Comparison of HPLC Methods for Determination of Anthocyanins and Anthocyanidins in Bilberry Extracts

An HPLC method and an acid hydrolysis HPLC method for the analysis of anthocyanins and
anthocyanidins in bilberry extracts have been developed. The HPLC method coupled with a mass
detector has identified 11 anthocyanins in bilberry extracts. The method provides anthocyanin profiles
that are very useful in verifying the identity of botanical raw materials, monitoring the consistency of
the raw material source, and quantitating the total anthocyanins. The acid hydrolysis HPLC method
greatly simplifies the anthocyanin profile in bilberry samples and converts anthocyanins to five major
anthocyanidin aglycones:  delphinidin, cyanidin, petunidin, peonidin, and malvidin. Each of these
aglycones can be separated completely and quantitated accurately with external standards. Various
extraction and hydrolysis conditions were investigated, and the advantages and disadvantages of
the HPLC and acid hydrolysis methods are discussed.
Keywords: HPLC; HPLC-MS; anthocyanin; anthocyanidin; bilberry extract

本研究开发了两种可用于分析越橘提取物中花青素与花色素的方法，分别为高效液相色谱法（High Performance Liquid Chromatography，HPLC）与酸水解-高效液相色谱法。该联用质谱检测器的高效液相色谱法已在越橘提取物中鉴定出11种花青素。该方法可获得花青素谱图，在验证植物源性原料的真实性、监控原料来源的一致性以及定量总花青素含量方面具备重要应用价值。酸水解-高效液相色谱法则可极大简化越橘样品中的花青素谱图，将花青素转化为五种主要花色素苷元：飞燕草素（delphinidin）、矢车菊素（cyanidin）、矮牵牛素（petunidin）、芍药素（peonidin）与锦葵素（malvidin）。上述每一种苷元均可通过外标法实现完全分离与准确定量。本研究考察了多种提取与水解条件，并对高效液相色谱法及酸水解-高效液相色谱法的优缺点进行了讨论。关键词：高效液相色谱法；高效液相色谱-质谱联用法；花青素；花色素；越橘提取物