Qu Ting, Gao Hui-Min, Chen Liang-Mian, Wang Zhi-Min, Zhang Qi-Wei, Cheng Yi-Yu
Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.
Zhongguo Zhong Yao Za Zhi. 2012 Oct;37(20):3086-91.
To kinds of establish a HPLC method for determining contents of indole alkaloids and bufadienolides contained in toad medicines, and analyze two kinds of components contained in toad venom, toad skin and toad periostracum.
As for alkaloids, Nucleosil C18 column was adopted with acetonitrile and water containing 0.5% potassium dihydrogen phosphate (6: 94, adjust pH to 3.2 with phosphate acid) as the mobile phase. The flow rate was 0.8 mL x min(-1), the detection wavelength was 275 nm, and the column temperature was 30 degrees C. As for bufadienolides, Alltima C18 column was adopted with acetonitrile and water containing 0.3% acetic acid (B) as the mobile phase. The gradient process was as follows: a linear gradient from 28% to 54% acetonitrile in the first 15 min, then kept at 54% for additional 20 min. The flow rate was 0.6 mL x min(-1), the detection wavelength was 296 nm, and the column temperature was 30 degrees C.
The linear ranges were 0.079 6-0.796 microg for serotonin, 0.097 2-1.945 microg for N-methylserotonin, 0.074 4-0.744 microg for N,N-dimethylserotonin, 0.103-2.05 microg for N,N,N-trimethylserotonin, and 0.067 2-0.672 microg for bufothionine, respectively. The average recoveries of serotonin and N-methylserotonin were 98.6% and 91.3%, respectively. The linear ranges of gamabufotalin, bufotalin, bufalin, cinobufagin and resibufogenin were 0.004 83-0.614, 0.007 9-1.006, 0.007 95-1.016, 0.009 7-1.24 and 0.009 6-1.22 microg, respectively, and their average recoveries were 101.6%, 102.5%, 101.0%, 99.1% and 98.9%, respectively.
Toad venom has the highest contents of indole alkaloids and bufadienolides, followed by toad skin, and toad periostracum showed the lowest contents and even no detection result.
建立高效液相色谱法测定蟾蜍类药材中吲哚生物碱和蟾毒配基的含量，并分析蟾酥、蟾皮及蟾衣中所含的这两类成分。
生物碱测定采用Nucleosil C18柱，以乙腈和含0.5%磷酸二氢钾的水（6∶94，用磷酸调pH至3.2）为流动相，流速为0.8 mL·min-1，检测波长为275 nm，柱温为30℃。蟾毒配基测定采用Alltima C18柱，以乙腈和含0.3%乙酸的水（B）为流动相，梯度洗脱程序为：前15 min乙腈由28%线性梯度至54%，然后在54%保持20 min，流速为0.6 mL·min-1，检测波长为296 nm，柱温为30℃。
血清素的线性范围为0.079 6～0.796 μg，N-甲基血清素为0.097 2～1.945 μg，N，N-二甲基血清素为0.074 4～0.744 μg，N，N，N-三甲基血清素为0.103～2.05 μg，蟾毒硫堇为0.067 2～0.672 μg；血清素和N-甲基血清素的平均回收率分别为98.6%和91.3%。脂蟾毒配基、蟾毒灵、蟾毒配基、华蟾酥毒基和脂蟾毒配质的线性范围分别为0.004 83～0.614、0.007 9～1.006、0.007 95～1.016、0.009 7～1.24和0.009 6～1.22 μg，平均回收率分别为101.6%、102.5%、101.0%、99.1%和98.9%。
蟾酥中吲哚生物碱和蟾毒配基含量最高，其次为蟾皮，蟾衣中含量最低，甚至未检出。
