lathyrism ethiopia tekle-haimanot et al nutr res 1993

7/29/2019 Lathyrism Ethiopia Tekle-Haimanot Et Al Nutr Res 1993

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u T, Zein AZ, Spencer PS.leurotoxic disorder. Int J Epid. Addis Ababa: Commercial)r the determination of a leurotoxin. Anal Biochem^Oxalvlamino-L-alanine, theie d ) -rass-pea: Threat andtion, 1989:128-132.^-aminopropionic acid, and>;15:1257-1259.alyl-L-a, -diaminopropioniclytochemistry 1977,16:1211-xicities of a- and -N-oxalyl-ett 1985;55:89-94.

- oxalyl derivatives ofneurotoxin in this and otherH, Yeshanew A. Grass Pea:thiopia. (A paper presentedlgladesh, 30 November- 3es. Central Statistic Office,new dimensions. Nutrition

oundation of Ind ia. 1984:19-

ITRITION RES EARC H, Vol. 13, pp. 1113-1126,19 9371-5317/93 $6.00 + .00 Printed in the USA.pyright (c) 1993 Pergamon Press Ltd. All rights reserved.

PATTERN OF LATHYRUS SATIVUS (GRASS PEA) CONSUMPTIONAND BETA-Af-OXALYL-a--DIAMINOPROPRIONIC ACID (-ODAP )CONTENT OF FOOD SAMPLES IN THE LATHYRISM ENDEMICREGION OF NORTHWEST ETHIOPIARedda Tekle-Haimanot*, Berhanu M. Abeg az", Elizabeth W uhib*",Angelina Kassina*", Yemane Kidane*", Naod Kebede",Tadesse Alemu* and Peter S. Spencer*"*

Faculty of Medicine, Addis Ababa University, P.O.Box, 4147,Addis Ababa, Ethiopia*: Faculty of Science, Addis AbabaUniversity, P.O.Box, 1176, Addis Ab aba "; Ethiopian NutritionInstitute, Addis Ababa"; and Oregon Health Sciences University,Portland, Oregon 97201, USA"**.

ABSTRACTGrass pea , a legume whose consum ption is associated w ith human lathyrism, isan important food crop among the poorer sectors of society in many parts ofEthiopia. A nutritional survey, focussing on the preparation and consumption ofgrass pea, in 224 families of two villages of a lathyrism endemic area in


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1114 R.TEKLE-HAIMANOTetal. LATHYRISMEpidemics of the disease have been reported following acute food shortages duringdrought or after floods (3-5). Recent studies have confirmed that -N-oxalyl-L-a, -diaminopropionic acid (-ODA P), [synonym : -N-oxalylamino-L-alanine (BOAA)] is theneurotoxic principle in L sativus responsible for the neurodegenerativo condition (6).Although lathyrism has in the past been reported from other p arts of the world , includingEurope, it is at present known to be endemic in India (7), Bangladesh (8) and Ethiopia(9). In Ethiopia, the disease occurs in the northern and central parts of the country withrecurrent outbreaks reported from the Lake Tana basin of the north-west AdministrativeRegion of Gondar.As part of a multidiscipilinary project for the improvement of Lathyrus sativus and thecontrol of lathyrism, we undertook an epidemiolqgical survey of the Dembia and Fogerasubdistricts of the Lake Tana basin (10). A nutritional survey was also carried out in twopeasant associations within the project area to find out the feeding habits of theinhabitants particularly in relation to the preparation an d consumption of grass pea. Foodsamples were collected and assayed for (-ODAP) levels. This paper reports on ourfindings.

