appreciable amount of curd and water, is in fact impure butter fat, more or less rancid when collected by the manufacturers. The ghee of the Indian market as well as of the Bengal villages is not merely melted out from butter and dried of water, but is heated to its own boiling temperature both to bring out the aroma for which it is valued and to destroy the curd in order to ensure its keeping quality. The manufacturers of ghee after making their collections from the houses of several villages boil their blend for these purposes. Then, as. has already been mentioned, the ghee is as a rule heated to its boiling point in cooking before it is taken as food. Bacharach made his experiment on a rat with a sample of ghee which he got from Bombay, and though the unsaponifiable matter extracted from this ghee gave the blue colour with antimony trichloride, the result on the rat was negative, but we do not know if this sample was genuine. This is the state of things, and we are fre- quently pressed for our opinion regarding the vitamine value of these articles and specially of ghee. I found ready to hand a brood of young albino rats belonging to my assistant Mr. N. K. Chatterji, and Mr. Chatterji at once /offered them to me for which I acknowledge /here my indebtedness to him. I hastened to / avail myself of this opportunity to test these fats on them. The following paragraphs give an account of the work and the results thereof. THE YITAMINE VALUE OF THE F00I7 In carrying out this experiment, I could not FATS OF BENGAL keep to the details of the very sound lines prescribed by the Accessory Food Factors Com- (A PRELIMINARY STUDY) mittee in their report to the League of Nations By B. B. BRAHMACHARI, d.p.h. Health Organisation as given in the Lancet, Director, Bengal Public Health Laboratory 1928, Vol. I, p. 148. The reasons for the will appear in their respective Mustard oil and ghee are the only two fats departures which enter as such into the dietary of the places. Rats.?The rats under the were people of Bengal. Mustard oil is taken through- experiment seven in all of the same out the province, in towns as well as in villages, number, litter, they were not 20 to 30 days old, but were already by the rich as much as by the poor. Ghee is 9 weeks old before the experiment could be much more used in the towns than in the villages arranged. They were between 40 and 50 and by the richer and the more well-to-do grammes in weight. than the mass. Both are used for culinary by Basal diet.?The basal diet used was as purposes, ghee particularly for cooking sweets. below :? Occasionally they are taken by mixing with boiled vegetables, and ghee also with, cooked Casein 19 rice, the latter always in such cases being first Starch 55 to the boil. of such dietetic brought Though Lard 16 importance, no study has been made, so far Yeast 5 as I know, on the vitamine contents of mustard mixture oil. The only works on ghee in this direction Salt 5 that I could find on record are those of Ghosh (1922) and of Bacharach (1930). Ghosh found This was practically the same as that of by feeding experiments on rats that ghee was Drummond and Coward (1920) used by I as it- equivalent in vitamine value to the average Dr. Ghosh. But omitted orange juice butter. But his ghee was not the ghee of the might contain vitamine A and further as rats Indian market nor yet of the villages of Bengal; are not known to suffer from scurvy (1929). it was the ghee of the villages of Upper India, I used Merck's soluble starch in place chiefly of the provinces of Allahabad, Agra and of rice starch and lard in place of the purified ' and for Oudh, it is the kacha' or underdone ghee, reduced vegetable oil of the formula, melted at a low temperature, containing an the yeast extract in it I also used Merck's 378 THE INDIAN MEDICAL GAZETTE [July, 1932 medicinal yeast. I purified the casein by heat- other vitamines in it. The other five rats were ing it at 120?C. in thin layers exposed to air given lard first. After the depletion of and towards the end of the experiment by vitamine A in them, as shown by fall of weight one and its in which treating it in a Soxhlet flask with petroleum in becoming steady three, ether. The salt mixture of the diet was after took twenty-three days, the whole of the lard the formula of McCollum and Simmonds (1918), was replaced by the same weight of the test fats and of the of the butter but by an oversight the calcium lactate in my control, i.e., fat, salt mixture was one-tenth of what is given in though the weight was still rising in the remain- the formula; this deficit of lime was however ing one. Two of them got mustard oil, two and the one which had lost a common factor in the diet of all my rats and, ghee, weight got butter fat. I could not three animals for as will be seen below, did not prevent normal get growth in the control animals. For anti- each of these groups, the five being all that were available. I did not rachitic vitamine I had to depend upon expos- begin by adding my ing the animals to the diffuse light of the setting test fats in small quantities, because my aim was not for the sun, not without success as is evident from the quantitative but, present, only growth of the control animals and from qualitative; my object was not to ascertain the examination of the bones of those which minimum dose of these fats which would restart succumbed. growth but to find out if they contain the Fats.?My test fats were of course mustard vitamine at all in amount commensurate to oil and ghee. For depletion of the vitamine A the physiological requirements. So the dose was the maximum of these fats which the as a preparation for the test I used lard. given Butter fat was used as control. The mustard rats could possibly take, forming as it did about 16 cent, of the food. In oil was the pure oil we got from a jail in this per by weight province. The ghee was a mixture of samples case the vitamine was found in them, my found genuine in our laboratory and blended object was to follow it up by the finding in of of in the usual way. The butter fat was the milk them of the quantity it. Lastly instead fat melted out of Keventer's butter on a water stopping at the end of the 4th week, I pro- the for over another week bath; I had no time to prepare my own butter. longed experiment on Keventer's butter is among the best in the as the animals kept butter fat and thriving market. on it suddenly got ill with diarrhoea and, in Procedure.?One pair of these rats were given consequence, it became necessary to prolong butter fat for the whole of their fat food the observations on them. throughout; these served as control for the Result.?The result of the experiment is completeness of and balance in the proximate given in the following table under the dates principles of the diet and sufficiency of the shown :?

