Effect of LED Lights on Secondary Metabolites and Antioxidant Activities in Red Pakchoi Baby Leaves.

Pakchoi (Brassica rapa subsp. chinensis) is one of the most widely consumed vegetables in Asian countries, and it is high in secondary metabolites. The availability, quantity, and quality of light play a critical role in the growth and development of plants. In this study, we investigated the effect of LEDs (light-emitting diodes; white, blue, red, and red + blue) on anthocyanin, glucosinolates, and phenolic levels in red pakchoi baby leaves. On the 24th day after sowing (DAS), red baby pakchoi leaves were harvested, and shoot length, root length, and fresh weight were measured. Among the different LED treatments, there was no significant difference in shoot length, whereas the highest root length was achieved in the red + blue LED treatment (23.8 cm). The fresh weight also showed a significant difference among the different LED treatments. In total, 12 phenolic and 7 glucosinolate individual compounds were identified using high-performance liquid chromatography (HPLC) analysis. The highest total glucosinolate (2937 µg/g dry wt) and phenolic (1589 µg/g dry wt) contents were achieved in baby leaves exposed to red + blue light. Similarly, the highest contents of total anthocyanins (1726 µg/g dry wt), flavonoids (4920 µg/g dry wt), and phenolics (5900 µg/g dry wt) were achieved in the red + blue treatment. Plants exposed to red + blue LED light showed the highest accumulation of anthocyanin, glucosinolates, and phenolic compounds. For antioxidant activity, DPPH (2,2-diphenyl-1-picrylhydrazylradical) free radical scavenging, ABTS (2,2-azinobis (3-ethylbenzothiazoline)-6-sulfonic acid) radical scavenging, and reducing power assays were performed, and the antioxidant activity of red pakchoi baby leaves grown under red + blue LED light was found to be the best. The metabolic profiling of the identified metabolites revealed distinct separation based on the secondary metabolites. This research will be helpful for farmers to choose the best LED light combination to increase the secondary metabolic content in pakchoi plants.

Metabolite profiles and biological activities of different phenotypes of Chinese cabbage (Brassica rapa ssp. Pekinensis).

Chinese cabbage is considered as one of the most important cruciferous vegetables in South Korea because of its use in salads, kimchi, and Korean cuisine. Secondary metabolites were quantified in three Chinese cabbage varieties: 65065, interspecific hybrid of Chinese cabbage × red cabbage exhibiting a deep purple color; 85772, interspecific hybrid of Chinese cabbage × red mustard exhibiting a reddish-purple color; and a typical Chinese green cabbage cultivar "CR Carotene" (Brassica rapa subsp. pekinensis cv. CR Carotene). A total of 54 metabolites (2 amines, 2 sugar alcohols, 2 sugar phosphates, 6 carbohydrates, 18 amino acids, 13 organic acids, 8 phenolic compounds, and 3 carotenoids) were detected in 85772. Of them, 52 metabolites excluding ß-carotene and 9-cis-ß-carotene, and 51 metabolites excluding leucine, ß-carotene, and 9-cis-ß-carotene, were detected in 65065 and CR Carotene, respectively. Amino acid content was the highest in 85772, followed by 65065 and CR Carotene. The cultivars 65065 and 85772 contained high levels of phenolic compounds and total anthocyanins. Cyanidin-, pelargonidin-, and petunidin-type anthocyanins were detected in 65065 and 85772. However, delphinidin-type anthocyanins which typically impart a deep purple color were identified only in the deep purple phenotype 65065. Furthermore, the total anthocyanin content was the highest in 85772 (4.38 ± 0.65 mg g -1 dry weight) followed by that in 65065 (3.72 ± 0.52 mg g-1 dry weight). Antibacterial and antioxidant analyses revealed remarkable antibacterial effects of the purple cultivars against pathogens Vibrio parahaemolyticus (KCTC 2471), Bacillus cereus (KCTC 3624), Pseudomonas aeruginosa (KCCM 11803), Staphylococcus aureus (KCTC 3881), Chryseobacterium gleum (KCTC 2094), and Proteus mirabilis (KCTC 2510)] and methicillin-resistant pathogenic strains of Pseudomonas aeruginosa (0826, 0225, 0254, 1113, 1378, 1731, p01827, and p01828) compared with the antibacterial effects of CR Carotene. Furthermore, 65065 and 85772 exhibited significantly higher antioxidant activity than that of the CR Carotene. Therefore, the novel purple Chinese cabbages (65065 and 85772), derived from interspecific hybridization, are potentially favorable alternatives to the typical green Chinese cabbage, given the higher content of amino acids, phenolic compounds, anthocyanins, and carotenoids, as well as an increased ability to scavenge free radicals and inhibit pathogen growth.

