Mechanism Of Quality Formation Research Articles

Transcriptome and metabolome reveal the molecular mechanism of quality formation in different planting areas of Ziziphus jujuba Mill. ‘Jingudazao’ fruit

Both environmental and genetic factors have important influences on the quality formation of jujube fruit. To elucidate the molecular mechanism of fruit quality formation with the effects of soil and meteorological factors, transcriptome and metabolome analysis was conducted on Ziziphus jujuba Mill. ‘Jingudazao’ (JG) fruits were harvested from the three areas, Hebei (HB), Shanxi (SX), and Xinjiang (XJ) at the half-red stage. Firstly, meteorological factors and soil conditions have significant effects on jujube fruit quality characteristics. In compared groups, 331, 295, and 297 differentially accumulated metabolites (DAMs) were identified, while the principal DAMs pertaining to carbohydrates, organic acids, amino acids, derivatives, etc. showed that JG_HB had fewer DAMs than JG_XJ and JG_SX. Among them, 20 differentially expressed genes (DEGs), 13 DAMs, and their derivatives participated in carbohydrates and acid metabolism. In addition, two DEGs and seven DAMs were involved in flavonoid biosynthesis. The results provide a valuable and theoretical foundation for the ecological planting and cultivation of jujube.

Changes in microbial diversity and volatile metabolites during the fermentation of Bulang pickled tea

The present study aimed to determine microbial community, short-chain fatty acids (SCFAs), and volatilome of Bulang pickled tea during fermentation. Sequencing of 16S rRNA and ITS revealed that Bualng pickled tea was dominated by Lactobacillus plantarum, unclassified Enterobacteriaceae, unclassified Debaryomyces, Candida metapsilosis, Cladosporium sphaerospermum, and unclassified Aspergillus. The overall contents of SCFAs increased, with acetic acid showing the highest content. A total of 398 differential volatile metabolites were detected using differential metabolomics analysis. Out of these different volatile compounds, ten key volatile compounds including (Z)-4-heptenal, 1-(2-thienyl)-ethanone, 5-methyl-(E)-2-hepten-4-one, 2-ethoxy-3-methylpyrazine, p-cresol, 2-methoxy-phenol, ethy-4-methylvalerate, 3-ethyl-phenol, p-menthene-8-thiol, and 2-s-butyl-3-methoxypyrazinewere were screened based on odor activity value (OAV). The Spearman correlation analysis showed a high correlation of SCFAs and volatile compounds with microorganisms, especially L. plantarum and C. sphaerospermum. This study provided a theoretical basis for elucidating the flavor quality formation mechanism of Bulang pickled tea.

UV-B Radiation Enhances Epimedium brevicornu Maxim. Quality by Improving the Leaf Structure and Increasing the Icaritin Content.

Epimedium brevicornu Maxim. is a herbal plant with various therapeutic effects, and its aboveground tissues contain flavonol compounds such as icaritin that can be used to produce new drugs for the treatment of advanced liver cancer. Previous studies have shown that ultraviolet-B (UV-B, 280-315 nm) stress can increase the levels of flavonoid substances in plants. In the current study, we observed the microstructure of E. brevicornu leaves after 0, 5, 10, 15, and 20 d of UV-B radiation (60 μw·cm-2) and quality formation mechanism of E. brevicornu leaves after 0, 10, and 20 d of UV-B radiation by LC‒ESI‒MS/MS. The contents of flavonols such as icariside I, wushanicaritin, icaritin, and kumatakenin were significantly upregulated after 10 d of radiation. The results indicated that UV-B radiation for 10 d inhibited the morphological development of E. brevicornu but increased the content of active medicinal components, providing a positive strategy for epimedium quality improvement.

Unveiling the quality formation mechanism of hand-stretched dried noodle based on typical flour and processing technology

This study examined the physicochemical properties of flour determining hand-stretched dried noodles (HSDN) and the mechanisms involved. The findings indicated that the flexural performance, cooking and eating quality of HSDN was primarily influenced by gluten characteristics. Additionally, porosity was significantly impacted by the addition of salt, while selecting flour with a higher glutenin to gliadin ratio and lower damaged starch content could potentially reduce the amount of salt required, contributing to the pore structure of HSDN. The HSDN obtained from long-term resting processes demonstrated superior cooking and eating quality in comparison to short-term ones. The extended resting period facilitated the expulsion of trapped air within the dough and the relaxation of the gluten network, resulting in improved dispersal and orientation of excessively aggregated gluten proteins during the stretching process. This led to the formation of more protein branches and stable β-sheet structures during subsequent resting periods.

