Evolution and functional divergence of glycosyltransferase genes shaped the quality and cold tolerance of tea plants.

Plant glycosyltransferases (UGTs) play a key role in plant growth and metabolism. Here, we examined the evolutionary landscape among UGTs in 28 fully sequenced species from early algae to angiosperms. Our findings revealed a distinctive expansion and contraction of UGTs in the G and H groups in tea (Camellia sinensis), respectively. Whole-genome duplication and tandem duplication events jointly drove the massive expansion of UGTs, and the interplay of natural and artificial selection has resulted in marked functional divergence within the G group of the sinensis-type tea population. In Cluster II of group G, differences in substrate selection (e.g., Abscisic Acid) of the enzymes encoded by UGT genes led to their functional diversification, and these genes influence tolerance to abiotic stresses such as low temperature and drought via different modes of positive and negative regulation, respectively. UGTs in Cluster III of the G group have diverse aroma substrate preferences, which contributes a diverse aroma spectrum of the sinensis-type tea population. All Cluster III genes respond to low-temperature stress, whereas UGTs within Cluster III-1, shaped by artificial selection, are unresponsive to drought. This suggests that artificial selection of tea plants focused on improving quality and cold tolerance as primary targets.

(Z)-3-Hexenol integrates drought and cold stress signaling by activating abscisic acid glucosylation in tea plants.

Cold and drought stresses severely limit crop production and can occur simultaneously. Although some transcription factors and hormones have been characterized in plants subjected each stress, the role of metabolites, especially volatiles, in response to cold and drought stress exposure is rarely studied due to lack of suitable models. Here, we established a model for studying the role of volatiles in tea (Camellia sinensis) plants experiencing cold and drought stresses simultaneously. Using this model, we showed that volatiles induced by cold stress promote drought tolerance in tea plants by mediating reactive oxygen species and stomatal conductance. Needle trap microextraction combined with GC-MS identified the volatiles involved in the crosstalk and showed that cold-induced (Z)-3-hexenol improved the drought tolerance of tea plants. In addition, silencing C. sinensis alcohol dehydrogenase 2 (CsADH2) led to reduced (Z)-3-hexenol production and significantly reduced drought tolerance in response to simultaneous cold and drought stress. Transcriptome and metabolite analyses, together with plant hormone comparison and abscisic acid (ABA) biosynthesis pathway inhibition experiments, further confirmed the roles of ABA in (Z)-3-hexenol-induced drought tolerance of tea plants. (Z)-3-Hexenol application and gene silencing results supported the hypothesis that (Z)-3-hexenol plays a role in the integration of cold and drought tolerance by stimulating the dual-function glucosyltransferase UGT85A53, thereby altering ABA homeostasis in tea plants. Overall, we present a model for studying the roles of metabolites in plants under multiple stresses and reveal the roles of volatiles in integrating cold and drought stresses in plants.

UGT89AC1-mediated quercetin glucosylation is induced upon herbivore damage and enhances Camellia sinensis resistance to insect feeding.

Quercetin is a key flavonol in tea plants (Camellia sinensis (L.) O. Kuntze) with various health benefits, and it often occurs in the form of glucosides. The roles of quercetin and its glucosylated forms in plant defense are generally not well-studied, and remain unknown in the defense of tea. Here, we found higher contents of quercetin glucosides and a decline of the aglucone upon Ectropis grisescens (E. grisescens) infestation of tea. Nine UGTs were strongly induced, among which UGT89AC1 exhibited the highest activity toward quercetin in vitro and in vivo. The mass of E. grisescens larvae that fed on plants with repressed UGT89AC1 or varieties with lower levels of UGT89AC1 was significantly lower than that of larvae fed on controls. Artificial diet supplemented with quercetin glucoside also reduced the larval growth rate, whereas artificial diet supplemented with free quercetin had no significant effect on larval growth. UGT89AC1 was located in both the cytoplasm and nucleus, and its expression was modulated by JA, JA-ILE, and MeJA. These findings demonstrate that quercetin glucosylation serves a defensive role in tea against herbivory. Our results also provide novel insights into the ecological relevance of flavonoid glycosides under biotic stress in plants.

