton, CA ) was used to renew the column after each tally. Flow rate was set at 0.44 188 mL/min and column temperature was set an initial temperature of 50 OC for 7 min and 75 OC for 189 the remainder of the trial. The initial solvent status was buffer Na-E. After 19.5 min the buffer was 190 switched to Na-F and at 30 min the buffer was switched to Na-D until 65 min. Then the column 191 was washed with solvent Na-R for 3 min before the dissolver was brought to the original 192 status. The flow rate of the ninhydrin ( Nin-RX, Beckman Coulter Inc. , Fullerton, CA ) was 193 set at 0.23 ml/min. A sample size of 20-100 L was injected during HPLC analysis. The amino 194 acerb composing was calculated from standard curves calibrated utilizing the 17 criterion amino 195 acids. 196

197

Determination of Phenolics. Entire phenoplasts of the all right calabash flesh, SCT, and seed of acrimonious 198 melons were determined by Folin-Ciocalteu method. One hundred mgs of each sample 199 was weighted into a screw-cap trial tubing and vortexed with 10 milliliters of methyl alcohol. The scattering 200 was heated in a H2O bath of 65 OC for 2h and allowed to chill at room temperature. To 1 milliliter of 201 the clear solution in a screw-cap trial tubing, 1.0 deionized H2O was added. The tubings were 202 vortexed and allowed to stand for 2h at room temperature. Absorption of the solution at 726 nm 203 was measured utilizing a spectrophotometer. The entire phenolic content was expressed as 204 chlorogenic acid equivalents in mg/g dry stuff. The phenolic acid components were analyzed 205 by HPLC [ 33 ] . 206

207

Antioxidant activity finding. Antioxidant activity was carried out by oxidising linoleic 208 acerb methyl ester ( MeLo ) in the presence of phenolic infusions as antioxidants [ 34 ] . Two milligram of 209 the infusions were dissolved in 10 milliliter of methyl alcohol. Five 100 microliters of the infusions 210

solution were added into 0.2 g of MeLo ( 500 ppm infusion in MeLo ) , and the methyl alcohol was 211 evaporated under a watercourse of N at ambient temperature. Five 100 microliters of 212 methyl alcohols were added into 0.2 g of MeLo for space as a mention. Oxidation of MeLo in the 213 nowadays of infusion was carried out in at 40 C for 72h. Two milligram of sample aliquots were taken at 214 the starting point ( zero clip ) and after 72h of oxidization ( at 40 C ) and dissolved in 10 milliliter of 215 2,2,4-trimethylpentane ( isooctane ) . The conjugated diene soaking up of the aliquots was read 216 utilizing a spectrophotometer ( Shimadzu Model UV-1601, Kyoto, Japan ) at a moving ridge length of 234 217 nanometer. The antioxidant activities were expressed as per centum suppression of conjugated diene 218 hydroperoxides formation of MeLo after 72h of oxidization comparing with space from MeLo as a 219 mention antioxidant as follows: % inhibition= [ ( AB ( 72h ) – AB ( 0h ) ) – ( AE ( 72h ) – AE ( 0h ) ) / 220 ( AB ( 72h ) – AB ( 0h ) ) ] x 100 ; where A= optical density, E= infusion, and B= space. 221

222

Antimutagenic activity finding. The antimutagenic activity of the methanolic infusion 223 from lyophilized bitter melon flesh and seed from assortments India white and China viridity was 224 determined by the method of Ames et Al. [ 35 ] . The histidine necessitating discolorations of Salmonella 225 typhimurium TA98 and TA 100 were used for this trial. Benzo ( a ) pyrene and Na azide were 226 used as mutagens for mutagenic and antimutagenic trials. The per centum suppression of 227 mutagenesis was calculated utilizing the undermentioned equation: suppression % = [ 1- ( figure of revertants 228 in the presence of fraction/number of revertants in the absence of fraction ) ] X 100. 229

230

Statistical analysis. All values are reported as agencies of three findings. Split secret plan 231 complete randomized design was conducted utilizing JMP 5 package bundle ( SAS 2002 ) and 232 Tukey HSD process was performed for the significance of differences at the 5 % degree. . 233

234

Consequences and Discussion 235

236

Nutritive, Protein and Moisture Content of Bitter Melon. The acrimonious melon fruit has higher 237 content of vitamin C, K, Ca, Mg, and dietetic fibre as comparison to some 238 other commercial veggies [ 14 ] ( Table 1 ) . It besides has niacin, vitamin B1, vitamin B2, vitamin A, 239 protein, organic acids and other foods. Physical separation of acrimonious melons resulted in flesh 240 and seed fractions. The protein and wet content of flesh and seeds of four assortments of acrimonious 241 melons are given in Table 2. Melon flesh contained about 93 % wet for all four 242 assortments, whereas the wet content of melon seeds ranged from 53.3 % in Indian Green to 243 75 % in Indian White. Bitter melon flesh contained 8.4 % to 9.8 % protein, whereas seed contained 244 27 % to 31 % protein. 245

