Izhar M, Shemesh M
Department of Hormone Research, Kimron Veterinary Institute, Bet Dagan, Israel.
J Reprod Fertil Suppl. 1989;37:37-44.
To determine whether luteotrophic activity is present in the bovine placental granules, fetal cotyledons from fetuses of 50-100 days of gestation were used. Enriched granules were prepared using a Percoll gradient. Active substances were obtained from the granules by freeze-thawing. The extracts thus obtained were then eluted on a Sephacryl S-300 column. The resultant fractions were then analysed by (1) a radioreceptor assay for hCG-like substances and (2) a bioassay using progesterone production by bovine luteal cells. There were two peaks of activity, one indicative of a high molecular weight substance and the second of a low molecular weight substance. Higher molecular weight substances were eliminated by using acidic extracts. The low molecular weight fraction was further analysed using reverse phase h.p.l.c. (acetonitrile:water gradient). The elution of this substance at 45% acetonitrile resulted in a 100-fold increase in luteotrophic activity in the bioassay compared to the Sephacryl fraction. The small molecular weight substance is heat-stable and not extracted to the organic phase when partitioned between methanol and chloroform.
为了确定牛胎盘颗粒中是否存在促黄体活性，使用了妊娠50 - 100天胎儿的胎儿绒毛叶。使用Percoll梯度制备富集颗粒。通过冻融从颗粒中获得活性物质。然后将如此获得的提取物在Sephacryl S - 300柱上洗脱。然后通过(1)针对hCG样物质的放射受体测定法和(2)使用牛黄体细胞产生孕酮的生物测定法对所得级分进行分析。有两个活性峰，一个表明是高分子量物质，另一个是低分子量物质。通过使用酸性提取物去除高分子量物质。使用反相高效液相色谱法（乙腈：水梯度）对低分子量级分进行进一步分析。与Sephacryl级分相比，该物质在45%乙腈处洗脱时，生物测定中的促黄体活性增加了100倍。该小分子物质对热稳定，在甲醇和氯仿之间分配时不被萃取到有机相中。