MATERIALS AND METHODSIn the subdistrict of Fogera of the Gondar Administrative Region, two peasantassociations were selected for the nutritional survey. Shina (pop. 6,142) is a village in agrass pea cultivating area, while Montura (pop. 2,560) situated 50 km north of Shina isa non-grass pea growing village. Both villages are about 600 km north of Addis Ababa.In Shina, most of the farmland invariably gets flooded during and after the heavy rainymonths of July to September. Farmers therefore resort to the cultivation of the hardy crop,grass pea. On the other hand, Montura which is situated on the highlands, has a morefertile and productive land where cereals like teff (Eragrotis teff), wheat, barley and milletare cultivated.For the nutritional survey of the two villages, a questionnaire was developed to get thefollowing essential information: supply and preparation of food, distribution andconsumption of food and infant feeding, with particular emphasis on grass pea. Thequestionnaire was carefully designed to obtain the comparative dietary pattern of thetarget populations of the two villages.The questionnaire, initially prepared in English, was translated into Amharic, the locallanguage spoken in the region. Based on pilot studies, the questionnaire was furthermodified to formulate the questions in accordance with the cultural background andeducational levels of the population.Ten female research assistants were recruited from among the secondary schoolgraduates of Woreta, a town situated between the two villages. These enumerators weretrained for twoweeks by nutritionists of the research team on the concept and techniquesof dietary field survey. As the enumerators w ere recruited locally, a fairly reliable rapportand confidence was developed between them and the respondents. As females, they

were readily accepted by housewives into the kitchens. The activities of the enumeiwere supervised closely by the three experienced nutritionists of the research ((March-April 1989).The purpose of the nutritional study was explained and discussed with the elder;local leaders of the peasant associations. After obtaining the consent and support


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1116 R. TEKLE-HAIMANOT et a l. LATHYRISM 11diluted one hundred fold prior to analysis.Absorbances were measured on a Beckman model 24 spectrophotometer.Diaminopropionic acid hydrochloride (DAP.HCI) and o/tftophthalaldehyde (99% and 97%respectively) were purchased from Aldrich; mercaptoethane was acquired from Hopkinand Williams. Water was redistilled from an all-glass set-up. Ethanol (60% aq.) was usedfor extraction.Preparation of reagent. Ort/70-phthalaldhyde (OPT) reagent was prepared by dissolving100 mg of the aldehyde in 1 ml ethanol and 0.1 ml mercaptoethane. This was added to99 ml of 0.1 M sodium tetraborate buffer. In all cases a freshly prepared reagent wasused.-ODAP analysis. The extracts were filtered and taken in duplicate (0.1 ml each), treatedwith 3N KOH (0.2 ml) and kept in boiling water bath for 30 min. After cooling to roomtemperature, 3N HCI (0.2 ml) was added and the final volume made to 1 ml with water.A sample blank was prepared with the extract (0.1 ml) and water (0.9 ml) to account forany interference from the reaction between the OPT reagent and other constituents of theextract.The OPT reagent (2 ml for each sample) was added to the hydrolysed and controlsamples and absorbance measured at i^^ - 476 nm after 30 min. Reagent blank wasprepared by diluting the OPT reagent (2 ml) with water (1 ml). -ODAP concentration wascalculated from a calibration curve prepared by using commercial DAP.HCI and assuming100% conversion of -ODAP to DAP at the given conditions of hydrolysis [12].Statistical methodThe t-test was employed for comparing two means.

RESULTSNumbers and types of meals (Table 1 )Most families (95%) in both peasant associations had two main meals during the day. Themeals were served in the middle of the day and in the evening. The main dishes in bothcommunities were enjera and wott. The enjera was most often prepared from finger m illetalone and sometimes millet was mixed with teff, corn, and sorghum. The wott made ofshiro was mainly prepared from grass pea alone or mixed with chickpea (Cicer anietinum)in both villages. In some families, lentil was also used for wott.In Montura, potato was added to wott (in addition to legumes). Other foods like nifro andkollo were prepared from grass pea and consumed as snacks. Kitta was also consumedby a few of the families studied.

