Identification of bacteriochlorophylls in lakes via reverse‐phase HPLC

Abstract

Reverse‐phase HPLC has been used successfully to quantify phytoplankton pigments in lakes and oceans. Here we extend the technique to photosynthetic bacterioplankton and demonstrate the identification of five bacteriochlorophylls (BChl a, b, c, d, e) extracted from pure cultures with 90% acetone. The technique was then applied to natural plankton samples from two oligotrophic‐mesotrophic lakes in northern Wisconsin. In both lakes, previously undetected layers of phototrophic bacteria were identified based on their pigment compositions. In one of the lakes, the bacterial layer previously had been misidentified as a deep zone of abundant pheopigment. These observations re‐emphasize concerns that traditional protocols (i.e. measuring the absorbance of mixed acetone extracts at 665 nm before and after acidification) have serious limitations in natural systems and can lead to misinterpretations of planktonic distributions and processes. The potential importance of phototrophic bacteria in dimictic temperate lakes is demonstrated. Without significantly modifying standard reverse‐phase HPLC protocols, unamibiguous determinations of eucaryotic and procaryotic chlorophylls and degradation products can be made simultaneously.