Weight in grammes

Rats On lard On test fat Test fat Remarks

Number Sex 23.7 3.8 9.8 15.8 22.8 29.8 5.9 12.9 19.9

I Male 50 70 75 50 60 70 85 80 75 Butter fat Died on 28th II Male 55 80 100 100 75 Mustard oil August, weight BO grm. 65 65 65 Died on 18th Sep- V Female .. 50 50 65 70 70 tember, weight 50 grm. Died on 15th IV Male 50 70 75 75 50 60 60 60 Ghee Sep- tember, weight 50 grm. 70 50 55 55 Died on 20th III Female .. 40 60 65 65 70 Sep- tember.

On butter fat throughout

Sex. 23.7 3.8 9.8 15.8 22.8 29.8 5.9 12.9 19.9

Male 50 80 100 110 120 140 140 150 150

110 160* 125 120 Female .. 50 80 85 95 130

* Was. pregnant: brought forth 9 young ones on the 6th September. July, 1932] VITAMINES IN BENGALI DIETARIES: BRAITMACHARI 379

The relative nourishment of the rats on these I the closing day of the experiment, and expired different- fats in the experiment will be seen at on the 23rd. (a) The diarrhoea was not accom- a glance from the following graph :?- panied by any suggestion of xerophthalmia, the animal looked about with VitamineVitamine valuevalue ofof butterbutter fat,fat. gheeghee andand mustardmustard oil.oil. throughout open and the were to [As[As foundfound onon rats].rats]. eyes eyes bright the last; (b) the disease also attacked simultaneously the other two rats on butter fat, the female one from it was LardLard suffering severely it; (c) limited ButterButter fatfat to these rats only, those on the other fats passed Ghee normal stools; on the outbreak of the disease, the food of these three rats for the week was Mustard oiloil found rather offensive smelling and it was further found that some food which had acci- dentally dropped on the ground was returned to the bottle; hence it is very likely that the disease was an infection carried through the food of these three rats.