Genome-wide association analysis discovered new loci and candidate genes associated with low-phosphorus tolerance based on shoot mineral elements concentrations in soybean.

Phosphorus (P) deficiency is one of the major limitations for soybean production. Moreover, it has been well reported P and other mineral elements function interdependently or antagonistically to control nutrients homeostasis in plants. Thus, it is urgently needed to understand the genetic mechanism of the accumulation of mineral elements in response to low-P stress. In this study, to identify single nucleotide polymorphisms (SNPs) and candidate genes controlling the accumulation of mineral elements suffering low-P stress in seedling stage of soybean plants, we measured concentrations of mineral elements, including P, Zn, Fe, Mn, Mg and Ca, in shoots of 211 soybean accessions under normal phosphorus (+P) and low phosphorus (-P) conditions in two hydroponic experiments. And genome-wide association study (GWAS) using high density NJAU 355K SoySNP array and concentrations of five of these mineral elements except P was performed. A total of 36 SNPs distributed on 13 chromosomes were identified to be significantly associated with low-P tolerance, and nine SNPs on chromosome 10 formed a SNP cluster. Meanwhile, the candidate gene GmFeB1 was found to serve as a negative regulator element involved in soybean P metabolism and the haplotype1 (Hap1) of GmFeB1 showed significantly higher shoot Fe concentration under -P condition than that of Hap2. In summary, we uncover 36 SNPs significantly associated with shoot mineral elements concentrations under different P conditions and a soybean low-P related gene GmFeB1, which will provide additional genetic information for soybean low-P tolerance and new gene resources for P-efficient soybean varieties breeding.

Functional study and elite haplotype identification of soybean phosphate starvation response transcription factors GmPHR14 and GmPHR32.

Phosphorus (P) is one of the important mineral elements required for plant growth and development. However, because of the low mobility in soil, P deficiency has been an important factor limiting soybean production. Here, we identified 14 PHR (phosphate starvation response) genes in soybean genome and verified that two previously unreported GmPHR members, GmPHR14 and GmPHR32, were involved in low-P stress tolerance in soybean. GmPHR14 and GmPHR32 were present in two diverged branches of the phylogenic tree. Both genes were highly expressed in roots and root nodules and were induced by P deficiency. GmPHR14 and GmPHR32 both were expressed in the nucleus. The 211 amino acids in the N terminus of GmPHR32 were found to be required for the transcriptional activity. Overexpressing GmPHR14 or GmPHR32 in soybean hairy roots significantly increased roots and shoots dry weight under low-P condition, and overexpressing GmPHR14 additionally significantly increased roots P concentration under low-P condition. GmPHR14 and GmPHR32 were polymorphic in soybean population and the elite haplotype2 (Hap2) for both genes was preferentially present in improved cultivars and showed significantly higher shoots dry weight under low-P condition than the other two haplotypes. These results suggested GmPHR14 and GmPHR32 both positively regulated low-P responses in soybean, and would shed light on the molecular mechanism of low-P stress tolerance. Furthermore, the identified elite haplotypes would be useful in P-efficient soybean breeding. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01301-z.

Herbivore-induced DMNT catalyzed by CYP82D47 plays an important role in the induction of JA-dependent herbivore resistance of neighboring tea plants.

