503 SPECIAL ARTICLES THE ANTIRACHITIC VITAMIN D. ANNOUNCEMENT OF A STANDARD BY THE MEDICAL RESEARCH COUNCIL. THE Medical Research Council’s Committee on Accessory Food Factors have long attempted to find generally acceptable conventions for the quantitative record of the activities characteristic of the different accessory factors, or so-called vitamins. As in other instances, however, any unit of activity based on the production of a stated degree of a biological effect, however carefully this may be defined, must be liable to wide variation with differences in the conditions under which the test is conducted. This is especially so in testing for a vitamin action, where the effect on which measurement is based is the correction of an abnormal condition produced by a deficient diet. The severity of the condition to be treated will vary with differences in the dieting and treatment of the i experimental animals, as well as with racial and individual differences in the stocks. Other bio- logical standards, such as those for sera, hormones, &c., held by the Council’s Department of Biological i Standards, are based on stable standard materials, in terms of each of which a unit of activity is defined which is independent of the details of the biological method used in its measurement. Here a measure- ment of activity in units is made on a strictly compara- I tive basis, with reference to the standard. I NEED FOR A BASIS OF MEASUREMENT. With regard to one of the vitamins, the anti- rachitic vitamin known as Vitamin D, this necessity for a common material standard was recently emphasised in the following recommendation by the Committee :— Any unit or standard based directly on the response of the experimental young rats to administration of anti- rachitic material is unsatisfactory, owing to the dispro- portionate variation in the response to equal doses of apparently similar animals, when tested at different seasons of the year or bred in different laboratories. The Vitamin D unit should rather be referred to a stated quantity of some standard antirachitic material, with which comparison can be made by any reliable biological method. The discovery of the method of producing this vitamin artificially from ergosterol has led to its manufacture on a large scale, its issue in many forms for therapeutic use, and its addition to various types of food materials. The proper desire of manu- facturers to use and to supply such preparations with a quantitative indication of activity has led to the appearance of several unofficial " units," mostly defined in terms of some biological effect, and having no clearly determined relation to one another. The need for a common basis of measurement is, therefore, accentuated, in this case, by an existing and growing confusion. One of the provisional units current in this country-namely, that adopted by the Pharma- ceutical Society’s Laboratories when reporting on the Vitamin D content of materials submitted to them for assay, has, indeed, been based on a material standard consisting of an oily solution of irradiated ergosterol, supplied to them for this purpose by the National Institute for Medical Research. Only a small quantity of this material had been prepared, however, and sufficient data were not available to justify the assumption that it would have the stability necessary for a standard and allow the Committee to recommend it for general use. PREPARATION OF THE NEW STANDARD. It was accordingly decided to arrange for the preparation by the National Institute of a similar solution, on a scale sufficient for its eventual distribu- tion to the numerous laboratories or institutions likely to require it, and to organise an investigation of its properties by different experts working on behalf of the Committee. A quantity of ergosterol from yeast, purified and dried by methods carefully recorded was subjected, in an accurately made alcoholic solu- tion, to irradiation with the rays from a mercury arc, the physical details of the procedure being measured and recorded with the greatest practicable accuracy. All conditions known to influence the amount of Vitamin D formed should, therefore, be reproducible at any future date, or in any other laboratory, leaving only the final adjustment of a new standard to equivalence with this original one to be made on the basis of comparative biological assay. The irradiation products were carefully freed from alcohol, dissolved in pure, unadulterated olive oil, and the volume adjusted to produce a known concentration, in terms of ergosterol used. The solution was then placed, and its main bulk has since been kept in the Institute’s refrigerating chamber at a temperature of - 4° to 0’ C. TESTS OF STABILITY. Evidence of the stability of such a preparation, under different conditions and over a sufficiently long period, being of central importance for a decision as to its use as a standard, the investigations promoted by the Committee have, of necessity, been of a corresponding length. The experimental results from the different contributing laboratories have recently been completed, and have now been carefully reviewed by the Committee, who have stated their conclusion from them in the following terms :- From these tests it appears that specimens of irradiated ergosterol, prepared in the manner described above, do retain their activity unchanged for two years if maintained at or below 0’ C. If they are kept at room temperature loss of activity may occur. COMMITTEE’S RESOLUTIONS. The evidence of the different investigators being uniformly favourable as to the suitability of this solution in other respects for use as a standard in comparative biological measurement of Vitamin D the Committee decided to recommend its adoption as a standard, and the definition of a unit in terms of it, and adopted the following resolutions :— I (a) That the standard solution of irradiated ergosterol prepared and maintained by the National Institute for Medical Research be recommended for adoption as a standard for the comparative estimation of Vitamin D. (b) That the unit of Vitamin D be defined as the anti- rachitic potency of a quantity of this preparation correspond- ing to 0.0001 mg. of the ergosterol used in its production. z, (c) That either the X ray method, the " Line test," or chemical analysis of the bones of the experimental animals, be recommended for use in the estimation of Vitamin D. (d) That supplies of the standard material be made available for general distribution from the National Institute for Medical Research, Hampstead. As the supply is limited Iit is hoped that users will themselves prepare standard solutions of irradiated ergosterol for use in individual tests, retaining the National Institute’s material for reference. It is important that all such standard materials be kept at temperatures not exceeding 0° C.