TABLE 1Dietary Habits of the Inhabitants of the L SativusGrowing Village of Shina and Montura

Type ofFoodEnjera MilletTeffSorgumCornBarleyL sativus NifroKolloKittaShiroVegetables' PotatoKale

Daily consumptiongm/person/day (% of total)SHINA MONTURA

578.3(50.8) 375(23.1)283.0 (24.8) 320 (19.6)160.5(14.1) 231 (14.2)117.5(10.3) 372(22.8)332 (20.4)91.3(49.5) 38(100)72.1 (39.1)21.0(11.4)consumed with sameevery meal

139(51.1)133(48.9)Different methods are used in the preparation of grass pea for shiro. The most commemethods used are as follows:

1. Aftr removal of foreign particles, grass pea is lightly toasted on a clay or meipan, transferred into cold water in a wooden trough or clay pot and thorougrwashed. The wet grass pea is toasted until browned and set aside for cooling.The husk is then removed by the use of a stone handmill {woftcho). The split grapea is again washed, set in the sun for a while, and then m ixed with peeled onicgarlic, ginger and basil. The mixture is pounded in a wooden mortar and set in tlsun for final drying. It is then ground into flour. This method is mostly usedMontura.2. Grass pea, free of all foreign particles, is added to boiling water on the sto\removed from heat after a few minutes and kept aside overnight. The followiday, the liquid is discarded and the grass pea toasted until browned. After coolifor a while, the toasted grass pea is dehusked and thoroughly washed. Twashed split grass pea is then set in the sun for a while. The rest of tpreparation until the final flour product is as described above (under 1).


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1118 R. TEKLE-HAIMANOT et al. LATHYRISM 1

3. Cleaned grass pea is toasted and dehusked. The split grass pea is washe d andset in the sun to dry. It is then ground into shiro.Grass pea, shiro wott preparation differed from house to house in the two peasantassociations. The most common methods are as follows:

1. Water is poured into a washed clay pot and placed on a stove. When the waterstarts to boil, grass pea shiro is added a little at a time while stirring the mixturecontinuously. Salt is added and the sauce cooked until done. When ready the wottis eaten warm or cold with enjera.

2. Peeled and chopped onion is fried in a clay pot by adding a little water at a timeuntil browned. If oil or water from crushed oil seeds is available, it is added whilef ry ing. Enough water is then added into the frying onion to boil. When the waterstarts to boil, shiro is added while stirring continuously. It is then cooked until done.This may take 20 to 30 minutes.Boiled grass pea (nifro) is most often consumed in Shina. Different methods are used fornifro preparation. The most common methods are as follows:

1. Grass pea is cleaned and washed with cold water. Water is added and the grasspea cooked for a short time. The water is then drained off. Fresh water is addedand the legume cooked until tender. Sometim es, chickpea or whole wheat is addedto the grass pea before adding water the second time.

2. Grass pea is washed 2-3 times with hot water. Cold water is added and thelegume cooked until it is soft enough to eat.3. Grass pea is soaked in cold water, left overnight, and the liquid discarded thefollowing day. It is then thoroughly washed, water added, and cooked.4. Cleaned grass pea is thoroughly washed with cold water. Enough water is addedand then the legume is cooked until done.

The roasted form of grass pea snack (kollo) was prepared in two alternative ways:1. Grass pea is added into boiling water and boiled for sometime. Excess water isdecanted and the grass pea is roasted over a hot clay or iron griddle.2. Grass pea is soaked in cold water for some time and excess water is decanted,roasting is carried out as in 1.

Very few people consumed grass pea unleavened bread (kitta) during the survey period.Kitta is eaten mainly at times of acute food shortages. It is prepared in the following

manner:Grass pea is cleaned and set in the sun for sometime. The husk is removed in istone mill {woftcho). The split grass pea is ground into fine flour. Water is addeca little at a time and the moistened flour kneaded to make a dough. Salt is addecto taste. The thick dough is spread over hot clay or metal griddle by hand ambaked on both sides for about 10 minutes.