246

Amino Acid Composition of Bitter Melon. Freeze dried melon flesh was high in lysine ( mole 247 per centum ) compared to soy protein isolate. Flesh was comparative lower in glutamic acid and 248 arginine ( Table 3 ) . Essential amino acids, including threonine, valine, methionine, isoleucine, 249 leucine, and phenylalanine are comparable in sum to soy proteins and other legume proteins. 250 On the other manus, oven dried flesh had a much lower per centum of lysine and a lower 251 per centum of arginine while other aminic acids were about similar. Amino acerb composings 252 of seeds are given in Table 4. Seed protein was higher in glutamic acid and arginine but lower in 253 lysine compared to flesh proteins. Glycine was besides higher in acrimonious melon compared to soy 254 proteins. Other aminic acids contents were similar degrees as flesh proteins. 255

256

Quality Criteria. The quality standards of acrimonious melon are fresh visual aspect with unvarying 257 colour and house without inordinate seed development. The acrimonious melons are harvested, 258 selected for size and uniformity of fruit surface features and normally packed in carton or 259 wood boxes incorporating 5, 10, or 20 kilogram of fruit. It is a chilling sensitive veggie, and may be 260 air-cooled to 10 OC-12 OC [ 36, 37 ] . Acrimonious melons are intermediate in perishableness. The common 261 postharvest defects are seed development, softening and maturing with internal /external colour 262 alteration. The recommended temperature and comparative humidness for postharvest handling of acrimonious 263 melon are 10-12.5 OC and 85-90 % , severally, with estimated shelf-life 7 to 14 yearss [ 36, 37 ] . 264

265

Phenolic contents. Entire phenolic content of oven and lyophilized bitter melon tissues from 266 four assortments are given in Table 5. Overall, phenolic content in oven-dried samples were 267 significantly higher than lyophilized samples. Phenolic contents of the oven dried and freeze-268 dried tissues ranged from 5.39-8.94 milligram of chlorogenic acid equivalent ( CAE ) /g dry affair and 269 4.64-8.90 mg/CAE.g dry affair, severally. Phenolic contents of seed, SCT, and flesh ranged 270 from 4.67-8.02, 4.64-8.94, and 5.36-8.90 mg/CAE dry affair, severally. Phenolic contents of 271 the flesh were significantly higher than those of the SCT and seed and phenolic contents of the 272 seed was the lowest among those of all the tissues. The entire phenolic contents of four assortments 273 were significantly different with the highest was Indian white followed by China white, China 274 Green, and Indian viridity. 275

276

Antioxidant activities of acrimonious melon infusion. There was no important difference in the 277 antioxidant activities ( % suppression ) of the methanolic infusions from acrimonious melons among 278 assortments and drying methods ( oven and lyophilized ) ( Table 6 ) . The antioxidant activities of 279

Indian viridity, Indian white, China green and China white ranged from 79-88, 79-87, 80-86, and 280 79-87 % suppression, severally. The antioxidant activities of the oven-dried samples and the 281 lyophilized samples were 79-88 and 79-86 % suppression, severally. The antioxidant activities 282 of the methanolic infusions of flesh and SCT were non significantly different, while they were 283 significantly higher than that of seeds. Shu-Jing and Lean-Teik [ 38 ] reported that acrimonious melon 284 infusions possess powerful antioxidant and free extremist scavenging activities. These antioxidant 285 activities could hold contributed, at least partially, to the curative benefits of the certain 286 traditional claims of wild BM. 287

288

Antimutagenicity of acrimonious melon infusion. Bitter melon assortments IW and CG showed higher 289 antimutagenic effects against benzo ( a ) pyrene with Salmonella TA98 ( 92-100 % suppression ) and 290 Salmonella TA100 ( 79-86 % suppression ) ( Table 7 ) but lower antimutagenic effects against Na 291 azide ( informations non shown ) . Similar determination was reported by several writers [ 39, 40 ] . Wattenberg 292 [ 41 ] discussed chemopreventive agents as barricading agents that prevent carcinogens from 293 making or responding with critical mark sites, and as stamp downing agents that prevent development of 294 the neoplastic procedure in cells that otherwise would go malignant. Bitter melon extract 295 contains both agents [ 40 ] . Although the exact mechanism of the chemopreventive effects of 296 acrimonious melon is non yet known, these findings suggest that acrimonious melon is a possible 297 chemopreventive agent against carcinogenesis. 298

299

Decisions 300

Bitter melon is an first-class beginning of phenolic compounds, antioxidant and antimutagen. This 301 can happen application in nutrient merchandises, and dietetic addendums. The phenolic infusions showed 302

high suppression consequence to Prevent lipid oxidization. These natural works phenoplasts can be a good 303 antioxidant which may be applied in many nutrient systems to keep the nutrient quality. More 304 Detailed probes on acrimonious Melon phenoplasts, peptides, and proteins are needed to supply 305 information for their nutraceutical values. Bitter melon has high demand in the cultural market 306 particularly in Asian, African and certain South American states. To the best of our cognition 307 in normal vegetable use of acrimonious melon no toxicity has been reported. However high dosage of 308 acrimonious melon capsule which is now available in the US every bit good as international market may do 309 some sort of toxic consequence, like over dosage of any other polyphenolics it is reported in some non 310 referred beginnings that higher dosage may do abortion. More Detailed probes on acrimonious 311 Melon phenoplasts, peptides, and proteins are needed to supply information for their nutraceutical 312 values. 313

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