Non-targeted metabolomic analysis reveals the mechanism of quality formation of citrus flower-green tea.

Citrus flower-green tea (CT) is a scented tea processed from green tea (GT) and fresh citrus flower, which is favored by consumers due to its potential health benefits and unique citrus flavor. This study evaluated the quality of CT and revealed the mechanism of its quality formation. The CT had a significant citrus flavor and a good antioxidant activity, and its sensory quality was superior to that of GT. Headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) analysis revealed that the scenting process resulted in a significant increase of alkenes such as β-pinene, trans-β-ocimene, α-farnesene, isoterpinolene, and γ-terpinene, as well as a significant decrease of alcohols such as α-terpineol, l-menthol, and linalool in CT in comparison with GT. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed that the levels of flavonoids (such as neohesperidin, hesperidin, tangeritin, hesperetin 5-O-glucoside, and nobiletin) and alkaloids (such as trigonelline and theobromine) in CT increased significantly after scenting process, while the levels of amino acids (such as valine and l-phenylalanine) and organic acids (such as ascorbic acid) decreased significantly. These observations showed that the scenting process promoted the absorption of aroma from citrus flowers by GT and the changes in its non-volatile metabolites, leading to the formation of citrus flavor quality in CT. © 2024 Society of Chemical Industry.

Unveiling the impact of harvest time on Dioscorea opposita Thunb. cv. Tiegun maturity by NMR-based metabolomics and LC-MS/MS analysis.

Dioscorea opposita Thunb. cv. Tiegun maturity (DM) is an important factor influencing its quality. However, there are few studies on the impact of harvest time on its maturation. In the present study, a NMR-based metabolomics approach was applied to investigate the dynamic metabolic changes of D. opposita Thunb. cv. Tiegun at six different harvest stages: stage 1 (S1), stage 2 (S2), Stage 3 (S3), stage 4 (S4), stage 5 (S5) and stage 6 (S6). Principal component analysis showed distinct segregation of samples obtained from S1, S2 and S3 compared to those derived from S4, S5 and S6. Interestingly, these samples from the two periods were obtained before and after frost, indicating that frost descent might be important for DM. Eight differential metabolites responsible for good separation of different groups were identified by the principal component analysis loading plot and partial least squares-discriminant analysis. In addition, quantitative analysis of these metabolites using liquid chromatography-tandem mass spectrometry determined the effects of harvest time on these metabolite contents, two of which, sucrose and allantoin, were considered as potential biomarkers to determine DM. The present study demonstrated that NMR-based metabolomics approach could serve as a powerful tool to identify differential metabolites during harvesting processes, also offering a fresh insight into understanding the DM and the potential mechanism of quality formation. © 2024 Society of Chemical Industry.

Identification and expression analysis of citrate synthase 3 gene family members in apple

As one of the key enzymes in cell metabolism, the activity of citrate synthase 3 (CS3) regulates the substance and energy metabolism of organisms. The protein members of CS3 family were identified from the whole genome of apple, and bioinformatics analysis was performed and expression patterns were analyzed to provide a theoretical basis for studying the potential function of CS3 gene in apple. BLASTp was used to identify members of the apple CS3 family based on the GDR database, and the basic information of CS3 protein sequence, subcellular localization, domain composition, phylogenetic relationship and chromosome localization were analyzed by Pfam, SMART, MEGA5.0, clustalx.exe, ExPASy Proteomics Server, MEGAX, SOPMA, MEME, WoLF PSORT and other software. The tissue expression and inducible expression characteristics of 6 CS3 genes in apple were determined by acid content and real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Apple CS3 gene family contains 6 members, and these CS3 proteins contain 473-608 amino acid residues, with isoelectric point distribution between 7.21 and 8.82. Subcellular localization results showed that CS3 protein was located in mitochondria and chloroplasts, respectively. Phylogenetic analysis divided them into 3 categories, and the number of genes in each subfamily was 2. Chromosome localization analysis showed that CS3 gene was distributed on different chromosomes of apple. The secondary structure of protein is mainly α-helix, followed by random curling, and the proportion of β-angle is the smallest. The 6 members were all expressed in different apple tissues. The overall expression trend from high to low was the highest relative expression content of MdCS3.4, followed by MdCS3.6, and the relative expression level of other members was in the order of MdCS3.3 > MdCS3.2 > MdCS3.1 > MdCS3.5. qRT-PCR results showed that MdCS3.1 and MdCS3.3 genes had the highest relative expression in the pulp of 'Chengji No. 1' with low acid content, and MdCS3.2 and MdCS3.3 genes in the pulp of 'Asda' with higher acid content had the highest relative expression. Therefore, in this study, the relative expression of CS3 gene in apple cultivars with different acid content in different apple varieties was detected, and its role in apple fruit acid synthesis was analyzed. The experimental results showed that the relative expression of CS3 gene in different apple varieties was different, which provided a reference for the subsequent study of the quality formation mechanism of apple.