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503
SPECIAL ARTICLES
THE ANTIRACHITIC VITAMIN D.
ANNOUNCEMENT OF A STANDARD BY THE
MEDICAL RESEARCH COUNCIL.
THE Medical Research Council’s Committee on
Accessory Food Factors have long attempted to findgenerally acceptable conventions for the quantitativerecord of the activities characteristic of the differentaccessory factors, or so-called vitamins. As in otherinstances, however, any unit of activity based on theproduction of a stated degree of a biological effect,however carefully this may be defined, must be liableto wide variation with differences in the conditionsunder which the test is conducted. This is especiallyso in testing for a vitamin action, where the effect onwhich measurement is based is the correction of anabnormal condition produced by a deficient diet.The severity of the condition to be treated will varywith differences in the dieting and treatment of the iexperimental animals, as well as with racial andindividual differences in the stocks. Other bio-
logical standards, such as those for sera, hormones,&c., held by the Council’s Department of Biological iStandards, are based on stable standard materials,in terms of each of which a unit of activity is definedwhich is independent of the details of the biologicalmethod used in its measurement. Here a measure-ment of activity in units is made on a strictly compara-
Itive basis, with reference to the standard. INEED FOR A BASIS OF MEASUREMENT.
With regard to one of the vitamins, the anti-rachitic vitamin known as Vitamin D, this necessityfor a common material standard was recentlyemphasised in the following recommendation by theCommittee :—
Any unit or standard based directly on the response ofthe experimental young rats to administration of anti-rachitic material is unsatisfactory, owing to the dispro-portionate variation in the response to equal doses ofapparently similar animals, when tested at different seasonsof the year or bred in different laboratories. The Vitamin Dunit should rather be referred to a stated quantity of somestandard antirachitic material, with which comparison canbe made by any reliable biological method.
The discovery of the method of producing thisvitamin artificially from ergosterol has led to itsmanufacture on a large scale, its issue in many formsfor therapeutic use, and its addition to varioustypes of food materials. The proper desire of manu-facturers to use and to supply such preparations witha quantitative indication of activity has led to theappearance of several unofficial " units," mostlydefined in terms of some biological effect, and havingno clearly determined relation to one another. Theneed for a common basis of measurement is, therefore,accentuated, in this case, by an existing and growingconfusion. One of the provisional units current inthis country-namely, that adopted by the Pharma-ceutical Society’s Laboratories when reporting on
the Vitamin D content of materials submitted to themfor assay, has, indeed, been based on a materialstandard consisting of an oily solution of irradiatedergosterol, supplied to them for this purpose by theNational Institute for Medical Research. Only a
small quantity of this material had been prepared,however, and sufficient data were not available tojustify the assumption that it would have the stability
necessary for a standard and allow the Committeeto recommend it for general use.
PREPARATION OF THE NEW STANDARD.
