Abstract:
Copper Storage in the Bovine Foetus W. J. PRYOR* SUMMARY Cu concentration of foetal organs are similar to those of the adult bovine with the exception of the liver and the spleen which are considerably greater. The liver Cu concentration of the foetus is independent of the age of the foetus within a range of 33-345 p.p.m. of Cu in the liver of dams. There is limited evidence to show that at lower dam levels, there is some correlation. Total foetal Cu is related to the age of the foetus, but appears also to be largely independent of liver Cu concentrations of the dam. INTRODUCTION Although there is aomprehensive information on the copper (Cu) content of the growing and adult bovine (Cunningham, 1931) and of factors influencing it (Underwood, 1956 and others), there is a paucity of such information for the bovine foetus. Cunningham records one analysis of an unaged foetus, whilst Gruzewska and Roussel (1936), using the benzoin oxime method of analysis, reported Cu concentrations of 90-180 p.p.m. D.W. in livers of foetuses aged 2-9 months old. Beneel (1933) obtained values of 50-80 p.p.m. D.W. for foetal livers 3-4 mo\nths old and reported 50-801 mgms. of Cu in bovine embryos, most of which were aged 3-4 months. Field (1957), and Allcroft and Uvarov (1959) have shown that treatment with Cu of cows with liver Cu concentra,tions of 5-60 p.p.m. resulted in increased liver storage in newborn calves. In certain areas it may be expected that this method of preventive treatment of Cu deficiency will be used fairly extensively. Information on Cu levels, of the foetus and factors influencing them appear necessary and are being investigated. This communication records some of this information so far obtained. MATERIALS AND ME' THODS (a) Source of Animal Material Entire bovine foetuses if weighing below a,pproximately 1500 gms. were collected in the uterus from an abattoirs together with part of the liver of the dam and transported to the laboratory where foetal organs were dissected out. When foetuses were heavier than 1500 gms. the viscera required were removed at the abattoirs, packed and transported with the adult liver as before. The crown-rump length (C.R.L.) of the foetus was measured in whichever place it was eviscerated. Weights of the smaller foetuses were also recorded but it was not possible to obtain breed or history of the cattle. (b) Material for Analysis The usual precautions to avoid Cu contamination were taken at all stages during the preparation of animal material for analysis. + Department of Animal Yeerongpilly, Queensland. Husbandry, University Veterinary School, 162 Samples were dried for at least 24 hours in an oven a,t 100�C. before digestion and analysis. ( c ) Copper Analysis Material was analysed for Cu concentration on the dry-matter basis by the method of Clare, Cunningham, and Perrin (1945)) using a calibrated Unicam SP.500 spectrophotometer. (a Total Copper Content This was determined in most cases by estimation. A technique devised for the small foetuses of the sheep and the pig where total Cu is determined by direct measurement was only practicable in the case of the smaller bovine foetuses. RESULTS of Cu in foetal organs The liver was established to be the main site of body copper. With the exception of the liver and spleen, values obtained from viscera, muscle, bone, skin, and brain were similar to those recorded for the adult bovine (Cunningham, 1931) and did not appear to vary appreciably with age. The mean Cu concentration of all foetal livers was approximately 2.7 times the adult mea,n . Mean foetal spleen values were a similar degree higher than published adult figures. (ii) The relationship between concentration of Cu in foetal liver and that in the adult liver at varying foetal ages using the C.R.L. as an index, is shown in Table 1. TABLE I. Between Foetal Age and Liver Cu Concentrations of Dam and Foetus. Number of Estimations Mean Liver Cu Concentration of Foetus (p.p.m. D.W.) Mean Liver .Cu Conocfenh-& t on (p.p.m. D.W.) ( i ) Concentration Relationship Foetus C.R.L. (mm. 1 40- 100 loo- 200 200- 300 300- 400 400- 500 500- 600 600- 700 700- 800 800- 900 900-1000 TOTAL ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... 3 12 12 5 IO 7 3 11 9 4 76 ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... 552 405 458 455 431 445 362 379 424 513 432 ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... ...... 189 202 202 173 184 177 43 94 97 118 161 Analyses of variance were carried out showing that for foetuses differences between C.R.L. groups were not significant (F = 1.3, P > 005). The analysis of variance for cows shows that differences between groups are highly significant (F = 3.36, P ,< 0.01). ` There is a correlation between foetal and dam liver levels of r = 0.2928 t 0.1256. This is significantly different at the 5% level from a correlation of zero. 163 If however, the four dams with liver Cu concentrations of less than 30 p.p.m. are eliminated the correlation between foetal and dam levels is r = 0.087 t 0.141 so that the correlation is not signif icantly different from zero. (iii) Total cu in the foetus at varying foetal weights is shown in Table II. The 2nd-5th values were obtained by direct measurement from f oetuses. TABLE II Total Cu in the Foetus and Percentage of Cu Contained in Liver at Differing Foetal Weights. Wt. of Foetus (gms) Total Cu in Foetus mgms. Estimated Percentage of Cu in Liver 11 130 161 308 478 5000 10000 19000 27000 .. . . ..