Combined metabolome and transcriptome analyses of quality components and related molecular regulatory mechanisms during the ripening of Huangjin Peach

Huangjin Peach is highly nutritious with cinsiderable market potential. Flavonoids, sugars, and acids are among the key compounds affecting the quality of yellow peaches. In this study, we analyzed the metabolites and gene expression of Huangjin Peach to explain the quality formation mechanism during the ripening period (including the young fruit, color change, and mature stages). We explored the biosynthetic pathways of flavonoids, sugars, and organic acids through metabolomics (LC-MS/MS) and transcriptomics (RNA-seq) approaches. A total of 864 metabolites were identified during the ripening of Huangjin Peach. Flavonoids increased significantly mainly at the color change stage and accumulated at the mature stage. During the combined transcriptomic and metabolomic analyses, CHS, F3′H, DFR, and A3GT were identified as the main enzyme genes regulating flavonoid synthesis, as well as the key factors affecting fruit color. Sugar alcohols accumulated significantly in Huangjin Peach during the ripening period, and organic and phenolic acids decreased significantly during the color change stage, which affect the quality and taste of fruits. Most metabolites and genes changed dramatically during the color change stage, indicating that the color change stage is critical for fruit quality formation. The expression of key genes was verified by qRT-PCR. Our results explained some of the key fruit quality formation mechanisms of Huangjin Peach, which provide a valuable foundation for the improvement and development of functional foods.

Comparative Transcriptome and Metabolome Analyses Revealed Quality Difference between Beauty Tea Processed through Indoor Withering and Outdoor Solar Withering.

Withering is the first processing procedure of beauty tea, and there are few reports on the impact of withering methods on the quality of beauty tea and its regulatory mechanisms. Through comparison of fresh tea leaves (FT) with the leaves at Indoor natural Withering 18h (IWT-18) and Outdoor Solar Withering 6h (OWT-6), which were collected at the end of the two withering processes, 17282 and 13984 differentially expressed genes (DEGs) were screened and 267 and 154 differential metabolites (DMs) were identified, respectively. Coexpression network revealed that a large number of DEGs and DMs were enriched in phenylpropanoid, flavonoid, and adenosine Triphosphate binding (ABC) transporter pathways, and the number of DMs and DEGs in IWT-18 vs FT exceeded that in OWT-6 vs FT. Both withering methods promoted a significant increase in content of phenylalanine and upregulation of β-glucoside (BGLU) expression in the phenylpropanoid metabolism pathway. The five proanthocyanidins related to theaflavins in the flavonoid synthesis pathway was more significant accumulation in FT vs IWT-18 than in FT vs OWT-6. Meanwhile, both withering methods can affect the ABC transporter pathway to promote the accumulation of amino acids and their derivatives, but different withering methods affect different ABC transporter families. Outdoor withering with more severe abiotic stress has a greater impact on the ABCG family, while indoor withering has a more significant effect on the ABCC family. Sensory evaluation results showed that the dry tea of IWT-18 was slightly better than that of OWT-6 because of longer withering time and more thorough substance transformation. In conclusion, the formation of honey flavor in beauty tea may be closely related to the DEGs and DMs in these three pathways. Our research provides theoretical data support for further revealing the mechanism of quality formation during the withering process of beauty tea. This article is protected by copyright. All rights reserved.

Untargeted Metabolomics and Physicochemical Analysis Revealed the Quality Formation Mechanism in Fermented Milk Inoculated with Lactobacillus brevis and Kluyveromyces marxianus Isolated from Traditional Fermented Milk.