Reliability check on chemical analysisThe results of assays obtained at the Addis Ababa University Department of Chemistron identical samples were in excellent agreement with those of a reference laboratory the Albert Einstein College of Medicine, New York.The Rao method involves an initial hydrolysis of -ODAP to 2,3- diaminopropionic aci


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1120 R. TEKLE-HAIMANOT et al. LATHYRISM 11TABLE 2-N-Oxalyl, -diaminopropionic acid (-ODAP) content in different grass pea foodsamples from Shina and Montura peasant associations

Sample typeWhole seedDecorlicataed seedShiro-flourShiro-sauceNifro-boiled

Size ofsamples6310501313

-ODAP content (mg/100g)Higest

81 069 042 933 870 4

Lowest28 223 915511370

Average45 544 427 019 047 1

TABLE 3-ODAP content of boiled grass pea prepared in the laboratory using differentmethods of boiling.Code

3637383940414243444546MeanS.d.

-ODAP content (mg/100g)Dry seed

488549387514394514528429359253514488.191.8

1282211338465380402394275246211387326.586.0

2239211378444373373429330389218380342.282.5

3261232465401359373420380450268373362.077.3

4331553408521331324345401317183408374.7101.7

1. Grass pea boiled for a short time, excess water drained. Then boiled with fresh water.2. Grass pea washed with boiling hot water 3 times and then boiled with fresh water.3. Grass pea soaked overnight, excess water discarded. Then boiled with fresh water.4. Grass pea washed with cold water and then boiled with fresh water.Simulated kitchen experiments confirmed that steeping grass pea in a large volume ofwater for 3 minutes and decanting the excess water leeched out approximately 30% of-ODAP. Improved detoxification was achieved when the boiled snack, nifro, wa sprepared from grass pea boiled directly with hot water as compared to that preparedfrom grass pea soaked overnight in cold water. This was statistically verified, (Tables3 and 4).Roasting grass pea preceded by steeping of the seeds achieved some degree of -ODAP reduction. On the other hand, when grass pea roasting was done after onlysoaking the seeds, there was an increase in the -ODAP level in six of 11 samples

(Table 5). Although the absolute differences were not statistically significant,neverthless the trend in the increase of ODAP level could not be ignored.Higher levels of -ODAP were observed in the grass pea bread produced by thekitchen experiment compared with the original grass pea samples. The increase,which was found in 14 out of 20 identical experiments, ranged from 4.1-48.9 % (Table6)< However, due to the high variability in ODAP levels of the bread samp les, thedifferences were not statistically significant.

TABLE 4-ODAP content of boiled grass pea {nifro) samples prepared in the kitchen laboratoryby two different methods: boiling directly and after soaking

Samples12345678:910Mean; S.d.

-ODAP content (mg/100 g)

Dry seeds9018311,042824739796591367475598716.4206.0

Soaked and boiled seedWet seed

739732936697789634380282252343

Discardedwater3.58.210.99.711.58.74.41.31.78.1

Boiledseed683676429634704514338334343264491.9107.9

Boiled seedDiscardedwater

300420598655662535275264387370

Boiledseed479324697408387331253246317229 :367.1139.8

DISCUSSIONiFood lgumes, including grass pea, form a major part of the protein source andnutrients of the Ethiopian diet. Grass pea, though considered a poor person's food, iscultivated quite extensively in areas of the country subject to adverse agriculturalconditions, particularly flooding and drought. Grass pea production is concentrated inthe northwest region of Ethiopia, and is also cultivated in northeast, central andsoutheast regions.


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1122 R. TEKLE-HAIMANOT et al. LATHYRISM 11

TABLE 5-ODAP content of grass pea roasted (kollo) by two different methods

Raw grass pea

Code50515253541/12/13/14/15/16/1MeanS.d.

