# Improved Functions of Fermented Coffee by Lactic Acid Bacteria
## 4 Discussion
However, one little clue on why CGA is increased in fermented coffee is that CGA is formed by the ester linkage of quinic acid and caffeic acid via possible esterification of the esterase enzyme in our selected strains [38]. It has been reported in previous studies that fermentation increases the total polyphenol content of fermented coffee beans because phenolic compounds are naturally coupled with sugar, which decreases their availability to the organism. During fermentation, the starter organism's proteolytic enzyme hydrolyzes phenolic complexes into simple, soluble-free phenols and biologically more active forms that are easily absorbed [21]. Chlorogenic acid (CGA) is a polyphenol that coffee contains, and it has proven to be an antimicrobial and antioxidative agent [39]. Therefore,the increased CGA content in fermented coffee extract might influence the consumer's health due to its antimicrobial, anti-inflammatory, and antioxidative potential. From several in vitro, animal, and human studies, it is somehow evident that CGA consumption can improve consumers' health due to its anti-inflammatory and anti-oxidative activity against various degenerative and non-degenerative diseases [38]. The phenolic compounds' antibacterial mechanism of action is complicated. They can exert antibacterial activity by altering the cytoplasmatic membrane structure, affecting the proton motive force, electron movement, and active transport. Furthermore, the mechanism of phenolic toxicity may include inhibition by oxidized chemicals, either by reactivity with sulfhydryl groups or more non-specific interactions with proteins [40]. A recent study revealed that CGA may exhibit its antibacterial effects through multiple mechanisms, including the downregulation of ribosomal subunits, modulation of lipid metabolism, and scavenging of intracellular ROS [41]. The increased antioxidant activity is also linked with the increased production of chlorogenic acid; however, during the fermentation process, lactic acid bacteria also secrete secondary metabolites, which are also oxidative agents, such as vitamin C and Vitamin B. The caffeine content of the fermented coffee extract decreased with the increase in fermentation time in our experiment. It was noted that the caffeine content of the fermented coffee extract showed an inverse relation with the fermentation time. After 12 h of fermentation, the caffeine content decreased while at the start of fermentation, its concentration was high. The decreased content of caffeine in the fermented coffee extract is related to the growth of bacteria involved in fermentation because they can use caffeine for their metabolism [42]. Previous studies suggest that LAB degrade caffeine into dimethylxanthine and then methylxanthine with the help of the caffeine demethylylase enzyme by removing the methyl group at N-x terminals. There are three types of dimethylxanthine, i.e., paraxanthine, theobromine, and theophylline. Food microorganisms mostly transform caffeine into paraxanthine via demethylation of a caffeine methyl group at N-3 and the paraxanthine is further demethylated at N-7 and produces 1-methylxanthine [43]. The LAB-degrading potential of caffeine into dimethylxanthine could be due to the toxicity of caffeine. One study suggests that paraxanthine toxicity is lower in animal caffeine models [44]. It could be assumed that low caffeine content is due to the degradation of caffeine into low toxic compounds as caffeine possesses some antimicrobial effects and these antimicrobial effects are reduced through the demethylation of caffeine by bacteria metabolism.