Herbivore-induced plant volatiles play important ecological roles in defense against stresses. However, if and which volatile(s) are involved in the plant-plant communication in response to herbivorous insects in tea plants remains unknown. Here, plant-plant communication experiments confirm that volatiles emitted from insects-attacked tea plants can trigger plant resistance and reduce the risk of herbivore damage by inducing jasmonic acid (JA) accumulation in neighboring plants. The emission of six compounds was significantly induced by geometrid Ectropis obliqua, one of the most common pests of the tea plant in China. Among them, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) could induce the accumulation of JA and thus promotes the resistance of neighboring intact plants to herbivorous insects. CsCYP82D47 was identified for the first time as a P450 enzyme, which catalyzes the final step in the biosynthesis of DMNT from (E)-nerolidol. Down-regulation of CsCYP82D47 in tea plants resulted in a reduced accumulation of DMNT and significantly reduced the release of DMNT in response to the feeding of herbivorous insects. The first evidence for plant-plant communication in response to herbivores in tea plants will help to understand how plants respond to volatile cues in response to herbivores and provide new insight into the role(s) of DMNT in tea plants.

Herbivore-induced volatiles influence moth preference by increasing the ß-Ocimene emission of neighbouring tea plants.

Herbivore-induced plant volatiles prime neighbouring plants to respond more strongly to subsequent attacks. However, the key volatiles that trigger this state and their priming mechanisms remain largely unknown. The tea geometrid Ectropis obliqua is one of the most devastating leaf-feeding pests of tea plants. Here, plant-plant communication experiments demonstrated that volatiles emitted from tea plants infested by E. obliqua larvae triggered neighbouring plants to release volatiles that repel E. obliqua adult, especially mated females. Volatile analyses revealed that the quantity of eight volatiles increased dramatically when plants were exposed to volatiles emitted by infested tea plants, including (Z)-3-hexenol, linalool, α-farnesene, ß-Ocimene and (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT). The results of behavioural bioassays demonstrated that ß-Ocimene strongly repelled mated E. obliqua females. Individual volatile compound exposure experiments revealed that (Z)-3-hexenol, linalool, α-farnesene and DMNT triggered the emission of ß-Ocimene from tea plants. Chemical inhibition experiments demonstrated that the emission of ß-Ocimene induced by (Z)-3-hexenol, linalool, α-farnesene and DMNT were dependent on Ca2+ and JA signalling. These findings help us to understand how E. obliqua moths respond to volatiles emitted from tea plants and provide new insight into volatile-mediated plant-plant interactions. They have potential significance for the development of novel insect and pest control strategies in crops.

Identification of loci and candidate gene GmSPX-RING1 responsible for phosphorus efficiency in soybean via genome-wide association analysis.

BACKGROUND: Phosphorus (P) is an essential element in maintaining high biomass and yield in crops. Soybean [Glycine max (L.) Merr.] requires a large amount of P during growth and development. Improvement of P efficiency and identification of P efficiency genes are important strategies for increasing soybean yield. RESULTS: Genome-wide association analysis (GWAS) with NJAU 355 K SoySNP array was performed to identify single nucleotide polymorphisms (SNPs) significantly associated with three shoot P efficiency-related traits of a natural population of 211 cultivated soybeans and relative values of these traits under normal P (+P) condition and P deficiency (-P) condition. A total of 155 SNPs were identified significantly associated with P efficiency-related traits. SNPs that were significantly associated with shoot dry weight formed a SNP cluster on chromosome 11, while SNPs that were significantly associated with shoot P concentration formed a SNP cluster on chromosome 10. Thirteen haplotypes were identified based on 12 SNPs, and Hap9 was considered as the optimal haplotype. Four SNPs (AX-93636685, AX-93636692, AX-93932863, and AX-93932874) located on chromosome 10 were identified to be significantly associated with shoot P concentration under +P condition in two hydroponic experiments. Among these four SNPs, two of them (AX-93636685 and AX-93932874) were also significantly associated with the relative values of shoot P concentration under two P conditions. One SNP AX-93932874 was detected within 5'-untranslated region of Glyma.10 g018800, which contained SPX and RING domains and was named as GmSPX-RING1. Furthermore, the function research of GmSPX-RING1 was carried out in soybean hairy root transformation. Compared with their respective controls, P concentration in GmSPX-RING1 overexpressing transgenic hairy roots was significantly reduced by 32.75% under +P condition; In contrast, P concentration in RNA interference of GmSPX-RING1 transgenic hairy roots was increased by 38.90 and 14.51% under +P and -P conditions, respectively. CONCLUSIONS: This study shows that the candidate gene GmSPX-RING1 affects soybean phosphorus efficiency by negatively regulating soybean phosphorus concentration in soybean hairy roots. The SNPs and candidate genes identified should be potential for improvement of P efficiency in future soybean breeding programs.

