HPLC Rotamer Separation and Determination of an Extract Fraction of a Microbial Pigment

• Published in: The FASEB journal 2013-04, Vol.27 (S1)
• Main Authors: Greenfield, Nathan, Wu, Zhiqian, Nimmanapalli, Ramadevi, Wang, Desuo
• Format: Article
• Language: English
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fasebj.27.1_supplement.lb67

Abstract only A bioactive red pigment fraction was extracted from a marine bacterium. The preliminary experimental data suggested that the extracted fraction may belong to a member of prodigiosin family. An HPLC rotamer separation method was developed to define the chemical identity of the isolated bacterial material. The chromatogram of the pigment showed a peak absorption at 530 nm and a typical retention time 5.6 min, same as the prodigiosin standard, when analyzed with a C‐18 column and an isocratic mobile phase of acetonitrile:water (80:20, v/v, 1ml/min, pH=7). Upon lowering the pH value of the mobile phase to 3 with HCl, the HPLC chromatogram of the nominally purified pigment showed two distinct absorption peaks. The separation of the parent (α rotamer) and daughter (β rotamer) peaks was pH‐ and temperature‐dependent. At the optimal condition (5°C and pH=3), the size of the β peak is approximately 30% of the α peak. All these parameters of rotamer separation of the unknown microbial pigment are identical to that of prodigiosin. Therefore, we suggest to use this simple and practical rotamer separation method to determine if an unknown red pigment belongs to prodigiosin family.