-ODAPmg/100gm

704500943789725608436606829681676681.6144.4

Roasted Sample 1Time(min)1.153.154.155.004.05

-ODAP (mg/100g)Discardedwater

9.97.021.816.912.7

seed651591831866667476488867656503596653.8145.3

Roasted sample 2Time(min)4.383.335.505.253.40

-ODAP (mg/100g)Discardedwater

8.66.314.622.88.7

seed7395841133894588770405732613433603681.3207.4

It ranks fifth in total area among Ethiopian food legumes with a mean of 60,030 ha ofland (8.7%) and 42, 200 t (7.6%) of grain (18). Probably as a reflection of therepeated food shortage situations in Ethiopia, the production of grass pea increasedby 20% between 1981 and 1987 (19). Sporadic cases of lathyrism continue to appearin the northwest part of the country (10).In the area studied here, grass pea is mainly used to produce the flour {shiro) used inthe preparation of the Ethiopian sauce or gravy, shiro wott. This was the case in bothvillages studied. The pulse is also used as snacks in the roasted and boiled forms,mainly in Shina, the village where lathyrism is endemic at a prevalence rate of 3%(10). The bread form is consumed widely only in extreme famine situations as was thecase during the major lathyrism epidemic of the Fogera and Dembia districts in1976/77 (5). The consistent view that transpires from interviews of consumers of grasspea is that the pulse has an excellent taste. In the flour form, it possesses the uniquewater-absorbing and expansive properties which are advantageous during thepreparation of the gravy, wott. As a result, in central parts of Ethiopia where grass peais not cultivated, the pulse is presently popularly used as an admixture in theproduction of shiro. Food adulteration, with Lathyrus sativus seeds have been reportedfrom India (20).There are only a few studies that have addressed the potential neurotoxicity of

different food preparations of grass pea with reference to -ODAP content. Dwivediand Singh have suggested that the Indian staple, ghotu, prepared by cooking amixture of Lathyrus sativus and rice in water to form a stiff porridge (paste balls)precipitated lathyrism more rapidly than chapati, the unleavened bread form. This wasbased on an epidemiological observation. The researchers further suggested that thepresence of water in the ghotu permitted the toxic component to be released insolution providing a more rapid absorption (21). To our knowledge -ODAP levelshave not been systematically assayed in the different grass pea food preparations,thus this hypothesis has not been tested.Our kitchen experiments have demonstrated that the Ethiopian unleavened bread,kitta, which in its form and preparation is equivalent to the Indian chapati, has higherdetectable levels of -ODAP than the dry grass pea seeds from which it is prepared.We have also epidemiological evidence that in the lathyrism outbreaks and epidemicsof'northwest Ethiopia, the bread {kitta) form was consumed in large quantities inaddition to other types of grass pea-derived foodstuffs. Gebre-ab et al. writing aboutthe 1976/77 lathyrism epidemic in northwest Ethiopia, when over 2500 persons wereaffected, observed that the commonest form of grass pea consumed by the affectedpopulation included kitta, nifro an d kollo. They also commented that the kitta form wasthen a new introduction into the area of the epidemic (5). In normal times, as was thecase during our survey, villagers in the lathyrism-endemic area used grass peacommonly as snacks in the boiled and roasted forms and very little as bread.Similarly, we have shown that the roasted form {kollo), if roasted without prior aqueousteeping, may contain higher levels of -ODAP than the native seeds. The conclusiondrawn from these experiments, which were designed to simulate the condition in thevillages, is that the level of -ODAP is maximal in the bread and roasted forms ofgrass pea. The level is lower in the boiled snacks, particularly when the process ofsteeping is employed. The flour form, shiro and the sauce prepared there from,contained relatively lower levels of -ODAP and thus may not be so hazardous.Detoxification of the grass pea through aqueous leeching of the toxic component (i.e.steeping) has been described previously (22). Mohan er al. have suggested theremoval of the toxin from L. sativus seeds through two alternative methods: steepingthe dehusked seeds in hot water for several hours and boiling'the seeds in water anddraining out the supernatant. This removed 70-80% of the neurotoxin (23).Moslehuddin et al. also found that washing the seeds partially removed the neurotoxitwhile fermentation, steaming or autoclaving appeared to have little effect (24).Ayyagari et ai, based on analysis of a limited number of samples, concluded that the-ODAP content did not decrease much in prepared food. Dry heating, roasting, deepf ry ing, or boiling did not alter the -ODAP levels (25). On the other hand, Rao et al.recorded that the roasting of grass pea for about 15-20 minutes at 140C renderedthe seed free of neurotoxin. Howevere, they observed that chapati making made the


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1124 R. TEKLE-HAIMANOT et al. LATHYRISM 112grass pea foodstuff more toxic (26). However, in the case of the roasting they d id notmention w hether the seeds were washed or boiled before roasting.TABLE 6-ODAP content of bread (kitta) prepared in the laboratory from different L sativussamples

Dry grass peaCODE

363738394041424344454667686970711/11/21/31/4MeanS.d.

