AbstractThis study is an evaluation of liquid state NMR as a tool for analyzing the lipid composition of algal cultures used for biodiesel production. To demonstrate the viability of this approach, 13C NMR was used to analyze the lipid composition of intact cells of the algal species, Neochloris oleoabundans (UTEX #1185). Two cultures were used in this study. One culture was “healthy” and grown in conventional media, whereas the other culture was “nitrogen‐starved” and grown in media that lacked nitrate. Triglyceride was determined to be present in both cultures by comparing the algal NMR spectra with published chemical shifts for a wide range of lipids and with a spectrum obtained from a triglyceride standard (glyceryl trioleate). In addition, it is shown that (1) the signal‐to‐noise ratio of the ∼29.5 ppm methylene peak is indicative of the lipid content and (2) the nitrogen‐starved culture contained a greater lipid content than the healthy culture, as expected. Furthermore, the nitrogen‐starved culture produced spectra that primarily contained the characteristic peaks of triglyceride (at ∼61.8 and ∼68.9 ppm), whereas the healthy culture produced spectra that contained several additional peaks in the glycerol region, likely resulting from the presence of monoglyceride and diglyceride. Finally, potential interferences are evaluated (including the analysis of phospholipids via 31P NMR) to assess the specificity of the acquired spectra to triglyceride. These results indicate that NMR is a useful diagnostic tool for selectively identifying lipids in algae, with particular relevance to biodiesel production. Biotechnol. Bioeng. 2010;106: 573–583. © 2010 Wiley Periodicals, Inc.