* .. .. .. .. . .. .. .. . .. .. .. .. .. ....... ..... .._.... ..... ....... ....... ....... ....... ..... . ..I < .. . . . . ..I .050 .235 .298 .562 1.0 25.0 47.5 94.0 130.0 DISCUSSION ,........... e....'..... ,........... *.......... ,........... ,. . . . . . . . . . . W...'..... ,....' . . . . . I.. . . . . . . . . . 84 81 79 77 76 66 55 50 35 In the foetus Cu is concentrated in the liver as is the case with the adult bovine, though at a considerably higher level. Ryley and Harvey (1960)) have shown that newly-born calves had significantly greater liver Cu concentration than their dams, though with increasing age these liver levels fell. The figures obtained for total Cu in the foetus and liver Cu concentration are considerably higher than those reported by Bence (1933) and Gruzewska and Roussel (1936). There would appear a possibility that previous methods of estimation used by these earlier workers may have been less sensitive than that of Clare, Cunningham and Perrin (1945). Much higher values for flesh and some viscera obtained by Rusoff (1941) lead him to obtain a total Cu content of 236mgms. for a new-born calf weighing 18,361 grams, of which 12.3% of body copper was present in the liver. The fact that liver Cu concentration of the foetus is independent of that of the dam provided she has a Cu liver level of above 30 p.p.m may at first sight appear at variance with the findings of Field (1957) and Allcroft and Uvarov (1959). However, the results did suggest that with very low dam levels there is some influence on the foetus though the overall relationship is not as direct as one might have expected. It should, however, be noted that the use of a figure of 30 p.p.m. to arbitrarily divide normal and deficient states in the adult bovine cannot be entirely sustained. Data of the type reported by Mylrea (1958) has shown that very low levels can occur without any sign of clinical abnormality. 164 The influence of copper reducing diets on foetal levels is currently being studied on sheep to endeavour to clarify this aspect. ` The fact that the liver Cu concentration of the foetus is independent of the age of the foetus indicates that Cu shortage in the foetus commences at a, very early stage of foetal life, the concentration never reaching a later peak. This is contrary to the view expressed by Underwood (1956) that the maximum concentrations occur at or shortly after birth. Foetuses with a C.R.L. greater than approximately 800 mm. would on the data of Maneely (1952) be virtually full-term. Results in this paper tend to support the view of Allcroft and Uvarov (1959) and Van der Grift (1955) obtained with living cows that the amount of Cu in the liver of pregnant cows decreased during the last four months of pregnancy. This would be a' pproximately equivalent to foetal C.R.L.` of greater than 380 mms. s Total foetal Cu on the other hand is as might be expected, closely related to, the age of the foetus, but though only limited observations have been made on this point it appears that it too, is not dependent on the copper status' of the dam at least unless possibly she is deficient. ACKNOWLED' GEMENTS I am indebted to the University of Queensland for a research grant; to M. C. Knott for technical assistance; to B. Hart for assistance in obtaining abattoir material, and to J. James who kindly analysed the data. REFERENCES Allcroft, R., and Uvarov, 0. (1959) Vet. Rec. 71 : 797. Bence, J . , Die Rolle des Kupfers in der Blutbilding. Ztschr, f. Klin. Med. 126 : 143-151. 193. (Cited by L. L. Rusoff, Florida Agr, expt. sta. tech. Bull. No. 356 [I9411 p. 13.) Glare, Cunningham, I. J., and Perrin (1945) N.Z.J. Sci.Tech. 26 : 340. Cunningham, I. J. (1931) Biochem J., 26 : 1267. Field, H. I. (1957) Vet. Rec. 69 : 788. Gruzewska, Z., and G. Roussel (1936) Compt. Rend. Sot. Biol. 122 : 13-14. (Cited by L. L. Rusoff, Florida, Agr. expt. sta. tech. Bull. No. 356 [1941] p. 13.) Maneely, R. B. (1952) Vet. Rec. 64 : 509. Mglrea, P . J . (1958) Aust. J. Agric. Res. 9 : 375. Ryley, J., and Harvey, J. (1960) pers. commUnderwood, E. J. (1956) ' Trace Elements in Human and Animal Nutrition' Academic Press Inc. New York. Van der Grift, J. (1955) Diss. Utrecht. (Cited by Allcroft R., and Uvarov, 0 [1959] Vet. Rec. 71 : 797.) DISCUSSION J. Lee (S.A.) stated that in experiments with sheep the concentration of copper in foetal livers had paralleled liver levels in the dams using the method of analysis referred to in this paper. This suggested a species difference. 165 Dr. E. A. Campbell (Qld.) .-The fact that the foetal liver levels are largely unrelated to dam liver levels is not surprising as foetal copper is derived from the dam' blood serum which has not been s shown to be related to the dam' liver levels. s Dr. N. T. M. Yeates (N.S.W.) .-The mechanism of control of the levels of copper indicate that there is a mechanism for its release. Dr. J. Harvey (Qld.) reported that in observations at Rocklea low levels had been found in the dams but high levels in their calves at birth. He suggested there may be some factor which prevents the dam from storing copper but which does not interfere with the transfer from the dam to the foetus. Interference factors must be considered. Answer.-There was still much to be learned as the copper story appears to be becoming even more complex on closer examination. 166