LED Lights Influenced Phytochemical Contents and Biological Activities in Kale (Brassica oleracea L. var. acephala) Microgreens.

Light-emitting diodes (LEDs) are regarded as an effective artificial light source for producing sprouts, microgreens, and baby leaves. Thus, this study aimed to investigate the influence of different LED lights (white, red, and blue) on the biosynthesis of secondary metabolites (glucosinolates, carotenoids, and phenolics) and the biological effects on kale microgreens. Microgreens irradiated with white LEDs showed higher levels of carotenoids, including lutein, 13-cis-ß-carotene, α-carotene, ß-carotene, and 9-cis-ß-carotene, than those irradiated with red or blue LEDs. These findings were consistent with higher expression levels of carotenoid biosynthetic genes (BoPDS and BoZDS) in white-irradiated kale microgreens. Similarly, microgreens irradiated with white and blue LEDs showed slightly higher levels of glucosinolates, including glucoiberin, progoitrin, sinigrin, and glucobrassicanapin, than those irradiated with red LEDs. These results agree with the high expression levels of BoMYB28-2, BoMYB28-3, and BoMYB29 in white- and blue-irradiated kale microgreens. In contrast, kale microgreens irradiated with blue LEDs contained higher levels of phenolic compounds (gallic acid, catechin, ferulic acid, sinapic acid, and quercetin). According to the total phenolic content (TPC) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) inhibition assays, the extracts of kale microgreens irradiated with blue LEDs had slightly higher antioxidant activities, and the DPPH inhibition percentage had a positive correlation with TPC in the microgreens. Furthermore, the extracts of kale microgreens irradiated with blue LEDs exhibited stronger antibacterial properties against normal pathogens and multidrug-resistant pathogens than those irradiated with white and red LEDs. These results indicate that white-LED lights are suitable for carotenoid production, whereas blue-LED lights are efficient in increasing the accumulation of phenolics and their biological activities in kale microgreens.

Impact of Light and Dark Treatment on Phenylpropanoid Pathway Genes, Primary and Secondary Metabolites in Agastache rugosa Transgenic Hairy Root Cultures by Overexpressing Arabidopsis Transcription Factor AtMYB12.

Agastache rugosa, otherwise called Korean mint, has a wide range of medicinal benefits. In addition, it is a rich source of several medicinally valuable compounds such as acacetin, tilianin, and some phenolic compounds. The present study aimed to investigate how the Tartary buckwheat transcription factor AtMYB12 increased the primary and secondary metabolites in Korean mint hairy roots cultured under light and dark conditions. A total of 50 metabolites were detected by using high-performance liquid chromatography (HPLC) and gas chromatography-time-of-flight mass spectrometry (GC-TOFMS). The result showed that the AtMYB12 transcription factor upregulated the phenylpropanoid biosynthesis pathway genes, which leads to the highest accumulation of primary and secondary metabolites in the AtMYB12-overexpressing hairy root lines (transgenic) than that of the GUS-overexpressing hairy root line (control) when grown under the light and dark conditions. However, when the transgenic hairy root lines were grown under dark conditions, the phenolic and flavone content was not significantly different from that of the control hairy root lines. Similarly, the heat map and hierarchical clustering analysis (HCA) result showed that most of the metabolites were significantly abundant in the transgenic hairy root cultures grown under light conditions. Principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) showed that the identified metabolites were separated far based on the primary and secondary metabolite contents present in the control and transgenic hairy root lines grown under light and dark conditions. Metabolic pathway analysis of the detected metabolites showed 54 pathways were identified, among these 30 were found to be affected. From these results, the AtMYB12 transcription factor activity might be light-responsive in the transgenic hairy root cultures, triggering the activation of the primary and secondary metabolic pathways in Korean mint.

The Effect of Light and Dark Treatment on the Production of Rosmarinic Acid and Biological Activities in Perilla frutescens Microgreens.

