**5. Conclusions**

*Pectins - Extraction, Purification, Characterization and Applications*

cost for producing flavonoids.

**4.2 Chemical synthesis of flavonoids**

for the mass production of flavonoids [3].

instability of an engineered biosystem itself [2, 108].

**4.3 Biosynthesis of flavonoids**

steps and plenty of time, labor, and organic solvents, which greatly increase the production cost. Moreover, different plant tissues often need to develop different approaches for processing, which makes the extraction more complicated and further increase the cost for the production of flavonoids. Therefore, this approach is not cost-effective, and it is crucial to develop alternative strategies to reduce the

Another approach for producing flavonoids is chemical synthesis. Basically, there are two strategies for chemical synthesis of flavones, that is, the chalcone route and the Baker-Venkataraman method [101]. Even though there are a few successful examples, chemical synthesis of flavonoids is often very complicated and involved in many steps [2]. It requires toxic reagents and extreme reaction conditions [3, 102]. Chiral synthesis and subsequent modifications further increase the difficulty of this approach in the production of flavonoids [3]. Moreover, the multistep chemical reactions often produce quite a number of intermediate products with a high similarity in structure, which further increases the difficulty in purification of the desired products. Therefore, chemical synthesis is not economically feasible

Since the biosynthetic pathway of flavonoids is largely elucidated in plants [20], other promising alternative strategies have been developed to produce these secondary compounds [2, 103–106]. One of these alternative strategies is to produce flavonoids in a microbial cell factory. It has been well known that *Escherichia coli* and *Saccharomyces cerevisiae* are the two most commonly used model organisms for the construction of a microbial cell factory. There are quite a few paradigms for the production of flavonoids using this strategy. For example, eriodictyol has been produced using l-tyrosine as a substrate in *E. coli* BL21(DE3) genetically modified by *TAL, 4CL, CHS, CHI, F3H*, and *F3'H* genes and the production can reach up to 107 mg/L by further introducing three other genes *acs*, *accBC*, and *dtsR1* to enhance the availability of malonyl-CoA [103]. Kaempferol has been produced in a microbial cell factory by introducing a *de novo* biosynthetic pathway into *S. cerevisiae*, and the biosynthesis has been further improved by introducing two more pathways to enhance the generation of acetyl-CoA and malonyl-CoA [107]. Obviously, this strategy circumvents some inherent disadvantages of traditional plant extraction and chemical synthesis. However, not all genetically modified microbes can produce desired products due to the well-known complexity of a microbial cell system, the incompatibility of artificially synthesized genetic elements in host cells, the growth inhibition of host cells by desired and intermediate products, and the

Recently, we have developed an *in vitro* platform to produce flavonoids by constructing a multienzyme synthetic system to convert naringenin into kaempferol in one pot [2]. After optimizing a series of reaction parameters, including the components and pH value of the buffer system, reaction temperature and time, and total amount and ratio of the enzymes, the production yield can reach up to 37.55 ± 1.62 mg/L within 40–50 min with a conversion rate of 55.89% ± 2.74% [2]. The advantages of this strategy are obvious. It is time- and labor-saving. The reaction conditions are easy to control accurately. Due to the clearness in the buffer components and the lack of complex physiological regulation as occurred in the microbial cell factory, it is possible to easily make further optimization in the future.

**152**

Pectins and flavonoids are two distinctive classes of bioactive secondary metabolites presented in the fruit peels and used in food industry. The flavonoids can be divided into six major subclasses, including the flavanols, flavones, flavonols, flavanones, anthocyanidins, and isoflavones, and their flavonoid biosynthetic pathway has been largely elucidated. These natural small compounds possess a wide range of health-beneficial properties and can be derived by traditional plant extraction via organic solvents, chemical synthesis, and biosynthesis by constructing a microbial cell factory or an *in vitro* multienzyme synthetic system.
