Evaluation of cell disruption methods for protein and coenzyme Q10 quantification in purple non-sulfur bacteria.

Abstract

A recent focus has been on the recovery of single-cell protein and other nutritionally valuable bioproducts, such as Coenzyme Q10 (CoQ10) from purple non-sulfur bacteria (PNSB) biomass following wastewater treatment. However, due to PNSB's peculiar cell envelope (e.g., increased membrane cross-section for energy transduction) and relatively smaller cell size compared to well-studied microbial protein sources like yeast and microalgae, the effectiveness of common cell disruption methods for protein quantification from PNSB may differ. Thus, this study examines the efficiency of selected chemical (NaOH and EDTA), mechanical (homogenization and bead milling), physical (thermal and bath/probe sonication), and combined chemical-mechanical/physical treatment techniques on the PNSB cell lysis. PNSB biomass was recovered from the treatment of gas-to-liquid process water. Biomass protein and CoQ10 contents were quantified based on extraction efficiency. Considering single-treatment techniques, bead milling resulted in the best protein yields (p < 0.001), with the other techniques resulting in poor yields. However, the NaOH-assisted sonication (combined chemical/physical treatment technique) resulted in similar protein recovery (p = 1.00) with bead milling, with the former having a better amino acid profile. For example, close to 50% of the amino acids, such as sensitive ones like tryptophan, threonine, cystine, and methionine, were detected in higher concentrations in NaOH-assisted sonication (>10% relative difference) compared to bead-milling due to its less disruptive nature and improved solubility of amino acids in alkaline conditions. Overall, PNSB required more intensive protein extraction techniques than were reported to be effective on other single-cell organisms. NaOH was the preferred chemical for chemical-aided mechanical/physical extraction as EDTA was observed to interfere with the Lowry protein kit, resulting in significantly lower concentrations. However, EDTA was the preferred chemical agent for CoQ10 extraction and quantification. CoQ10 extraction efficiency was also suspected to be adversely influenced by pH and temperature.