This study aimed to investigate the effect of light [a long-day photoperiod (16 h light/8 h dark cycle)] and dark treatment on the production of rosmarinic acid in P. frutescens microgreens and to determine its antioxidant and antibacterial activities. Microgreens of P. frutescens were grown under light and dark conditions and harvested after 10, 15, 20, and 25 days of each treatment. Although dry weight values of microgreens gradually increased from 10 to 25 days of both treatments, the microgreens grown under light treatment possessed slightly higher levels of dry weight than those grown in the dark. Rosmarinic acid and total phenolic content (TPC) were also analyzed using high-performance liquid chromatography (HPLC) and Folin-Ciocalteu assay. The accumulation patterns of rosmarinic acid and TPC gradually increased and decreased, respectively, in P. frutescens microgreens grown in continuous darkness. The highest accumulation was observed in microgreens grown for 20 days. However, rosmarinic acid and TPC values were not significantly different in microgreens grown under light conditions. According to the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical inhibition assay, the extracts of P. frutescens microgreens were confirmed to be strong antioxidants, and their ability to scavenge DPPH radicals was positively correlated with the total phenolic content in the microgreens after 10, 15, 20, and 25 days of both treatments. Considering the relatively higher values of dry weight, rosmarinic acid, TPC, and DPPH assay, P. frutescens microgreens after 20 days of darkness and 20 days of light treatment, respectively, were selected for screening antibacterial activity using nine pathogens. Both microgreen extracts showed strong antibacterial activity against pathogens. In particular, the extracts of microgreens grown for 20 days under light treatment showed higher antimicrobial effects. Therefore, the light treatments for 20 days, as well as the darkness treatment for 20 days, were the best conditions for P. frutescens microgreen production because of their high levels of dry weight, phenolics, and biological activities.

Effects of Carbohydrates on Rosmarinic Acid Production and In Vitro Antimicrobial Activities in Hairy Root Cultures of Agastache rugosa.

Agastache rugosa (popularly known as Korean mint) belongs to the Lamiaceae family and comprises 22 species of perennial aromatic medicinal species native to East Asian countries, such as Korea, Taiwan, Japan, and China. A. rugosa contains many phenolic compounds that exhibit pharmacological and physiological activities, including antioxidant, anticancer, antiviral, antifungal, and antibacterial activities. The highest concentrations of rosmarinic acid and its isomers have been reported in the roots of A. rugosa. In this in vitro study, hairy roots of A. rugosa were obtained and the carbohydrates (sorbitol, mannitol, glucose, maltose, galactose, mannose, and sucrose) were evaluated to determine those that were optimal for rosmarinic acid production and hairy root growth. Antioxidant and antibacterial activities of extracts of A. rugosa were also assessed. The best carbon source for A. rugosa hairy root cultures was sucrose, considering biomass productivity (0.460 ± 0.034 mg/30 mL), rosmarinic acid production (7.656 ± 0.407 mg/g dry weight), and total phenolic content (12.714 ± 0.202 mg/g gallic acid equivalent). Antioxidant and antimicrobial activities were displayed by A. rugosa hairy roots cultured in liquid medium supplemented with 100 mM sucrose. Twenty-five bacterial strains, including multidrug-resistant bacteria and one pathogenic yeast strain, were used for antimicrobial screening of A. rugosa hairy roots. The hairy root extracts displayed antibacterial activity against Micrococcus luteus (KCTC 3063) and Bacillus cereus (KCTC 3624). The inhibition of these bacteria was greater using A. rugosa hairy roots with the highest levels of phenolic compounds cultured in the presence of sucrose, compared to hairy roots with the lowest levels of phenolic compounds cultured in the presence of fructose. Considering hairy root biomass, phenolic compound production, and antibacterial activity, sucrose is the best carbon source for A. rugosa hairy root cultures.

Effects of Chilling Treatment on Baicalin, Baicalein, and Wogonin Biosynthesis in Scutellaria baicalensis Plantlets.

When plants are exposed to stressful conditions, they modulate their nutrient balance by regulating their primary and secondary metabolisms to adapt. In this study, changes in primary and secondary metabolites elicited by chilling stress treatment and the effects of treatment duration were examined in roots of Scutellaria baicalensis (S. baicalensis) plantlets. The concentrations of most sugars (maltose, glucose, sucrose, and fructose) and of several amino acids (proline and GABA), which are crucial regarding plant defense mechanisms, increased with increasing duration of chilling stress. Furthermore, salicylic acid levels increased after two-day chilling treatments, which may enhance plant tolerance to cold temperatures. The concentrations of flavones (baicalin, baicalein, and wogonin) increased during chilling stress, and those of phenolic acids (ferulic acid and sinapic acid) increased after two-day chilling treatments. The concentrations of these flavones were positively correlated with sucrose levels which acted as energy sources.

