![]() PAL catalyzes the conversion of l-phenylalanine to cinnamic acid, and 4CL catalyzes the subsequent conversion of cinnamic acid into CoA ester, which is reduced by CCR and CAD to cinnamyl alcohol. As shown in Additional file 1: Figure S1, four enzymes involves in the synthesis of cinnamyl alcohol from l-phenylalanine in this pathway, including phenylalanine ammonia-lyase (PAL, EC 4.3.1.5), 4-coumarate: CoA ligase (4CL, EC 6.2.1.12), cinnamoyl-CoA reductase (CCR, EC 1.1.1.44) and cinnamyl alcohol dehydrogenase (CAD, EC 1.1.1.195). Cinnamyl alcohol is also synthesized via the monolignol pathway in plants and the biotechnological production of natural cinnamyl alcohol has been reported by introducing the plant monolignol pathway into microbial strains. In addition, cinnamyl alcohol is also a versatile chemical applied in the synthesis of various valuable compounds, such as cinnamyl esters (which are flavor and fragrance agents), flunarizine (which is used for fungal infection treatment), Taxol (which is a cancer treatment drug) and cinnamyl glycosides (which enhance immune function). Cinnamyl alcohol not only is used in the food and cosmetic industries due to its sweet-spicy odor and cinnamon taste, but also demonstrates good anti-inflammatory and antimicrobial activities. Ĭinnamyl alcohol, a natural aromatic alcohol, is the skeleton of monolignols. Industrial biosynthesis offers a promising alternative as it allows for scalable production of natural monolignols from bioresources. However, most natural monolignols are found in low concentrations in plants, and their extraction is limited by plant growth and expensive downstream processing costs. They have received much attention with the development of market requirements. Monolignols, which are units of plant lignin, are important aromatic compounds that can be used to generate a wide range of high-value chemicals of commercial interest. Recently, due to the rapid depletion of fossil fuels and the increasing demand for aromatic compounds, the production of these compounds from plant resources has increased in interest. We developed an efficient one-pot two-step biosynthesis system for cinnamyl alcohol production, which opens up possibilities for the practical biosynthesis of natural cinnamyl alcohol at an industrial scale. This process also demonstrated robust performance when it was integrated with the production of cinnamic acid from l-phenylalanine. Up to 17.4 mM cinnamic acid in the aqueous phase was totally reduced to cinnamyl alcohol with a yield of 88.2%, and the synthesized cinnamyl alcohol was concentrated to 37.4 mM in the organic phase. With the use of a dibutyl phthalate/water biphasic system, not only was product inhibition removed, but also the simultaneous separation and concentration of cinnamyl alcohol was achieved. Thus, a biphasic system was proposed to overcome the inhibition of cinnamyl alcohol via in situ product removal. Severe product inhibition was found to be the key limiting factor for cinnamyl alcohol biosynthesis. The strain could convert cinnamic acid into cinnamyl alcohol without overexpressing alcohol dehydrogenase or aldo–keto reductase. Herein, we constructed a recombinant Escherichia coli BLCS coexpressing carboxylic acid reductase from Nocardia iowensis and phosphopantetheine transferase from Bacillus subtilis. It is therefore necessary to develop an efficient, green and sustainable biosynthesis method. At present, the preparation of cinnamyl alcohol depends on plant extraction and chemical synthesis, which have several drawbacks, including limited scalability, productivity and environmental impact. Cinnamyl alcohol is not only a kind of flavoring agent and fragrance, but also a versatile chemical applied in the production of various compounds.
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