It was accordingly decided to arrange for the
preparation by the National Institute of a similar
solution, on a scale sufficient for its eventual distribu-tion to the numerous laboratories or institutions
likely to require it, and to organise an investigationof its properties by different experts working on behalfof the Committee. A quantity of ergosterol fromyeast, purified and dried by methods carefully recordedwas subjected, in an accurately made alcoholic solu-tion, to irradiation with the rays from a mercury arc,the physical details of the procedure being measuredand recorded with the greatest practicable accuracy.All conditions known to influence the amount ofVitamin D formed should, therefore, be reproducibleat any future date, or in any other laboratory, leavingonly the final adjustment of a new standard to
equivalence with this original one to be made on thebasis of comparative biological assay. The irradiationproducts were carefully freed from alcohol, dissolvedin pure, unadulterated olive oil, and the volumeadjusted to produce a known concentration, interms of ergosterol used. The solution was thenplaced, and its main bulk has since been kept in theInstitute’s refrigerating chamber at a temperature of- 4° to 0’ C.
TESTS OF STABILITY.
Evidence of the stability of such a preparation,under different conditions and over a sufficientlylong period, being of central importance for a decisionas to its use as a standard, the investigations promotedby the Committee have, of necessity, been of a
corresponding length. The experimental results fromthe different contributing laboratories have recentlybeen completed, and have now been carefully reviewedby the Committee, who have stated their conclusionfrom them in the following terms :-
From these tests it appears that specimens of irradiatedergosterol, prepared in the manner described above, doretain their activity unchanged for two years if maintainedat or below 0’ C. If they are kept at room temperatureloss of activity may occur.
COMMITTEE’S RESOLUTIONS.
The evidence of the different investigators beinguniformly favourable as to the suitability of thissolution in other respects for use as a standard incomparative biological measurement of Vitamin Dthe Committee decided to recommend its adoption asa standard, and the definition of a unit in terms of it,and adopted the following resolutions :—
I (a) That the standard solution of irradiated ergosterolprepared and maintained by the National Institute forMedical Research be recommended for adoption as a standardfor the comparative estimation of Vitamin D.
(b) That the unit of Vitamin D be defined as the anti-rachitic potency of a quantity of this preparation correspond-ing to 0.0001 mg. of the ergosterol used in its production.
z, (c) That either the X ray method, the " Line test," orchemical analysis of the bones of the experimental animals,be recommended for use in the estimation of Vitamin D.
(d) That supplies of the standard material be madeavailable for general distribution from the National Institutefor Medical Research, Hampstead. As the supply is limitedIit is hoped that users will themselves prepare standardsolutions of irradiated ergosterol for use in individual tests,retaining the National Institute’s material for reference.It is important that all such standard materials be kept attemperatures not exceeding 0° C.
504
THE PROPOSED UNIT.
It may be mentioned that the unit recommendedhas the same value, in terms of the weight of theoriginal ergosterol represented, as that used for someyears past in the Pharmaceutical Society’sLaboratories, on the basis of the provisional standardsolution supplied to them. The standard solutionnow recommended for general use cannot be assumedto be exactly equivalent to this, though comparativetests have not revealed a measurable difference.
Apart from such difference as may exist, the unitnow proposed, will, therefore, correspond to one ofthose already current. It is of convenient size, asrepresenting the order of dose required for dailyadministration to a rat, to produce healing of ricketsin a curative test ; that is to say, the daily dosesuitable for the conditions of a particular test willbe neither a large multiple nor a small fraction ofthe unit proposed.The Committee mention three biological methods
of estimating antirachitic effect, which they are ableto recommend. Their recommendations on thismatter are obviously not meant to be exclusive.They have mentioned the methods in which, fromexperience of their own investigations, they haveconfidence, but their statement is not intended topreclude the use of other biological methods thatalready exist, or of better ones that further investiga-tion may produce, provided that in every case thetest is based on a comparison with the fixed standard.
DISTRIBUTION OF THE STANDARD.
As indicated in another of the Committee’srecommendations, the quantity of the standard solu-tion available will not allow it to be circulated in
large quantities. Its distribution will, of necessity,be limited to those institutions which have the staffand equipment needed for its accurate applicationin biological assay, including proper facilities for its ipreservation without loss of activity. Unless thenumber of institutions requiring it prove to be muchgreater than that to which other standards are
distributed, it should be possible to supply samplesonce or twice a year to each regular recipient for someyears to come, before the preparation of a new batchof the standard solution will be required. It is
proposed to make the first issue about the end ofSeptember of this year, and a memorandum givingdetails for the use of the standard will be sent witheach first sample. It will greatly facilitate the
completion of arrangements if applications frominstitutions wishing to receive the standard are madeas early as possible, so that the Department may beable to estimate the number of separate samples likelyto be required for the first and subsequent issues.All such applications should be made to the Directorof the Department of Biological Standards, NationalInstitute for Medical Research, Hampstead,London, N.W. 3.