Metabolic Analyses and Evaluation of Antioxidant Activity in Purple Kohlrabi Sprouts after Exposed to UVB Radiation.

Various metabolites act as plant defense molecules due to their antioxidant abilities. This study aimed to investigate the influence of UVB irradiation on the accumulation of metabolites, including primary metabolites (sugar, sugar alcohols, amino acids, organic acids, and an amine) and secondary metabolites (anthocyanins, fatty acids, and phenolic acids), and its synergistic antioxidant ability, in purple kohlrabi sprouts. Metabolite analyses revealed a total of 92 metabolites in the sprouts. Specifically, the levels of most amino acids increased after 24 h of UVB treatment, and then slightly decreased in the kohlrabi sprouts. The levels of most sugars and sugar alcohols increased after 24 h of UVB treatment and then decreased. The levels of TCA cycle intermediates and phenolic acids gradually increased during the UVB treatment. Furthermore, the levels of some fatty acids gradually increased during the UVB treatment, and the levels of the other fatty acids increased after 6 h of UVB treatment and then decreased. In particular, the levels of most anthocyanins, known to be strong antioxidants, gradually increased after 24 h of UVB treatment. In the in vitro ABTS scavenging assay, UVB-treated purple kohlrabi sprouts showed increased scavenging ability. This may be attributed to the increased accumulation of metabolites acting as antioxidants, in response to UVB treatment. This study confirmed that UVB irradiation induced the alteration of primary and secondary metabolism in the kohlrabi sprouts.

Gene Expression and Metabolic Analyses of Nontransgenic and AtPAP1 Transgenic Tobacco Infected with Potato Virus X (PVX).

Potato virus X (PVX), a species of the genus Potexvirus, is a plant pathogenic virus that causes severe symptoms such as mild mosaic, crinkling, necrosis, and mottling on leaves. The objectives of the present study were to investigate the effect of PVX virus infection on the metabolic system in nontransgenic and Arabidopsis thaliana production of anthocyanin pigment 1 (AtPAP1) transgenic tobacco using transcript expression analysis and metabolic profiling. Potato virus X inoculation increased the gene expression of phenylpropanoid and flavonoid biosynthesis and the production of chlorogenic acid, p-coumaric acid, benzoic acid, rutin, quercetin, and kaempferol in nontransgenic tobacco leaves. However, in the AtPAP1 transgenic tobacco leaves, PVX inoculation decreased the expression of AtPAP1 and phenylpropanoid and flavonoid biosynthesis genes, and the production of phenolics and anthocyanin also declined. In contrast, the levels of amino acids and tricarboxylic acid (TCA) cycle intermediates increased after infection in the AtPAP1 transgenic plant leaves. To date, these results have not been reported previously. We suggest that PVX infection decreases AtPAP1 expression, leading to the downregulation of phenylpropanoid and flavonoid biosynthesis in transgenic plants.

Identification, Characterization, and Expression Analysis of Carotenoid Biosynthesis Genes and Carotenoid Accumulation in Watercress (Nasturtium officinale R. Br.).