-ODAPmg/100gm48854938751439451452842935925351 4367475452598299416416416416439.285.5

% moisture70.876.563.165.462.870.862.769.867.070.871.466.765.364.377.5'65.3

Bread sample-ODAPmg/100gm

394426429584521535786583472331401510542590678502493244224227473.6145.5

grass peawet sample(gm)8080808080808080808080

Bread preparationwater formixing(ml)

6060606060756060605560

baking time(min)7777677101087

However, a serious and challenging question that will arise out of our findings will be why havehigher -ODAP levels been recorded in the bread and roasted forms of grass pea? How doesdirect heat increase -ODAP content of the grass pea?A plausible explanation relates to changes in the extractability of the -ODAP by thermallyintroduced processes. Other workers who have been studying the extractability of the toxin byimbibing with water over prolonged periods and determining the levels of -ODAP in the imbibedseeds, as well as in the fluid, have found that the two values when added up differ from the levelof toxin found by direct analysis of seeds [Lambien, F. personal communication]. Heat treatmentmay induce enzymatic changes, damage lipid membranes or even alter the nature of thecounterions that may be associated with the toxin. It Is possible to speculate changes in thecomposition and availability of counterions, K+, Na+, Mg++ that may be induced by heat. For

instance, the potassium salt of -ODAP is more extractable than the sodium salt. Little or noinformation is available on the relative extractabilities of other divalent metal salts of the toxin.Although there have been suggestions that the reduction of over 65 % in -ODAP content couldbe considered safe, the non-toxic levels of -ODAP have not yet been established (20). In fact,it will not be easy to set such a cut of standard because of the different concentration of -ODAPin germplasm lines of L. sativus. For instance, in the field samples we collected the -ODAPcontents varied from 128 to 985mg/100 gm of dry seeds. Thirty or thirty-five per cent of the mosttoxic sample may still contain a significant amount of -ODAP.

ACKNOWLEDGEMENTSThese studies were made possible by the grant from Band Aid which was obtained through ThirdWorld Medical Research Foundation (TWMRF). BA is grateful for additional support from theSwedish Agency for Research Cooperation with Developing Countries (SAREC). Aregay Waktola,Principal Investigator of the Lathyrism P roject in Ethiopia, Valerie Palmer of TWMRF and PennyJenden of Band-Aid are thanked for their interest and support. We are grateful to AsmeromKidane, Professor of Statistics, AAU, for helpful comm ents. We are grateful to the enumeratorsthat participated in field surveys.

REFERENCES1. Prasad LS, Sharan RK. Lathyrism. In: Vinken P J, Bruyen G W. (eds). Handbook ofClinical Neurology: Intoxication of the Nervous System Amsterdam: North HollandPublishing Co., 979: 505-14.2. Spencer PS, Schaumburg HH . Lathyrism: A neurotoxic disease. NeurobehavioralToxicology and Teratology 1983;5:625-9.3. Dwivedi MP and Mishra SS. Recent outbreak of lathyrism and experience withpropagation of detoxified L sativus. Proc Nat Soc Ind 1974;19:23-30.4. Kulkarni SW, Attal HC, Choubey BS. An epidemiologlc study of lathyrism in AmgaonBlock, Bahandra district. Ind Med J Res 1977;66:602-10.5. Gebre-Ab T, Wolde-Gabriel Z, Maffi M, Ahmed Z, Ayele TM, Fanta H. Neurolathyrism- areview and a report of an epidemic. Ethiopian M ed J 1978;16:1-11.6. Spencer PS , Roy DN , Ludolph A, Hugon J, Schaumburg HH. Lathyrism: Evidence for roleof the neuroexcitatory amino acid BOAA. Lancet 1986li:1066-67.7. Dwivedi MP. Epidemiological aspects of lathyrism in India- a changing scenario. In:Spencer P S (ed). Grass-pea: Threat and Promise. New York: Third world Medical