(2) Mustard oil failed to restart growth. The two rats 011 it had photophobia and rough coats. In the male which had remained steady at 100 grammes in the last week of the lard diet the weight dropped to 75 grammes in one ? week, and in another six days the animal died, weighing 50 grammes. The female rat had still been putting on weight and was at 70 grammes when she got the mustard oil, she remained at 70 grammes for one week, then 23rd 3rd 9th 15th 22nd 29th 5th 12th 19th steady July to 65 the next con- Aug. Aug. Aug. Aug. Aug. Sept. Sept. Sept. ; dropped grammes by week, tinued at that weight for the succeeding two weeks and died after 6 more with the (1) Butter fat.?The curve of growth of the days at 50 grammes. Both these animals two rats kept from the beginning on butter fat weight had no from and for the fat food is the same as that of the symptoms apart photophobia the deaths were rather sudden. normal growth of Osborne and Mendel given on rough coat, On the organs were found page 123 of Funk's Vitamines (1922). Their post-mortem normal, the intestines and the bones well formed rats as well as ours started from 50 grammes, empty and not soft. their rats grew to 150 grammes in 49 days, our male rat reached that weight on the 51st, (3) Ghee.?The male rat which had steadied our female rat on the 44th 160 day weighed at 75 grammes on lard developed inflammation but she was on the grammes, pregnant and of the respiratory passages, photophobia and birth to ones. following day gave nine young dcpilation and dropped to 50 grammes in the The for rats was therefore well ration these first week on ghee, recovered from the respi- balanced. ratory inflammation and put on 10 grammes in course of the next continued at that The result of the butter fat on the rat week, two next weeks depleted of the vitamine is also remarkable. level for the succeeding and to 50 and died in another This rat after rising from 50 grammes at dropped grammes three The of avitamosis were the start to 70 grammes the 17th day days. symptoms by in this suddenly developed xerophthalmia with photo- very much pronounced rat, the depilation was the fur was the phobia and depilation specially round the eyes, advanced, very rough, ended in destruction of the had inflammation of the respiratory passages xerophthalmia cornea and the became irri- and lost 20 grammes in 6 days. The dyspnoea blindness, temper the animal was and moved round was intense with the head thrown up with each table, snappish and round in with recesses of rest. The respiration and the animal was in the last frenzy bones of this animal too no of stage when the lard was replaced with butter showed signs rickets. The female which had with fat. The effect of the change was marvellous; began 40 last in three days the dyspnoea disappeared; in the grammes halted at 65 grammes in the week on on 5 in the first course of a week the animal opened its eyes; lard, put grammes week on the ulcer wThich had formed on the left cornea ghee, continued at 70 grammes through the next to 50 in third healed up, the fur improved and in three weeks week, dropped grammes rose to 55 in another the body weight went up from 50 grammes to week, again grammes week and was still 55 on the last 85 grammes. At this stage, it had diarrhoea grammes day of the The of avita- of a severe type, passed into a low condition, experiment. symptoms mosis were marked. It died 12 after with lost weight dropping to 80 grammes in seven days extreme days and 75 grammes on the 19th September xerophthalmia. 380 THE INDIAN MEDICAL GAZETTE [July, 1932

Conclusions.?-From the above experiment we may fairly conclude :? (1) that mustard oil, if used as the only source of vitamine A for a growing animal, can neither promote growth nor even support life; (2) that though butter fat contains a fair amount of vitamine A, ghee is little better than mustard oil in this respect. But in connection with ghee we must remember the above experi- ment bears on (?) the genuine ghee of the Bengal Food Adulteration Act, i.e., ghee that passes the standards of the Act. It cannot to 1 apply (?) the kacha' or imperfectly melted out ghee of up-country villages, which, as I have already pointed out, is practically impure butter fat more or less rancid, and which, no wonder, Dr. Ghosh found to be equivalent in vitamine value to the average butter. It may not also apply to (c) the inferior ghees of the market which are among other adulterated,' ' perhaps things with the kacha ghee, or are heated just suffi- ciently to put off the decomposition till passed on to the customers, with the fraudulent object ' that the water and the curd of the kacha' ghee add to the weight of the commodity, It must have been with some adulterated ghee like this, that Dr. Ghosh could, restart growth in rats while he failed to do so with his remelted ghee.

References Bacharach, A. L. (1930). Brit. Med. Journ., Vol. II, p. 141. Drummond, J. C., and Coward, K. H. (1920). Biochem. Journ., Vol. XIV, p. 661. Funk, C. (1922). The Vitamines, p. 123. Baltimore: Williams and Wilkins Co. Ghosh, S. N. (1922). Biochem. Journ., Vol. XVI, p. 35. Martindale, W. II., and Westcott, W. W. (1929). Extra-Phcirmacopccia, 19th Ed., Vol. II, p. 104. London: H. K. Lewis and Co. . McCollum and Simmonds (1918). Journ. Biol. Chem., Vol. II, pp. 33, 55, 104.