Watercress (Nasturtium officinale R. Br.) is an important aquatic herb species belonging to the Brassicaceae family. It has various medicinal properties and has been utilized for the treatment of cancer and other diseases; however, currently available genomic information regarding this species is limited. Here, we performed the first comprehensive analysis of the carotenoid biosynthesis pathway (CBP) genes of N. officinale, which were identified from next-generation sequencing data. We identified and characterized 11 putative carotenoid pathway genes; among these, nine full and two partial open reading frames were determined. These genes were closely related to CBP genes of the other higher plants in the phylogenetic tree. Three-dimensional structure analysis and multiple alignments revealed several distinct conserved motifs, including aspartate or glutamate residues, carotene-binding motifs, and dinucleotide-binding motifs. Quantitative reverse transcription-polymerase chain reaction results showed that the CBP was expressed in a tissue-specific manner: expression levels of NoPSY, NoPDS, NoZDS-p, NoCrtISO, NoLCYE, NoCHXE-p, and NoCCD were highest in the flower, whereas NoLCYB, NoCHXB, NoZEP, and NoNCED were highest in the leaves. Stems, roots, and seeds did not show a significant change in the expression compared to the leaves and flowers. High-performance liquid chromatography analysis of the same organs showed the presence of seven distinct carotenoid compounds. The total carotenoid content was highest in the leaves followed by flowers, seeds, stems, and roots. Among the seven individual carotenoids, the levels of six carotenoids (i.e., 13-Z-ß-carotene, 9-Z-ß-carotene, E-ß-carotene, lutein, violaxanthin, and ß-cryptoxanthin) were highest in the leaves. The highest content was observed for lutein, followed by E-ß-carotene, and 9-Z-ß-carotene; these carotenoids were much higher in the leaves compared to the other organs. The results will be useful references for further molecular genetics and functional studies involving this species and other closely related species.

Molecular Characterization, Expression Analysis of Carotenoid, Xanthophyll, Apocarotenoid Pathway Genes, and Carotenoid and Xanthophyll Accumulation in Chelidonium majus L.

Chelidonium majus L. is a perennial herbaceous plant that has various medicinal properties. However, the genomic information about its carotenoid biosynthesis pathway (CBP), xanthophyll biosynthesis pathway (XBP), and apocarotenoid biosynthesis pathway (ABP) genes were limited. Thus, the CBP, XBP, and ABP genes of C. majus were identified and analyzed. Among the 15 carotenoid pathway genes identified, 11 full and 4 partial open reading frames were determined. Phylogenetic analysis of these gene sequences showed higher similarity with higher plants. Through 3D structural analysis and multiple alignments, several distinct conserved motifs were identified, including dinucleotide binding motif, carotene binding motif, and aspartate or glutamate residues. Quantitative RT-PCR showed that CBP, XBP, and ABP genes were expressed in a tissue-specific manner; the highest expression levels were achieved in flowers, followed by those in leaves, roots, and stems. The HPLC analysis of the different organs showed the presence of eight different carotenoids. The highest total carotenoid content was found in leaves, followed by that in flowers, stems, and roots. This study provides information on the molecular mechanisms involved in CBP, XBP, and ABP genes, which might help optimize the carotenoid production in C. majus. The results could also be a basis of further studies on the molecular genetics and functional analysis of CBP, XBP, and ABP genes.

Integrated Analysis of Transcriptome and Metabolome and Evaluation of Antioxidant Activities in Lavandulapubescens.

Lavandula pubescens, belonging to the Labiatae family, is a newly discovered strongly aromatic species of lavender that is potentially beneficial for human health. Given the economic importance of lavender species, we sought in this study to characterize the terpenoid biosynthesis of L. pubescens by obtaining transcriptomic and metabolic datasets. Transcriptome analysis of L. pubescens grown aseptically in tissue culture medium yielded 124,233 unigenes with an average length of 470 bp and N50 value of 522 bp from 9,476,122,928 raw reads. In order to provide relevant biological information, the unigenes were annotated using the following public databases: National Center for Biotechnology Information (NCBI) nucleotide (NT) and non-redundant protein (NR), Brassica (BRAD), Arabidopsis Information Resource (TAIR), Clusters of Orthologous Groups (COG), and Gene Ontology (GO). NR annotation results revealed that L. pubescens is genetically closely related to Sesamum indicum. On the basis of the transcriptome data, a total of 14 cDNA clones encoding the terpene biosynthetic genes LpDXS, LpMCT, LpMCS, LpHDR, LpIDI, LpAACT, LpHMGS, LpHMGR, LpMVK, LpPMK, LpMVD, LpGPPS, LpSQS, and LpGGPPS were identified in L. pubescens. These were quantified in the roots, stems, and leaves of L. pubescens using quantitative real-time polymerase chain reaction (qRT-PCR), which revealed that the gene expression levels were higher in the leaves and stems than in the roots, which was found to be consistent with the levels of ursolic and oleanolic acids in the different organs using high-performance liquid chromatography (HPLC). A total of 48 hydrophilic metabolites were identified and quantified in the organs using gas chromatography time-of-flight mass spectrometry (GC-TOFMS). Furthermore, the antioxidant activity of an ethyl acetate extract of L. pubescens leaves was examined using different methods to determine the potential therapeutic properties. A reducing power assay revealed that the absorbance values increased in a concentration-dependent manner, whereas a 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay indicated the strong activity (60.4 ± 0.9%) of the ethyl acetate extract at a concentration of 100 µg/mL, which also showed strong hydrogen peroxide (57.4 ± 2.7%), superoxide radical (62.1 ± 0.7%), and hydroxyl radical (58.6 ± 0.4%) scavenging activities.

