Optimize the measurement of Poly-hydroxy-alkanoates (PHA) in biomass

Abstract

Poly-hydroxy-alkanoate (PHA) is an intracellular polymer that can be used as an energy and carbon source by microorganisms. Measuring PHA is important for understanding the microbial metabolism of enhanced biological phosphorus removal (EBPR) and aerobic granular sludge (AGS) systems. There is a commonly used method to measure PHA, which is based on organic solvent extraction and gas chromatography (GC). However, there are different versions of the same method with different parameters, but the role of some of these parameters is unclear. When different types of biomass are analyzed, there is a requirement to understand the parameters and obtain an optimal protocol. In this study, the effect of various digestion times, different alcohols and organic solvents, and acid concentrations were tested to obtain the optimal protocol. The results showed that a minimum digestion time was required to get the maximum yield of PHA, and the time might differ when using different types of biomass. Methanol was shown to be better for GC separation than propanol. Using different organic solvents didn’t affect the final concentration, and an optimal acid concentration was required to determine by comparison. The GC temperature program optimization showed that lower oven temperature in GC is more beneficial for peak separation. From the analysis, it would be suggested to use methanol and chloroform for digestion and keep the digestion time for 24 hours.