Abstract
Recently, leaf protein concentrate (LPC) has gained increased attention in response to the constantly growing protein demand. Green biorefineries can become more economical by valorizing their by-products and reducing environmental risks. The current study describes the variations in the antioxidant capacity and phytochemical composition of a liquid by-product (referred to as brown juice (BJ)) obtained during the extraction of leaf protein concentrate (LPC) from the fresh biomass of alfalfa (Medicago sativa L.). Four varieties of alfalfa were investigated during three harvest times, i.e., August 2017 (first harvest), September 2017 (second harvest), and June 2018 (third harvest). Also, the fresh BJ was lacto-fermented to extend its preservation period but also modifying its composition. The results of different general phytochemical composition analyses and antioxidant assays revealed similar tendencies across different alfalfa varieties and harvest times. Most of the phytochemicals in the BJ identified by HPLC-MS/MS can be classified as flavonoids/flavonoid derivatives, e.g., apigenin, naringenin, luteolin, formononetin. Substantially, the lacto-fermentation process induced a switch into aglycones, e.g., apigenin content increased by an order of magnitude, while apigenin-7-O-glucuronide content was halved after lacto-fermentation. Additionally, several B vitamins were detected, including B2, B3, and B7. These results could provide a basis for various ways of industrial valorization but need to be strengthened by data generated from large-scale production.