Traditional fermented milk from the western Sichuan plateau of China has a unique flavor and rich microbial diversity. This study explored the quality formation mechanism in fermented milk inoculated with Lactobacillus brevis NZ4 and Kluyveromyces marxianus SY11 (MFM), the dominant microorganisms isolated from traditional dairy products in western nan. The results indicated that MFM displayed better overall quality than the milk fermented with L. brevis NZ4 (LFM) and K. marxianus SY11 (KFM), respectively. MFM exhibited good sensory quality, more organic acid types, more free amino acids and esters, and moderate acidity and ethanol concentrations. Non-targeted metabolomics showed a total of 885 metabolites annotated in the samples, representing 204 differential metabolites between MFM and LFM and 163 between MFM and KFM. MFM displayed higher levels of N-acetyl-L-glutamic acid, cysteinyl serine, glaucarubin, and other substances. The differential metabolites were mainly enriched in pathways such as glycerophospholipid metabolism, arginine biosynthesis, and beta-alanine metabolism. This study speculated that L. brevis affected K. marxianus growth via its metabolites, while the mixed fermentation of these strains significantly changed the metabolism pathway of flavor-related substances, especially glycerophospholipid metabolism. Furthermore, mixed fermentation modified the flavor and quality of fermented milk by affecting cell growth and metabolic pathways.

Dynamic changes in the major chemical and volatile components during the “Ziyan” tea wine processing

Among all types of tea beverage, tea wine uniquely combines the flavor of tea and wine, and its consumption is increasing in recent years. “Ziyan” tea wine is a new type of fermented tea wine rich in anthocyanins. The information about the dynamic changes of major chemical and volatile components in its processing is scarce. This study aimed to reveal the changes in sugar, total acids, catechins, anthocyanins and volatile components during “Ziyan” tea wine processing. Results showed that the total sugar, anthocyanin, and total catechins contents of the fermented tea wine were dramatically decreased, while epigallocatechin and epigallocatechin gallate retention remained high and the contents of caffeine and amino acids were not significantly impacted. 27 volatile fragrance compounds were generated during the fermentation process. Most of the volatile compounds in the tea leaves remain in the tea wine. The unique sweet and fruity aroma of “Ziyan” tea wine is mainly due to phenylethanol, octanoic acid, citric acid, and phenylacetaldehyde, which contain rosin, alcohol, etc. Overall, the findings provide systematic insights into the quality formation mechanism of “Ziyan” tea wine, especially in terms of aroma.

RETRACTED: Study on the quality formation mechanism of Zao chili with enhanced fermentation by Lactipllantbacillus plantarum 5-1

Zao Chili (ZC) is a traditional fermented pepper, which plays an important role in Chinese cooking. The aim of this study was to elucidate the effect of Lactipllantbacillus plantarum 5-1 on the physicochemical properties, metabolite and microbiota profiling of ZC. The physicochemical factors changed regularly with the fermentation time. In the microbial communities, Lactobacillus, Weissella, Enterobacter, Gibberella, Fusarium, Zygosaccharomyces and Pichia were the dominant genera. 7 kinds of organic acids were detected in the whole fermentation process of ZC, but only 5 kinds changed significantly. Based on the OPLS-DA model with VIP>1 and ANOVA with P<0.05, 33 volatile flavor compounds with significant differences were screened out of 89. According to the redundancy analysis (RDA), fungi mainly contributed to soluble solids, while bacteria mainly contributed to pH. Lactobacillus, Weissella, Enterbacter and Zygosaccharomyces may be the potential flavor contributing microorganisms in the fermentation process of ZC by the Spearman correlation coefficient. A total of 11 main metabolic pathways were obtained by KEGG enrichment analysis of 89 volatile flavor compounds and 7 organic acids. Therefore, this study further enhanced our understanding of the flavor quality formation mechanism of Lactipllantbacillus plantarum in ZC, and providing a theoretical basis for improving the flavor quality of ZC.

