Optimised “green solvent” extraction of long-chain menaquinones (Vitamin K2) from wet Lactococcus lactis biomass

Abstract

Menaquinone (vitamin K2) is an essential fat-soluble vitamin for bone and cardiovascular health. Long-chain menaquinones are exclusively synthesised by bacteria, and Lactococcus lactis appears promising as a microbial production platform. Menaquinone exists in the bacterial cytoplasmic membrane protected by a thick cell wall, thus requiring efficient cell lysis and extraction for its recovery. In this work, several potential solvents that are ranked as “recommended” for pharmaceutical manufacture were screened for their effectiveness, with the aid of external heating, to simultaneously disrupt cells and extract menaquinone from wet biomass. These results were compared with those obtained by traditional two-step strategies and extraction approaches using dry biomass. Ethanol appeared to be the most effective extractant, and the technique was then optimised using response surface methodology. Under optimal conditions (75 °C, biomass concentration of 0.199 g mL−1 and extraction time of 36.8 min), the extraction yields for menaquinone-7 and menaquinone-8 were 6.73 ± 0.17 and 179.6 ± 3.59 µg g−1 dry cell weight, respectively. In addition, the use of ethanol as the extraction solvent coupled with microwave heating and bead-beating was investigated. Enhanced extraction was achieved using microwave-assisted extraction at an elevated temperature of 125 °C for 5 min, with a 1.15- and 1.2-fold increase in menaquinone-7 and menaquinone-8 yields, respectively. This work proposes the development of an integrated single-step process for the sustainable and efficient recovery of menaquinone from biomass, with the advantage of using ethanol as a green extraction solvent.