We characterized the intact polar lipid (IPL) composition in the surface waters of 22 lakes from Minnesota and Iowa, ranging in trophic state between eutrophic and oligo-mesotrophic, to investigate the impact of trophic state on IPL composition. A high diversity of IPL classes was detected. Most IPL classes were detected in all lakes, but the eutrophic lakes contained a significantly higher relative abundance of lysophosphatidylcholine (PC) than the oligo-mesotrophic lakes, which in turn were characterized by significantly higher relative abundance of hydroxymethyltrimethyl-alanine/trimethyl-homoserine (DGTA/DGTS) betaines, ornithine lipids and the recently discovered trimethyl ornithine (TMO) lipids. The higher relative abundance of ornithines and TMOs may relate to a higher contribution of heterotrophic bacteria relative to phytoplankton while the higher abundance of the DGTA/DGTS betaines may relate to substitution by microorganisms of these non-P lipids for PC under P-stress, as has been observed in other environments. We also detected a variety of heterocyst glycolipids (HGs) derived from N 2 -fixing heterocystous Cyanobacteria in all lakes, suggesting the presence of these Cyanobacteria in the full range of trophic conditions. Correlation of HG abundance with environmental data showed that high productivity lakes have high HG abundances, while other distributional differences in HGs, which did not correlate with environmental parameters, are likely due to differences in species composition. We conclude that the significant differences in IPL composition between the eutrophic and oligo-mesotrophic lakes are either due to adaptation of the membrane composition to nutrient conditions or due to general divergences in microbial composition under the different conditions.