Study on the differential changes of primary and secondary metabolites in the flesh of green ripe jujube based on principal component analysis
Ziziphus jujuba var. spinosa belongs to the Rhamnaceae family and Ziziphus genus of woody plants. Its fruit pulp, seeds, and leaves have high medicinal value. The “Shennong Bencao Jing” records that the flesh of sour jujube can be used as medicine, which has the function of stopping bleeding and diarrhea, and is mainly used to treat symptoms such as bleeding and diarrhea. Compared to sour jujube kernels, sour jujube flesh not only has medicinal value but also higher nutritional value. Sour jujube flesh contains abundant vitamin C, soluble sugars, proteins, amino acids, small molecule organic acids, terpenes and saponins, flavonoids, alkaloids and other components. Among them, the content of triterpenoid saponins with sedative and hypnotic activities is higher than that of sour jujube kernels.
Jujube fruit pulp has pharmacological effects such as improving sleep, protecting the liver, anti-tumor, regulating immunity, and improving cardiovascular and cerebrovascular diseases, as well as antioxidant properties. It can be used to develop products such as sweet taste inhibition, antidepressant, liver protection, and antibacterial.
Previous studies have found that the types and contents of components in traditional Chinese medicinal materials undergo significant changes at different growth stages. The reason for this phenomenon may be due to the differences in light, water, and external stimuli during the growth process of medicinal plants or fruits, which in turn affect the synthesis of secondary metabolites in their bodies. Xiang et al. measured different growth stages of Hypericum perforatum and used PCA analysis to determine the optimal harvesting stage for Hypericum perforatum at different growth stages; Yan et al. detected the nutrient content of jujube fruits at different maturity stages and analyzed the changes in nutrient content during the ripening process. The results can systematically and comprehensively evaluate the quality of medicinal materials and provide scientific basis for their suitable harvesting period. Determining the types of components in medicinal herbs at different growth stages can determine their harvesting period, increase the content of the main active ingredients in the herbs, enhance their medicinal efficacy, and provide reference for the quality of medicinal herbs. Jujube fruit pulp is mainly used as a juice liquid beverage, and the types of components in jujube fruit pulp at different maturity levels also have a significant impact on the flavor of the beverage. Therefore, analyzing the differences in the composition types of jujube fruit pulp at different maturity stages is of great significance for component analysis, quality control, and harvesting period.
This study harvested ripe red fruit jujube and green fruit jujube. Using UPLC-Q-TOF-MS/MS technology to analyze the main component types of green and ripe jujube fruit pulp, and combining PCA principal component analysis method, calculate the differences in primary metabolites between the two green and ripe jujube fruit pulp, determine the content of differential primary metabolites and secondary metabolites, and finally analyze the biosynthetic pathway through the differences in metabolite content.

Analysis and validation of the biosynthetic pathways of flavonoids in sour jujube. According to the characterization of component “2.1”, five flavonoids were obtained, namely rutin, quercetin, hyperoside, catechin, and epicatechin. The synthesis of flavonoids is mainly based on L-phenylalanine and L-tyrosine as precursors, which are synthesized through different branching pathways (see Figure 6). By analyzing the metabolites in the flesh of green jujube and ripe jujube, it was found that there were differences in the content of L-phenylalanine and L-tyrosine between the two fruit pulp. The content of the two metabolites in ripe jujube fruit pulp was higher than that in green jujube fruit pulp. Through content determination, it was found that the content of rutin, hyperoside, and quercetin in ripe jujube fruit pulp was higher than that in green jujube fruit pulp, which further verified the significant changes in flavonoid components during the ripening process from green jujube to ripe jujube fruit pulp. However, the content of catechins and epicatechins decreased in the flesh of ripe jujube fruit, mainly because catechins and epicatechins were mainly distributed in the fruit peel. Epicatechins have a strong astringency, while catechins have both astringency and bitterness, which are the main substances causing fruit astringency. Therefore, when the fruit is immature, it contains more catechins and epicatechins. During the fruit ripening process, the content of catechins and epicatechins gradually decreases, and the content is the lowest when the fruit is fully ripe. Therefore, the content of catechins and epicatechins changes inversely with the content of other flavonoids during the transition from green jujube to ripe jujube. Due to the fact that catechins and epicatechins determine the bitter taste of the fruit, it indicates that they can provide scientific basis for the consumption of jujube flesh.

Analysis and validation of terpenoid components in sour jujube. According to the characterization of component “2.1”, four triterpenoids were obtained, namely jujube seed saponin B, ursolic acid, oleanolic acid, and betulinic acid. Through content determination, it was found that the content of ursolic acid and oleanolic acid in ripe jujube fruit flesh was lower than that in green jujube fruit flesh. The reason for this phenomenon is that these two metabolites contribute to plant defense. For example, triterpenoids have antifungal properties and can resist toxin production, which determines that the content of these two metabolites in green fruit is higher than that in ripe fruit.

In summary, the flavor changes of fruit are determined by the content of catechins and epicatechins. During the ripening process of jujube, catechins and epicatechins gradually decrease, but other flavonoid components show an overall upward trend, indicating that the sedative and hypnotic effects of ripe jujube may be stronger than those of green jujube; The content of triterpenoid components such as oleanolic acid and ursolic acid in the flesh of green jujube is higher than that in ripe jujube, indicating that some triterpenoid components in green jujube may have stronger pharmacological activity than ripe jujube. The above research methods and results can provide technical support and scientific basis for the analysis of components, quality evaluation, and harvesting period of jujube fruit pulp with different maturity levels.