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題名 | 應用高效液相層析法檢測牛尿中孕酮之可行性探討=Quantitation of Bovine Urinary Progesterone by Using High Performane Liquid Chromatography |
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作者 | 楊健仁; 張維正; 李善男; 董Θ霙; 林仁壽; 吳兩新; Yang, C. J.; Chang, W. J.; Lee, S. N.; Dong, Y. Y.; Lin, J. H.; Wu, L. S.; |
期刊 | 動物園學報 |
出版日期 | 19980600 |
卷期 | 10 1998.06[民87.06] |
頁次 | 頁85-90 |
分類號 | 437.22 |
語文 | chi |
關鍵詞 | 牛尿; 孕酮; 高效液相層析法; Cow urine; Progesterone; HPLC; |
中文摘要 | 本實驗乃嘗試利用高效液相層析法檢測牛尿中游離孕酮。其方法為以C18萃取管 分離出尿中脂溶性物質,經40﹪甲醇預洗,消除尿中膽色素及其他可能的干擾鹽類。於C□ 分離管、移動相CH□CN:MeOH:H□O(1:1:1,v╱v╱v)及254 nm紫外光波長下, 在15.4分鐘時沖提出孕酮。本法之靈敏度達0.25 ng╱m1,而添加5 ng╱ml 及10 ng╱ml 孕 酮標準品,測得之回收率分別為70.3﹪及74.6﹪。 由酵素免疫分析法檢測經乙醚萃取之動情週期乳牛尿孕酮量,其濃度為0.6 ng╱ml~1.0 ng╱ml,此值低於高效液相層析法所能測得之下限,故本法無法供檢測動情週期間乳牛尿 孕酮之用。 |
英文摘要 | The progesterone content in tissue fluids and excreta can be a reliable marker for monitoring ovulation and corpus luteal function in animals. The immunoassay, such as radio immunoassay (RIA) and enzyme immunoassay (EIA) with the high sensitivity is commonly used for detecting progesterone levels. However, the radioactive hazard caused by the former restricts its usage, and the latter is limited to a few laboratories. High performance liquid chromatograph (HPLC) method measured the absolute value of steroids may provide on alternative choice. A rapid, sensitive method of HPLC to measured free progesterone in bovine urine without complex preparing procedure was developed. One ml of urine sample was extracted by C18 cartridge, then 40% methanol was added to expel the interfered substances from urine. Progesterone in the eluent was determined by reverse phase HPLC. The results revealed that: retention time, 15.4 min; the sensitivity of urinary sample was 0.25 ng/20 ul with the condition of 254 nm uv wavelength, a mobile phase of acetonitrile /methanol/water (1:1:1,v/v/v) and a flow rate of 0.8 ml/min. and the recovery rates were 70.3%, 74.6% for 5 ng/ml P□ and 10 ng/ml P□ respectively. This HPLC system for urinary progesterone determination was unable to detect the urinary free progesterone of estrous dairy cow, due to the low concentrations. After ether extract urinary sample, the free progesterone were between 0.6 ng/ml~1.0 ng/ml, lower than the sensitivity of the HPLC system. |
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