Metabolic Analysis of Root, Stem, and Leaf of Scutellaria baicalensis Plantlets Treated with Different LED Lights.

Light emitting diodes (LEDs) have recently been considered an efficient artificial light source in plant factories for enhancing plant growth and nutritional quality. Accordingly, this study aimed to review blue, red, and white LED light sources for efficiency and length of the growing period to produce seedlings of Scutellaria baicalensis with high nutritional value. The roots, stems, and leaves of S. baicalensis seedlings were grown under different LED lights and harvested after two and four weeks, and analyzed using high-performance liquid chromatography and gas chromatography time-of-flight mass spectrometry to identify and quantify primary and secondary metabolites. Roots, particularly in the seedlings treated with white LEDs were determined to contain the greatest concentrations of the representative compounds present in S. baicalensis: baicalin, baicalein, and wogonin, which show highly strong biological properties compared to the other plant organs. A total of 50 metabolites (amino acids, sugars, sugar alcohols, organic acids, phenolic acids, and amines) were detected in the roots, stems, and leaves of S. baicalensis seedlings, and the concentrations of primary and secondary metabolites were generally decreased with the increasing duration of LED illumination. Therefore, this study suggests that white LED light and a 2-week growing period are the most efficient conditions for the production of baicalin, baicalein, and wogonin.

Effect of Light and Dark on the Phenolic Compound Accumulation in Tartary Buckwheat Hairy Roots Overexpressing ZmLC.

Fagopyrum tataricum 'Hokkai T10' is a buckwheat cultivar capable of producing large amounts of phenolic compounds, including flavonoids (anthocyanins), phenolic acids, and catechin, which have antioxidant, anticancer, and anti-inflammatory properties. In the present study, we revealed that the maize transcription factor Lc increased the accumulation of phenolic compounds, including sinapic acid, 4-hydroxybenzonate, t-cinnamic acid, and rutin, in Hokkai T10 hairy roots cultured under long-photoperiod (16 h light and 8 h dark) conditions. The transcription factor upregulated phenylpropanoid and flavonoid biosynthesis pathway genes, yielding total phenolic contents reaching 27.0 ± 3.30 mg g-1 dry weight, 163% greater than the total flavonoid content produced by a GUS-overexpressing line (control). In contrast, when cultured under continuous darkness, the phenolic accumulation was not significantly different between the ZmLC-overexpressing hairy roots and the control. These findings suggest that the transcription factor (ZmLC) activity may be light-responsive in the ZmLC-overexpressing hairy roots of F. tataricum, triggering activation of the phenylpropanoid and flavonoid biosynthesis pathways. Further studies are required on the optimization of light intensity in ZmLC-overexpressing hairy roots of F. tataricum to enhance the production of phenolic compounds.

Metabolic profiling and antioxidant activity during flower development in Agastache rugosa.

Our previous study showed that flowers of Agastache rugosa had higher phenolic levels and higher antibacterial and antioxidant capacity compared to those of the leaves and stems. The aim of this study was to provide information on the variation in primary and secondary metabolites during flower development in A. rugosa by using high performance liquid chromatography (HPLC) and assays of total anthocyanin (TAC), flavonoid (TFC), and phenolic content (TPC), as well as gas chromatography time-of-flight mass spectrometry (GC-TOFMS) analysis. Assays of TPC, TAC, and TFC showed that the floral bud (stage I) contained higher TPC than did the partially open flower (stage II) and fully open flower (stage III). However, the TFC was the highest at stage II, and the highest TAC was observed at stage III. Furthermore, HPLC analysis revealed that the level of total phenylpropanoids, including rosmarinic acid, tilianin, acacetin, 4-hydroxybenzoic acid, caffeic acid, chlorogenic acid, trans-cinnamic acid, rutin, (-)-epicatechin, quercetin, and kaempferol, was higher in stages I and II, but the concentrations of rutin and rosmarinic acid were highest in stage III. A total of 43 compounds, including amino acids, organic acids, phenolic compounds, sugars, photorespiration-related compounds, and intermediates of the tricarboxylic acid cycle, were identified through GC-TOFMS analysis. Of these compounds, most amino acids decreased during flower development. In contrast, the increase in concentrations of glucose and sucrose were observed from stages I to III. In this study, health-beneficial compounds were identified and quantified in flowers of A. rugosa. Accordingly, our results suggests that A. rugosa flowers can potentially be used as biomaterials for pharmaceuticals, cosmetics, food, and related industries. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at (10.1007/s12298-021-00945-z).