UGT85A53 promotes flowering via mediating abscisic acid glucosylation and FLC transcription in Camellia sinensis.

Uridine diphosphate (UDP)-dependent glycosyltransferases catalyse the glycosylation of small molecules and play important roles in maintaining cell homeostasis and regulating plant development. Glycosyltransferases are widely distributed, but their detailed roles in regulating plant growth and development are largely unknown. In this study, we identified a UDP-glycosyltransferase, UGT85A53, from Camellia sinensis, the expression of which was strongly induced by various abiotic stress factors and its protein product was distributed in both the cytoplasm and nucleus. Ectopic overexpression of CsUGT85A53 in Arabidopsis resulted in an early-flowering phenotype under both long- and short-day conditions. The transcript accumulation of the flowering repressor genes FLC and ABI5, an activator of FLC in ABA-regulated flowering signaling, were both significantly decreased in transgenic Arabidopsis compared with wild-type plants. The decreased expression level of FLC might be associated with an increased level of DNA methylation that was observed in CsUGT85A53-overexpressing (OE) plants. Biochemical analyses showed that CsUGT85A53 could glucosylate ABA to form inactive ABA-glycoside in vitro and in planta. Overexpression of CsUGT85A53 in Arabidopsis resulted in a decreased concentration of free ABA and increased concentration of ABA-glucoside. The early-flowering phenotype in the CsUGT85A53-OE transgenic lines was restored by ABA application. Furthermore, CsUGT85A53-OE plants displayed an ABA-insensitive phenotype with higher germination rates compared with controls in the presence of low concentrations of exogenous ABA. Our findings are the first to identify a UGT in tea plants that catalyses ABA glucosylation and enhance flowering transition as a positive regulator.

The identification of key candidate genes mediating yellow seedling lethality in a Lilium regale mutant.

Leaf color mutants are ideal materials for exploring plant photosynthesis mechanisms, chlorophyll biosynthetic pathways and chloroplast development. The yellow seedling lethal mutant lrysl1 was discovered from self-bred progenies of Lilium regale; however, the mechanism of leaf color mutation remains unclear. In this study, the ultrastructural and physiological features and de novo RNA-Seq data of a L. regale leaf color mutant and wild-type L. regale were investigated. Genetic analysis indicated that the characteristics of the lrysl1 mutant were controlled by a recessive nuclear gene. The chlorophyll a, chlorophyll b and carotenoid contents in the mutant leaves were lower than those in the wild-type leaves. Furthermore, the contents of the chlorophyll precursors aminolevulinic acid (ALA), porphobilinogen (PBG), protoporphyrin IX (ProtoIX), Mg-protoporphyrin IX (Mg-ProtoIX), and protochlorophyll (Pchl) decreased significantly in mutant leaves. Transcriptome data from the mutant and wild type showed that a total of 892 differentially expressed genes were obtained, of which 668 and 224 were upregulated genes and downregulated genes in the mutant, respectively. Almost all genes in the photosynthesis pathway and chlorophyll biosynthetic pathway were downregulated in the mutant, which corroborated the differences in the physiological features mentioned above. Further research indicated that the chloroplasts of the mutant leaves exhibited an abnormal morphology and distribution and that the expression of a gene related to chloroplast development was downregulated. It was concluded that abnormal chloroplast development was the main cause of leaf color mutation in the mutant lrysl1 and that LrGLK was a gene related to chloroplast development in L. regale. This research provides a foundation for further research on the mechanism by which LrGLK regulates chloroplast development in L. regale.