Research Foundation, 1980:1-26.8. , Haque A, Mannan MA. The problems of lathyrism in Bangladesh. In: Spencer PS (ed).Grass-pea: Threat and Promise. New York: Third World Medical Research Foundation,1989:27-35.9. Tekle Haimanot R. Lathyrism in Ethiopia. In: Spencer PS (ed). Grass-pea: Threat andPromise. New York: Third World Medical Research Foundation, 1989:36-40.


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1126 R. TEKLE-HAIMANOT et al.10. Tekle-Haimanot R, Kidane Y, Wuhib E, Kalissa A, Alemu T, Zein AZ, Spencer PS.Lathyrism in rural Northwestern Ethiopia: a highly prevalent neurotoxic disorder. Int J Epid1990;19:664-672.11. Ethiopian Nutrition Institute. Ethiopian Traditional Recipes. Addis Ababa: CommercialPrinting Press, 1980.12. Rao SLN. A sensitive and specific calorimetric method for the determination of oc..diaminopropionic acid and the Lathyrus sativus neurotoxin. Anal Biochem1978:864:386-95.13. Abegaz B, Kebede N, Asmelash S. HPLC analsysis of -N-Oxalylamino-L-alanine, theneurotoxin in the legume Lathyrus Sativus. In: Spencer P S (ed). Grass-pea: Threat andPromise. New York: Third W orld Medical Research Foundation, 1989:128-132.14. Wu G, Bowlus SB, Kim KS, Haskell BE. L-2-Oxalylamino--aminopropionic acid, andisomer of Lathyrus Sativus neurotoxin. Phytochemistry 1976;15:1257-1 59.15. Harrison FL, Nunn PB , Hill RR. Synthesis of a- and -N-oxalyl-L-cc, -diaminopropionicacids and their isolation from seeds of Lathyrus Sativus.Phytochemistry 1977:16:1211-1215.16. Chase RA, Pearson S, Nunn PB, Lantos PL. Comparative toxicities of a- and -N-oxalyl-L-a, -diaminopropionic acids to rat spinal cord. Neurosci Lett 1985;55:89-94.17. Bell EA, O'Donovan JP. The isolation of a- and - oxalyl derivatives ofa,,oxalylalminopropionic acid which occurs together with neurotoxin in this and otherspecies. Phytochemistry 1966;5:1211-1219.18. Wolde-Amlak A, Asgelil D, Bekele H, Regassa E, Wasie H, Yeshanew A. Grass Pea(Lathyrus Sativus) Research and its production potential in Ethiopia. (A paper presentedat the 3rd Triennial Colloquium of INILSEL, Dhaka, Bangladesh, 30 November- 3December 1991).19. People's Democratic Republic of Ethiopia: Facts and Figures. Central Statistic Office,1987.20 . Golpalan C. The Lathyrism problem: current status and new dimensions. NutritionFoundation of India, 1984:12-14.21 . Dwivedi MP, Singh SP. The lathyrism problem. In: Nutrition Foundation of India. 1984:19-33 .22 . Jha K. Effect of the boiling and decanting method of Khesari (Lathyrus sativus)detoxification, on changes in selected nutrients. Arch Lathinoma Nutr; 1987 Mar;37(1 ):101 -23. Mohan VS, Nagarajan V, G opalan C. Simple practical procedures for the removal of toxicfactors in Lathyrus sativus (Khesari dal), lnd J Med Res 1966:54:410-9.24. Moslehuddin ABM, Hang YD, Stoewsand GS. Evaluation of the Toxicity of processed

Lathyrus Sativus seeds in chicks. Nutrition R eports International 1987;36:851-855.26. Ayyagari R, Rao BSN, Roy DN . Lectins, trypsin inhibitors, BOAA and tannins in legumesand cereals and the effects of processing. Food Chemistry 1989:34:229-238.Accepted for publication June 16, 1993.

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