Inter-specific differences of medicinal bioactive products are correlated with differential expressions of key enzyme genes in Scutellaria baicalensis and Scutellaria viscidula

Revealing the dynamic accumulation and synthesizing mechanism of medicinal bioactive products is critical for the evaluation and utilization of crops in medicinal industry. Huangqin ( Scutellaria baicalensis Georgi) and Nianmao-Huangqin ( Scutellaria viscidula Bge) are important medicinal crops containing flavonoids with good bioactivities. Previous studies have found significant differences of flavonoid contents between the two species. Nevertheless, the dynamic changes of bioactive products and related differential synthesizing mechanism remains unknown. In this study, an inter-specific comparative study was reported on the dynamic accumulation of eight bioactive products (scutellarin, scutellarein, baicalin , wogonoside, apigenin , baicalein, wogonin, and chrysin) and the expression of seven key enzyme genes in different organs (root, stem, and leaf) during different growth stages (May, June, July, August, September, and October) of S. baicalensis and S . viscidula . The results showed that the eight bioactive products were differentially accumulated in different growth stages for the two species. Specifically, the contents of five ingredients (except wogonoside, scutellarein and apigenin) in roots of S. baicalensis were significantly higher than those in S. viscidula in harvest stage (October). Meanwhile, the relative expression of seven key enzyme genes in baicalin synthesizing pathway showed significant spatio-temporal specificities. The expression of UBGAT in downstream baicalin synthesis was significantly higher in roots of S. baicalensis than that of S. viscidula ( P < 0.05). Moreover, there exist significant correlations between the content of bioactive products and the expression of key enzyme genes. The expression of upstream ( CLL-7 , FNSII-1 , FNSII-2 ) and downstream ( F6H and UBGAT ) key enzyme genes were significantly positively correlated with the contents of baicalin in both S. baicalensis and S. viscidula. On the contrary, the expression of GUS was significantly negatively correlated with the content of baicalin in the two species. Collectively, these results give clear evidences that the expression of key enzyme genes (e.g., FNSII-2 , GUS , and UBGAT ) in baicalin synthesis is probably the main reason for the difference of bioactive products, suggesting that S . viscidula can be used as an alternative to S. baicalensis in pharmaceutical industry. Furthermore, these findings will provide a theoretical basis for exploring the inter-specific differences of bioactive products in medicinal crops within the same genus, thereby revealing the molecular mechanism of quality formation of Chinese medicinal materials. • Eight bioactive products showed content difference between two Scutellaria crops. • Seven key genes were differentially expressed in S. baicalensis and S. viscidula . • Difference of product content was correlated with difference of gene expression. • S. viscidula can be used as alternative medicinal source to S. baicalensis . • Molecular mechanism of differential bioactive product contents was revealed.

Effects of Microbial Action and Moist-Heat Action on the Nonvolatile Components of Pu-Erh Tea, as Revealed by Metabolomics.

Microbial action and moist-heat action are crucial factors that influence the piling fermentation (PF) of Pu-erh tea. However, their effects on the quality of Pu-erh tea remain unclear. In this study, the effects of spontaneous PF (SPPF) and sterile PF (STPF) on the chemical profile of Pu-erh tea were investigated for the first time, and sun-dried green tea was used as a raw material to determine the factors contributing to the unique quality of Pu-erh tea. The results indicated that the SPPF-processed samples had a stale and mellow taste, whereas the STPF-processed samples had a sweet and mellow taste. Through metabolomics-based analysis, 21 potential markers of microbial action (including kaempferol, quercetin, and dulcitol) and 10 potential markers of moist-heat action (including ellagic acid, β-glucogallin, and ascorbic acid) were screened among 186 differential metabolites. Correlation analysis with taste revealed that metabolites upregulated by moist-heat and microbial action were the main factors contributing to the staler mellow taste of the SPPF-processed samples and the sweeter mellow taste of the STPF-processed samples. Kaempferol, quercetin, and ellagic acid were the main active substances formed under microbial action. This study provides new knowledge regarding the quality formation mechanism of Pu-erh tea.

Explore the interaction between root metabolism and rhizosphere microbiota during the growth of Angelica sinensis.

