Assay of 18 amino acids in Chuanmingshen violaceum roots by pre-column derivative HPLC

Abstract

The roots of Chuanmingshen violaceum is a commonly used Chinese herb and food, which contains rich amino acids. However, the kinds and amounts of amino acids are variety in this herb among the geographical location and ecological environment. Therefore, this study firstly developed a new pre-column derived HPLC method to quantify the levels of 18 amino acids in Ch. violaceum roots. Then 24 Ch. violaceum samples were harvested from its main cultivating areas in Sichuan, China. These samples were divided into 4 producing areas based on their geographical sites. The 18 kinds of amino acids were quantified in these sample by the developed method. The differences of these amino acids were further analyzed among these herbal samples and the 4 producing areas by t-test and principal component analysis(PCA). The result indicated the peaks of the 18 kinds of amino acids were separated well in 70 min.The correlation coefficients between peak areas and concentration of these amino acids were more than 0.999 1(n=6). All of their recoveries were in the range of 97.38%-101.3%(n=6).Their detection limit was in the range of 0.003-0.379 μg·mL~(-1).It demonstrates that the developed HPLC method can accurately quantify the amounts of multi-amino acids in this herb. The results of t-test analysis showed the contents of histidine, cystine, leucine, valine, tryptophan, phenylalanine and threonine were significantly different(P<0.05) among the 4 producing areas. But the differences of other amino acids were not significant.The first five factors were extracted by PCA to calculate the comprehensive score. The order of comprehensive score for the 4 producing areas was B(0.603, n=10), C(0.206, n=3), A(-0.283, n=7) and D(-1.167, n=4). The total content of amino acids in Ch. violaceum collected in B producing area was largest(12.5 mg·g~(-1)). It is concluded the Ch. violaceum contains multi-kinds of amino acids. On the basis of amino acid amount, Langzhong city and Cangxi county in Sichuan province(producing area B) is the suitable areas for cultivating Ch. violaceum.