THE PRAISE OF HINDU CUSTOMS.
TN the June number of the Medical Review ofReviews, published in Calcutta, Dr. NaliniranjanSengupta takes us to task for some criticisms of Hinducustoms which appeared in an annotation on con-ditions of childbirth in India in THE LANCET lastyear. 1 The main propositions in our survey, however,have long been conceded by Indian reformers them-selves, and notably by Mr. Gandhi. We agree withDr. Sengupta that there has been a tendency recently
1 THE LANCET, 1929, i., 140.
to leave out the economic factor in discussing thisand similar questions, and that much of the ill-healthin India is due to poverty ; but when he proceeds toargue in favour of purdah and child marriage, onthe ground that they are safeguards against syphilis,and can apparently find nothing to criticise adverselyin the present customs, we may safely leave him tohis own more enlightened countrymen. There is,however, another side to Dr. Sengupta’s argument,and a very interesting one. He protests that at thepresent time it seems impossible for an Englishmanto find virtue in any Indian custom, and proceedsto give an account of certain time-honoured practices,based on rules laid down in the old Hindu scriptures,which are often admirable. Dr. Sengupta holds thatsome of the maligned social customs have actuallydone much to preserve the populat,ion under con-ditions which, if not mitigated in any way, mighteasily have proved fatal. Considering the chronicstate of semi-starvation in which many of the Indianpeople habitually live, he ascribes their comparativeimmunity from rickets in many parts of India tothe immemorial custom of rubbing infants with oiland exposing them at intervals to the sun’s rays.Similarly, he believes the low incidence of gastro-intestinal diseases in children, noted in a recent reportof the health officer of Calcutta, to be due to the veryancient practice, which used to be almost universal,of boiling the household milk immediately on receipt.It is true also that the ancient scriptures of theHindus inculcate habits of cleanliness which, howeverperfunctorily they may be kept up by many devoteesto-day, are far in advance of the practice of mostWestern races. The rules insist on the early morningbath, the habitual use of a fresh toothbrush (con-sisting of a twig of the neem tree), careful washingof the hands before and after meals, and ablutionsafter visits to the latrine. Specific rules are, moreover,laid down for periodic sexual abstention ; intercourseis forbidden during menstruation and pregnancy,for several weeks after parturition, and at variousother stated times. Dr. Sengupta justly adds thatthere is much that is admirable in the family life ofhis co-religionists, and quotes the late Sir GeorgeBirdwood, I.M.S., who praised the Mahratha Brahminladies as perfect daughters, wives, and mothers,whose virtues were deeply rooted in religious con-victions. Dr. Sengupta contends that certain factorsin the deterioration of the health of the Indian peopleare of European introduction. Syphilis, he says,comes from abroad, and is accordingly known asthe "feringi" (foreign) disease. The adulterationand impoverishment of foodstuffs is regarded as thedirect outcome of so-called civilising influences, themodern milling of rice, and blending of ghee withvegetable products having deprived the people oftheir vitamins. There is something to be said fromhis point of view, but when he blames Europe uncon-ditionally for malaria he is going too far. To statethat this disease was practically unknown in Indiabefore the ’sixties of the last century is a grotesqueexaggeration which leads one to suspect the accuracyof more reasonable statements. The fact thatmalaria has been spread by otherwise beneficentrailway and irrigation schemes is one of the tragedieswhich are liable to attend any efforts to improvecommunications and to safeguard the country againstfamines, but in working out plans for the future itought to be possible to minimise this danger. Dr.Sengupta is a staunch upholder of everything Indian,but like so much of the pro-Indian propaganda atthe present time, his article makes no attempt togive credit where credit is due.
ST. PETER’S HOSPITAL.-Last year 589 patientswere admitted to the wards of this Covent Garden hos-pital for stone and other urinary disesases. It is the onlyinstitution of the kind with a supply of radium. A longlease of premises nearly opposite the hospital in Henrietta-street has been acquired, and a pathological departmenthas been established there. The hospital itself is beingre-arranged and modernised.