Selection of reference genes for quantitative real-time PCR analysis of photosynthesis-related genes expression in Lilium regale.

Photosynthesis is closely related to the growth of plants. A stable reference gene is fundamental for studies of the molecular mechanism of photosynthesis in Lilium regale. Therefore, it is very important to select a suitable reference gene for qRT-PCR analysis on genes of photosynthetic system, chlorophyll biosynthetic pathway and chloroplast development in Lilium regale. Three kinds of tissues, leaves and bulbs (abnormal leaves) of tissue culture plantlets and cotyledons of seedlings of the wild-type and mutant Lilium regale were selected as materials for qRT-PCR. Six housekeeping genes were selected as candidate genes from transcriptome sequencing data of the wild-type and yellow seedling lethal mutant of Lilium regale. Finally, the expression stability of six candidate reference genes was analyzed using geNorm, NormFinder, and BestKeeper software, the comparative ∆Ct method, and the RefFinder program. The results showed that LrActin2 was the best reference gene for qRT-PCR analysis of photosynthesis-related genes expression in leaves of tissue culture plantlets and seedlings of Lilium regale. This study provided useful data for further research on molecular mechanism of photosynthesis in the Lilium.

A genome-wide expression profile analysis reveals active genes and pathways coping with phosphate starvation in soybean.

BACKGROUND: Phosphorus is one of the most important macronutrients that is required for plant growth and development. However, stress under low-P conditions has become a limiting factor that affects crop yields and qualities. Plants have developed strategies to cope with this, while few genes associated with low-P tolerance have been identified in soybean. RESULTS: Genome-wide analyses were performed on the roots and leaves of a low-P-tolerant accession and a low-P-sensitive accession which were identified by hydroponic experiments under different P treatments. Through comparative analyses on the differently expressed genes, we explored 42 common genes that were highly correlated to low-P stress. The functional classification of these genes revealed 24 Gene Ontology (GO) terms of biological process including response to oxidation reduction, hormone stimuli, and biotic and abiotic stimuli. Additionally, three common pathways were identified. CONCLUSIONS: These results could not only promote the work on the molecular regulation mechanism under low-P stress in soybean, but also facilitate the cultivation of high-phosphorus-acquisition and high-phosphorus-utilization soybean varieties.

Association analysis for detecting significant single nucleotide polymorphisms for phosphorus-deficiency tolerance at the seedling stage in soybean [Glycine max (L) Merr].

Tolerance to low-phosphorus soil is a desirable trait in soybean cultivars. Previous quantitative trait locus (QTL) studies for phosphorus-deficiency tolerance were mainly derived from bi-parental segregating populations and few reports from natural population. The objective of this study was to detect QTLs that regulate phosphorus-deficiency tolerance in soybean using association mapping approach. Phosphorus-deficiency tolerance was evaluated according to five traits (plant shoot height, shoot dry weight, phosphorus concentration, phosphorus acquisition efficiency and use efficiency) comprising a conditional phenotype at the seedling stage. Association mapping of the conditional phenotype detected 19 SNPs including 13 SNPs that were significantly associated with the five traits across two years. A novel cluster of SNPs, including three SNPs that consistently showed significant effects over two years, that associated with more than one trait was detected on chromosome 3. All favorable alleles, which were determined based on the mean of conditional phenotypic values of each trait over the two years, could be pyramided into one cultivar through parental cross combination. The best three cross combinations were predicted with the aim of simultaneously improving phosphorus acquisition efficiency and use efficiency. These results will provide a thorough understanding of the genetic basis of phosphorus deficiency tolerance in soybean.