Profiles of Secondary Metabolites (Phenolic Acids, Carotenoids, Anthocyanins, and Galantamine) and Primary Metabolites (Carbohydrates, Amino Acids, and Organic Acids) during Flower Development in Lycoris radiata.

This study aimed to elucidate the variations in primary and secondary metabolites during Lycorisradiata flower development using high performance liquid chromatography (HPLC) and gas chromatography time-of-flight mass spectrometry (GC-TOFMS). The result showed that seven carotenoids, seven phenolic acids, three anthocyanins, and galantamine were identified in the L. radiata flowers. Most secondary metabolite levels gradually decreased according to the flower developmental stages. A total of 51 metabolites, including amines, sugars, sugar intermediates, sugar alcohols, amino acids, organic acids, phenolic acids, and tricarboxylic acid (TCA) cycle intermediates, were identified and quantified using GC-TOFMS. Among the hydrophilic compounds, most amino acids increased during flower development; in contrast, TCA cycle intermediates and sugars decreased. In particular, glutamine, asparagine, glutamic acid, and aspartic acid, which represent the main inter- and intracellular nitrogen carriers, were positively correlated with the other amino acids and were negatively correlated with the TCA cycle intermediates. Furthermore, quantitation data of the 51 hydrophilic compounds were subjected to partial least-squares discriminant analyses (PLS-DA) to assess significant differences in the metabolites of L. radiata flowers from stages 1 to 4. Therefore, this study will serve as the foundation for a biochemical approach to understand both primary and secondary metabolism in L. radiata flower development.

Comparative Analysis of Secondary Metabolites and Metabolic Profiling between Diploid and Tetraploid Morus alba L.

We profiled and quantified primary (amine, organic acids, tricarboxylic acid cycle intermediates, amino acids, and carbohydrates) and secondary metabolites (triterpenoids, phenolic acids, carotenoids, flavonoids, and anthocyanins) in the edible parts (leaves and fruits) of the diploid and tetraploid cultivar Morus alba L. 'Cheongil.' Through comprehensive metabolic profiling, the tetraploid mulberry cultivar was able to produce diverse metabolites supported by higher accumulation patterns of primary and secondary metabolites in their edible parts. In particular, the edible parts of the tetraploid showed higher accumulation patterns of most metabolites (amino acids, carbohydrates, carotenoids, and anthocyanins) than the diploid, which was supported by the results of principal component analyses (PCAs) showing a clear separation between the diploid and tetraploid groups. Additionally, this metabolome study comprehensively described the correlation between primary and secondary metabolites in the edible parts of diploid and tetraploid mulberry cultivars and provided information useful for plant breeding strategies to improve metabolite biosynthesis using polyploidy.

Enhancement of the flavone contents of Scutellaria baicalensis hairy roots via metabolic engineering using maize Lc and Arabidopsis PAP1 transcription factors.

Baicalin, baicalein, and wogonin are valuable natural flavonoid compounds produced by Scutellaria baicalensis. In this study, we showed that the maize transcription factor Lc can enhance the production of these three flavonoids in hairy root cultures of S. baicalensis by comprehensively upregulating flavonoid biosynthesis pathway genes (SbPAL1, SbC4H, and Sb4CL) and baicalein 7-O-glucuronosyltransferase (UBGAT), ultimately yielding total flavonoid contents of up to 80.5 ± 6.15 mg g-1 dry weight, which was 322% greater than the average value of total flavonoid contents produced by three GUS-overexpressing lines. Similarly, the Arabidopsis transcription factor PAP1 was found to enhance flavonoid accumulation by upregulating SbPAL1, SbPAL2, SbPAL3, SbC4H, Sb4CL, SbCHI, and UBGAT, ultimately yielding total flavonoid contents of up to 133 ± 7.66 mg g-1 dry weight, which was 532% greater than the average value of total flavonoid contents produced by three GUS-overexpressing lines. These findings indicate that metabolic engineering in S. baicalensis can be achieved using Agrobacterium rhizogenes-mediated transformation and that the production of baicalin, baicalein, and wogonin can be enhanced via the overexpression of ZmLc and AtPAP1 in hairy root cultures. These results also indicate that ZmLc and AtPAP1 can be used as positive regulators of the flavonoid biosynthetic pathway of S. baicalensis hairy root cultures.