Angelica sinensis is a medicinal plant widely used to treat multiple diseases in Asia and Europe, which contains numerous active components with therapeutic value. The interaction between root and rhizosphere microorganisms is crucial for the growth and quality formation of medicinal plants. But the micro-plant-metabolite regulation patterns for A. sinensis remain largely undetermined. Here, we collected roots and rhizosphere soils fromA. sinensisin seedling stage (M) and picking stage (G), respectively cultivated for one year and two years, generated metabolite for roots, microbiota data for rhizospheres, and conducted a comprehensive analysis. Changes in metabolic and microbial communities ofA.sinensisover growth were distinct. The composition of rhizosphere microbes in G was dominated by proteobacteria, which had a strong correlation with the synthesis of organic acids, while in M was dominated by Actinobacteria, which had a strong correlation with the synthesis of phthalide and other organoheterocyclic compounds, flavonoids, amines, and fatty acid. Additionally, co-occurrence network analysis identified thatArthrobacterwas found to be strongly correlated with the accumulation of senkyunolide A and n-butylidenephthalide.JGI 0001001.H03was found to be strongly correlated with the accumulation of chlorogenic acid. Based on rhizosphere microorganisms, this study investigated the correlation between root metabolism and rhizosphere microbiota of A. sinensis at different growth stages in traditional geoherb region, which could provide references for exploring the quality formation mechanism of A. sinensis in the future.

Integrative transcriptomic, proteomic and metabolomic analysis reveals the dynamic regulation of secondary metabolism upon development of Prunella vulgaris L.

Prunella vulgaris L. (P. vulgaris, Labiatae) is a perennial medicinal and edible plant widely used in China, Korea, Japan and Europe. The reddish brown spica of P. vulgaris (Prunellae Spica), which is collected in summer, has been commonly used in traditional medicine and food industry, while it is also used with whole grass in Europe and Taiwan. To clarify the regulatory pathways and mechanism of quality formation in P. vulgaris, targeted metabolomic, transcriptomic, and proteomic analyses of Prunellae Spica samples from five consecutive developmental stages were carried out. The results showed that terpenoids were mainly synthesized in the maturity stage of Prunellae Spica, with the key enzymes and coding genes in downstream pathways being mainly expressed during ripening, while related enzymes in the upstream pathway showed the opposite pattern. Flavonoids mainly accumulated before ripening, with highly expressed pathway enzymes and coding genes. The accumulation of phenylpropanoids was relatively active throughout the development process. Rosmarinic acid (RA) and its synthetic intermediate products mainly accumulated via more active pathway enzymes and coding genes before ripening. The regulatory factors and metabolites related to RA synthesis were mainly enriched in phenylpropanoid biosynthesis, plant hormone signal transduction, plant pathogen interaction, oxidative phosphorylation, and endoplasmic reticulum protein processing pathways.

Pile-fermentation of dark tea: Conditions optimization and quality formation mechanism

This study aimed to optimize the dark tea pile-fermentation conditions through single factor and orthogonal experiments. The optimal conditions consisted of 38 g/100g moisture content of wet green tea, 45 °C of temperature, and 85 g/100g humidity. On this basis, the dynamic changes of flavor components during pile-fermentation were systematically explored by chromatography, mass spectrometry and other methods. During pile-fermentation, changes were observed in the internal ratio of the following various substances, with a decrease in the content of tea polyphenols (especially ester catechins), amino acids, and soluble sugars, as well as the formation of new substances of theaflavin, thearubigin, and theabrownin, thereby affecting the brew color, flavor, and taste of dark tea. After pile-fermentation, a gradual decrease was detected in the floral aroma components, such as phenethyl alcohol, linalool oxides, and linalool, in contrast to a gradual increase in the content of aged fragrance components, including methoxy aromatic compounds like 1,2,3- and 1,2,4-trimethoxybenzene. During pile-fermentation, the important aroma substances such as benzaldehyde, β-ionone, and dihydroactinidiolide showed an uptrend first and then a downtrend. The changes in the ratio of these substances contributed jointly to the aroma quality formation of dark tea.

Transcriptome analysis of walnut quality formation and color change mechanism of pellicle during walnut development