The effect of carbon-polyaniline hybrid coating on high-temperature electrochemical performance of perovskite-type oxide LaFeO3 for MH-Ni batteries.

An efficient carbon-polyaniline (PANI)-coated method was applied for perovskite-type oxide LaFeO3 to enhance its high-temperature electrochemical performance. Transmission electron microscopy (TEM) results reveal that LaFeO3 particles are evenly coated with carbon and PANI hybrid layers after carbon-PANI treatment. The carbon layers prevent the nanosized LaFeO3 particles from aggregation and allow the electrolyte to penetrate in every direction inside the particles. The PANI layers also enhance the electrocatalytic activity, facilitating hydrogen protons transferring from the electrolyte to the electrode interface. The cooperation of carbon and PANI hybrid layers results in a significant enhancement of the electrochemical performance at high temperatures. At an elevated temperature (60 °C), the maximum discharge capacity of the LaFeO3 electrodes remarkably increases from 231 mA h g(-1) to 402 mA h g(-1) and the high rate dischargeability at a discharge current density of 1500 mA g(-1) (HRD1500) increases from 22.7% to 44.3%. Moreover, the hybrid layers mitigate the corrosion of LaFeO3 electrodes by reducing the loss of active materials in the alkaline electrolyte, leading to increase in the capacity retention rate from 67.1% to 77.6% after 100 cycles (S100).

Construction of sodium alginate/trehalose/wheat starch ternary complex and its effects on storage stability of frozen dough system.

In order to improve the quality of frozen dough, a calcium alginate-coated sodium alginate/trehalose/wheat starch ternary complex was designed in this paper. The ternary complex was added to dough, and the dough quality were measured after 0-30 d of frozen storage. The XRD and FT-IR results showed the ternary complex was mainly starchy crystal. The TGA curves showed the starting (To), peak (Tp) and termination temperature (Tc) were increased. The interaction between sodium alginate and trehalose enhanced the thermal performance of ternary complex. As the ternary complex addition to dough increased, the maximum ice crystal formation zone of the frozen dough passed faster, resulting in more uniform and smaller ice crystals. The dough with 0.8% addition contained more bound water and had better hardness, springiness and cohesiveness. In conclusion, the study provides a novel insight and understanding for the development of ternary complex as food additives in frozen food industry.

Effects of sodium carboxymethyl cellulose on storage stability and qualities of different frozen dough.

Hydrocolloids as Additives have been used for improving the quality of frozen dough for a long time. In this work, the effects of sodium carboxymethyl cellulose (CMC) on quality changes of frozen dough in storage were studied. The water loss rate of the dough and water holding capacity were measured. Rheological and texture properties of the frozen dough were measured by a rheometer and a texture analyzer, respectively. Scanning electron microscopy (SEM) was used to characterize surface network structure and protein structure changes of the frozen dough. Our results reveal that the addition of CMC can inhibit the formation of ice crystals and recrystallization, thus effectively stabilizing the molecular structure of starch, and resulting in more uniform moisture distribution in the frozen dough. When 3% addition of CMC, the water holding capacity of the two kinds of dough reached the best, and the water loss rate of corn dough reached the lowest. The cohesion of the two kinds of dough reaches the maximum with 3 wt% addition of CMC, while the hardness and chewiness of wheat and corn multigrain dough reaches the maximum with 3 wt% and 4 wt% addition of CMC, respectively. The results show proper CMC addition (3 wt% and 4 wt%) finally improves the stability and qualities of the frozen dough. The research concerning the effects of CMC on quality of frozen dough provides better understanding for the frozen food industry.

Distribution of and Temporal Variation in Volatiles in Tea (Camellia sinensis) Flowers during the Opening Stages.