Metabolite Profiling and Comparative Analysis of Secondary Metabolites in Chinese Cabbage, Radish, and Hybrid xBrassicoraphanus.

We profiled and quantified primary and secondary metabolites in the leaves and roots of xBrassicoraphanus (Baemuchae), Brassica campestris ssp. pekinensis (Chinese cabbage), and Raphanus sativus (radish). We obtained 72 metabolites from leaves and 68 metabolites from both leaves and roots of xBrassicoraphanus, Chinese cabbage, and radish. The metabolic profiles in this study revealed intermediate-level production of most metabolites from different parts of Baemuchae compared with that from different parts of Chinese cabbage and radishes. This was supported by the results of principal component analyses for the detected metabolites, which indicated that the Baemuchae group was located between the Chinese cabbage and radish groups. In particular, several amino acids (phenylalanine, tryptophan, and methionine) played the main role in phenylpropanoid and glucosinolate biosynthesis and were positively correlated with phenolic compounds, indolic glucosinolates, and aliphatic glucosinolates, respectively, in different parts. Furthermore, analysis of different species revealed the presence of 10 different glucosinolates, 10 phenolics, and 7 carotenoids, and their levels varied in the roots and leaves of the studied species. Among the leaves of the three species, Chinese cabbage had the highest total glucosinolate level, which was 3.14 times higher than the lowest level observed in radish. Baemuchae had the highest total phenolic compound level, which was 2.87 times higher than the lowest level found in Chinese cabbage, and radish had the highest carotenoid level, which was 12.41 times higher than the lowest one observed in Chinese cabbage. In the roots of Baemuchae, Chinese cabbage, and radish, glucosinolate levels did not vary significantly. Chinese cabbage contained the highest total phenolic compound level, which was 2.38 times higher than the lowest level found in radish, and the highest total carotenoid level, which was 2.49 times higher than the lowest level observed in Baemuchae. This metabolomic study provided chemical composition information that can be applied to future breeding strategies and comprehensively described the relationship among metabolites detected in the three plant species.

Metabolic Profiling of Primary Metabolites and Galantamine Biosynthesis in Wounded Lycoris radiata Callus.

Plants are continuously exposed to abiotic and biotic factors that lead to wounding stress. Different plants exhibit diverse defense mechanisms through which various important metabolites are synthesized. Humans can exploit these mechanisms to improve the efficacy of existing drugs and to develop new ones. Most previous studies have focused on the effects of wounding stress on the different plant parts, such as leaves, stems, and roots. To date, however, no study has investigated the accumulation of primary and galantamine content following the exposure of a callus to wounding stress. Therefore, in the present study, we exposed Lycoris radiata calli to wounding stress and assessed the expression levels of several genes involved in metabolic pathways at various time points (0, 3, 6, 12, 24, 48, 72, and 96 h of exposure). Furthermore, we quantify the primary and galantamine content using gas chromatography-time-of-flight mass spectrometry and the high-performance liquid chromatography qRT-PCR analysis of eight galantamine pathway genes (LrPAL-2, LrPAL-3, LrC4H-2, LrC3H, LrTYDC2, LrN4OMT, LrNNR, and LrCYP96T) revealed that seven genes, except LrN4OMT, were significantly expressed following exposure to wounding stress. Galantamine contents of calli after 3, 6, 12, 24, 48, 72, and 96 h of exposure were respectively 2.5, 2.5, 3.5, 3.5, 5.0, 5.0, and 8.5 times higher than that after 0 h of exposure. Furthermore, a total of 48 hydrophilic metabolites were detected in the 0 h exposed callus and 96 h exposed callus using GC-TOFMS. In particular, a strong positive correlation between galantamine and initial precursors, such as phenylalanine and tyrosine, was observed.