Walnuts (including those covered with a pellicle) are loved for their rich nutritional value. And the popular varieties of walnut cultivation are Juglans sigillata L. The pellicle (seed coat) of these walnut cultivars has different colors and has an indispensable influence on the walnut quality formation. However, there are few reports on the pellicle color and quality formation in different developmental stages of walnut (Juglans sigillata L.). Therefore, in this study, three walnut cultivars (F, Q, and T) with different pellicle colors were selected for transcriptome sequencing and physiological index analysis of the color and quality formation mechanisms at different development stages. The results showed that with the development of walnut fruit, the starch sucrose metabolism pathway in the pellicle was activated and promoted starch hydrolysis. Meanwhile, the expression levels of genes related to the alpha-linolenic acid metabolism pathway were significantly increased during walnut maturation, especially in F2. Some physiological indicators related to lipid oxidation were also detected and analyzed in this study, such as MDA, CAT, POD and DPPH. These results were similar to the expression patterns of corresponding regulatory genes in the RNA-Seq profile. In addition, lignin synthesis genes were up-regulated in the phenylpropanoid metabolic pathway, while key genes enriched in the flavonoid and anthocyanin synthesis pathways were down-regulated. The results were consistent with the results of total anthocyanins and flavonoid content detection during walnut development. Therefore, this experiment suggested that with the maturation of walnut pellicle, the gene expression in the phenylpropanoid metabolic pathway flowed to the branch of lignin synthesis, especially in the Q variety, resulting in lower flavonoid and anthocyanin content at the maturity stage than immature. This is also the main reason for the pale pellicle of the three walnut varieties after mature. The findings of this study showed that changes in the expression levels of regulating genes for lipid, starch, sugar, and flavonoid synthesis during walnut development influenced the accumulation of the related metabolite for walnut quality formation and pellicle color. The results of this experiment provided the molecular basis and reference for the breeding of high nutritional quality walnut varieties.

Screening of reference genes for quantitative real-time PCR in Artemisia argyi

Artemisia Argyi Folium, a traditional Chinese medicine of important medicinal and economic value, sees increasing demand in medicinal and moxibustion product market. Screening stable and reliable reference genes for quantitative real-time PCR(qRT-PCR) is a prerequisite for the analysis of gene expression in Artemisia argyi. In this study, eight commonly used reference genes, Actin, 18s, EF-1α, GAPDH, SAND, PAL, TUA, and TUB, from the transcriptome of A. argyi, were selected as candidate genes. The expression of each gene in different tissues(roots, stems, and leaves) of A. argyi and in leaves of A. argyi after treatment with methyl jasmonate(MeJA) for different time(0, 4, 8, 12 h) was detected by qRT-PCR. Then, geNorm, NormFinder, BestKeeper, ΔCT, and RefFinder were employed to evaluate their expression stability. The results demonstrated that Actin was the most stable reference gene in different tissues and in leaves treated with MeJA, and coming in the second was SAND. Furthermore, the expression of DXS and MCT which are involved in terpenoid backbone biosynthesis was detected in different tissues and after MeJA treatment. The results showed that the expression patterns of DXS and MCT in different tissues and under MeJA treatment calculated with Actin and SAND as internal reference genes were consistent, which validated the screening results. In conclusion, Actin is the most suitable reference gene for the analysis of gene expression in different tissues of A. argyi and after MeJA treatment. This study provides valuable information for gene expression analysis in A. argyi and lays a foundation for further research on molecular mechanism of quality formation of Artemisia Argyi Folium.

Quality Evaluation of Taxilli Herba from Different Hosts Based on Simultaneous Determination of Multiple Bioactive Constituents Combined with Multivariate Statistical Analysis.

Taxilli Herba (TAXH) is an important traditional Chinese medicine with a long history, dating from the Eastern Han Dynasty to the present times. However, the active constituents in it that parasitize different hosts vary, affecting its clinical efficacy. Given the complexity of the host origins, evaluating the quality of TAXH is critical to ensure the safety and effectiveness of clinical medication. In the present study, a quantitative method based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) was established, which simultaneously determined the content of 33 active constituents, including 12 flavonoids, 4 organic acids, 12 amino acids, and 5 nucleosides in 45 samples. Orthogonal partial least squares discriminant analysis (OPLS-DA) was employed to classify and distinguish between TAXH and its adulterants, Tolypanthi Herba (TOLH). A hierarchical clustering analysis (HCA) was conducted combined with a heatmap to visually observe the distribution regularity of 33 constituents in each sample. Furthermore, gray relational analysis (GRA) was applied to evaluate the quality of samples to get the optimal host. The results demonstrated that TAXH excelled TOLH in quality as a whole. The quality of TAXH parasitizing Morus alba was also better, while those that were parasitic on Cinnamomum camphora and Glyptostrobus pensilis had relatively poor quality. This study may provide comprehensive information that is necessary for quality control and supply a scientific basis for further exploring the quality formation mechanism of TAXH.