Effect of different forms of alkali treatment on specific fermentation inhibitors and on the fermentability of lignocellulose hydrolysates for production of fuel ethanol.

Treatment with alkali, particularly overliming, has been widely used as a method for the detoxification of lignocellulose hydrolysates prior to ethanolic fermentation. However, the mechanisms behind the detoxification effect and the influence of the choice of cation have not been well understood. In this study, a dilute acid hydrolysate of spruce and an inhibitor cocktail consisting of six known inhibitors were used to investigate different alkali detoxification methods. The various treatments included the addition of calcium hydroxide, sodium hydroxide, potassium hydroxide, and ammonia to pH 10.0 and subsequent adjustment of the pH to 5.5 with either sulfuric or hydrochloric acid as well as treatment with the corresponding amounts of calcium, sodium, and potassium as sulfate or chloride salts at pH 5.5. An RP-HPLC method was developed for the separation of 18 different inhibitors in the hydrolysate, including furaldehydes and phenolics. Detection and quantification were carried out by means of UV, DAD, and ESI-MS in negative mode. Treatment of the spruce hydrolysate with alkali resulted in up to approximately 40% decrease in the concentration of furaldehydes. The effects on the aromatic compounds were complex. Furthermore, SFE was performed on the precipitate formed during alkali treatment to evaluate the inhibitor content of the precipitate, and the following RP-HPLC analysis implied that potential inhibitors were removed mainly through conversion rather than through filtration of precipitate. Parallel experiments in which sulfuric acid or hydrochloric acid was used for acidification to pH 5.5 after alkali treatment indicated that the choice of anion did not affect the removal of inhibitors. Detoxification with calcium hydroxide and ammonia resulted in better fermentability using Saccharomyces cerevisiae than detoxification with sodium hydroxide. The results from the experiments with the inhibitor cocktail indicated that the positive effects of alkali treatment are difficult to explain by removal of the inhibitors only and that possible stimulatory effects on the fermenting organism warrant further attention.