Tea (Camellia sinensis) flowers emit a large amount of volatiles that attract pollinators. However, few studies have characterized temporal and spatial variation in tea floral volatiles. To investigate the distribution of volatiles within tea flowers and their variation among opening stages, volatile components from different parts of tea flowers and different opening stages were collected by headspace solid-phase microextraction and analyzed by gas chromatography-mass spectrometry. A total of 51 volatile compounds of eight chemical classes were identified in the tea flowers. Volatile compounds were most abundant in tea flowers of the Shuchazao cultivar. Acetophenone, 1-phenylethanol, 2-phenylethanol, and benzyl alcohol were the most abundant volatiles. Terpenes were common in the sepals, and benzoids were common in the stamens. The fatty acid derivatives were mainly distributed in the pistils and receptacles and were less abundant in the petals, sepals, and stamens. During the opening phase of tea flowers, the volatile content increased 12-fold, which mainly stemmed from the increase in benzoids. These results enhance our understanding of the formation of volatiles in tea flowers.

Characterization of Key Odorants in Xinyang Maojian Green Tea and Their Changes During the Manufacturing Process.

Xinyang Maojian (XYMJ) green tea is a famous high-grade Chinese green tea, but the key odorants contributing to its aroma have been poorly understood. In this study, solid-phase microextraction and solvent-assisted flavor evaporation were used for sample preparation, and gas chromatography-mass spectrometry (GC-MS) and gas chromatography-olfactometry (GC-O) were used for both qualitative and quantitative analysis. A total of 50 volatile compounds of five chemical classes were identified in XYMJ tea infusion. Among them, nine odorants including nonanal, ß-ionone, octanal, E-nerolidol, linalool, cis-3-hexenyl hexanoate, geraniol, decanal, and ß-cyclocitral were identified as key odorants of XYMJ based on GC-O, odor activity values, and aroma combination experiments. Changes in the content of these aroma-active compounds during the manufacturing process of XYMJ (fresh leaves, fixing, rolling, shaping, and drying) were also determined. Most aroma-active compounds decreased after the fixation process, with the exception of cis-3-hexenyl hexanoate. This is the first study to investigate the key odorants in XYMJ using the sensomics approach. The findings of this study provide novel information on the aroma quality of XYMJ.

Scaling up of high-speed countercurrent chromatographic apparatus with three columns connected in series for rapid preparation of (-)-epicatechin.

In the present study, compact high-speed countercurrent chromatographic apparatus was constructed with three columns connected in series. Two sets of columns were prepared from 10 mm and 12 mm I.D. tubing to form 12 L and 15 L capacities, respectively. Performance of these columns was compared for the separation of (-)-epicatechin gallate (ECG) from a tea extract by flash countercurrent chromatography (FCCC). In each separation, 200 g of the tea extract in 1600 mL of mobile phase was loaded onto the column. The 12 L column gave 7.5 L (35 g of ECG) and the 15 L column gave 9 L (40 g of ECG) of ECG solution without impurities. The ECG solution was directly hydrolyzed by tannase into (-)-epicatechin. The hydrolysate was purified by flash chromatography on AB-8 macroporous resin to give 52 g of EC (purity 99.1%). This scaled up apparatus could be used for the industrial separation of natural products.

Isolation of the retinal isomers from the isomerization of all-trans-retinal by flash countercurrent chromatography.

The isolation of the retinal isomers from all-trans-retinal was performed by flash countercurrent chromatography. In each separation, isomerization reaction solution of 200mg all-trans-retinal could be loaded on a 1200 mL of high-speed countercurrent chromatographic column with 5mm bore, eluted by a mobile phase flow rate of 25 mL/min, resulting in 63 mg of 11-cis-retinal, 24 mg of 13-cis-retinal and 26 mg of 9-cis-retinal with purities more than 95%. n-Hexane-acetonitrile (3:1) was used as the solvent system which possesses the advantages of simplicity, re-use of the solvent and multiple injections. This method could be used to prepare 13-cis-retinal, 11-cis-retinal and 9-cis-retinal for the photoisomerization investigation, such as the effect of 11-cis-retinal in the visual system.