The Effects of Body Fat Reduction through the Metabolic Control of Steam-Processed Ginger Extract in High-Fat-Diet-Fed Mice.

The prevalence of metabolic syndrome is increasing globally due to behavioral and environmental changes. There are many therapeutic agents available for the treatment of chronic metabolic diseases, such as obesity and diabetes, but the data on their efficacy and safety are lacking. Through a pilot study by our group, Zingiber officinale rhizomes used as a spice and functional food were selected as an anti-obesity candidate. In this study, steam-processed ginger extract (GGE) was used and we compared its efficacy at alleviating metabolic syndrome-related symptoms with that of conventional ginger extract (GE). Compared with GE, GGE (25-100 µg/mL) had an increased antioxidant capacity and α-glucosidase inhibitory activity in vitro. GGE was better at suppressing the differentiation of 3T3-L1 adipocytes and lipid accumulation in HepG2 cells and promoting glucose utilization in C2C12 cells than GE. In 16-week high-fat-diet (HFD)-fed mice, GGE (100 and 200 mg/kg) improved biochemical profiles, including lipid status and liver function, to a greater extent than GE (200 mg/kg). The supplementation of HFD-fed mice with GGE (200 mg/kg) resulted in the downregulation of SREBP-1c and FAS gene expression in the liver. Collectively, our results indicate that GGE is a promising therapeutic for the treatment of obesity and metabolic syndrome.

Phenylalkyl Glycosides from the Flowers of Brugmansia arborea L. and Their Radical Scavenging Effect and Protective Effect on Pancreatic Islets Damaged by Alloxan in Zebrafish (Danio rerio) Larvae.

The study aimed to investigate the antioxidant and antidiabetic activity of Brugmansia arborea L. flower extracts, solvent fractions, and isolated compounds. B. arborea L flowers were extracted with aqueous methanol, and concentrated extract was successively partitioned into EtOAc, n-BuOH, and H2O fractions. Repeated silica gel and octadecyl silica gel column chromatographies for EtOAc and n-BuOH fractions led to the isolation of a new phenylalkyl glycoside (6), along with five known ones. Several spectroscopic data led to the structure determination of one new phenylalky glycoside as brugmansioside C (named) (6) and five known ones as benzyl-O-ß-D-glucopyranoside (1), benzyl-O-ß-D-glucosyl-(1→6)-ß-D-glucopyranoside (2), 2-phenylethyl-O-ß-D-glucopyranoside (3), 2-phenylethyl-O-ß-D-glucosyl-(1→6)-ß-D-glucopyranoside (4), and 3-phenylpropyl-O-ß-D-glucopyranoside (5). The five known ones (1-5) were isolated from B. arborea flowers for the first time in this study. The extract, solvent fractions, and all isolated compounds showed radical scavenging activities using ABTS radical, and EtOAc fraction showed the highest scavenging capacity, whereas compounds 2, 4, and 6 did not display the capacity to use the DPPH radical. The extract, solvent fractions, and all isolated compounds showed a protective effect on pancreatic islets damaged by alloxan treatment in zebrafish larvae. The pancreatic islet size treated with EtOAc, n-BuOH fractions, and all compounds significantly increased by 64.0%, 69.4%, 82.0%, 89.8%, 80.0%, 97.8%, 103.1%, and 99.6%, respectively, compared to the alloxan-induced group. These results indicate that B. arborea flowers and their isolated compounds are useful as potential antioxidant and antidiabetic agents.

Synthesis of Alpha-Linked Glucosides from Soybean Isoflavone Aglycones Using Amylosucrase from Deinococcus geothermalis.

Soybean isoflavone aglycones (SIAs) have many biological activities but are poorly water-soluble in the human body. Glycosylation provides structural diversity to SIAs and can alter their physicochemical properties, including water solubility. An alpha-linked glucosylation of SIA was achieved using amylosucrase from Deinococcus geothermalis. A total of 13 alpha-linked glucosyl SIAs were obtained, and their colors in solution were confirmed. The structures of the isolated compounds were identified by mass spectrometry and multidimensional nuclear magnetic resonance spectroscopy. The amylosucrase transglycosylation formed new isoflavone glycosides with alpha glycosidic bonds at C-7 and/or C-4' of SIAs, followed by the production of isoflavone glycosides with alpha (1 → 6) glycosidic bonds. The products with a glucosyl moiety attached to the C-4' of SIAs were found to be more water-soluble than their counterparts attached to the C-7 and/or beta-linkages. This study suggests a strategy for the synthesis of bioactive compounds with enhanced water solubility through alpha-linked glucosylation.

8-paradol from ginger exacerbates PINK1/Parkin mediated mitophagy to induce apoptosis in human gastric adenocarcinoma.

Gastric cancer (GC) occurs in the gastric mucosa, and its high morbidity and mortality make it an international health crisis. Therefore, novel drugs are needed for its treatment. The use of natural products and their components in cancer treatments has shown promise. Therefore, this study aimed to evaluate the effect of 8-paradol, a phenolic compound isolated from ginger (Zingiber officinale Roscoe), on GC and determine its underlying mechanisms of action. In this study, repeated column chromatography was conducted on ginger EtOH extract to isolate gingerol and its derivatives. The cytotoxicity of the eight ginger compounds underwent a (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide) tetrazolium reduction (MTT) assay. 8-paradol showed the most potent cytotoxicity effect among the isolated ginger compounds. The underlying mechanism by which 8-paradol regulated specific proteins in AGS cells was evaluated by proteomic analysis. To validate the predicted mechanisms, AGS cells and thymus-deficient nude mice bearing AGS xenografts were used as in vitro and in vivo models of GC, respectively. The results showed that the 8-paradol promoted PINK1/Parkin-associated mitophagy, mediating cell apoptosis. Additionally, the inhibition of mitophagy by chloroquine (CQ) ameliorated 8-paradol-induced mitochondrial dysfunction and apoptosis, supporting a causative role for mitophagy in the 8-paradol-induced anticancer effect. Molecular docking results revealed the molecular interactions between 8-paradol and mitophagy-/ apoptosis-related proteins at the atomic level. Our study provides strong evidence that 8-paradol could act as a novel potential therapeutic agent to suppress the progression of GC by targeting mitophagy pathway.

Phenolic Compounds from the Fruits of Prunus davidiana (Rosaceae) and Their Antioxidant Activities.

This research was supported by Cooperative Research Program for Agriculture Science & Technology Development (Project No. PJ014204032019) and the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2020R1A6A3A01100042).

Steppogenin suppresses tumor growth and sprouting angiogenesis through inhibition of HIF-1α in tumors and DLL4 activity in the endothelium.

BACKGROUND: Hypoxia is a characteristic feature of many solid tumors. As an adaptive response to hypoxia, tumor cells activate hypoxia-inducible factor-1α (HIF-1α). Under hypoxic conditions, angiogenesis mediated by HIF-1α is involved in the growth and metastasis of tumor cells. During the angiogenic process, differentiated tip endothelial cells (ECs) characterized by high expression of DLL4 promote angiogenic germination through filopodia. Inhibitors of HIF-1α or DLL4 have been widely studied PURPOSE: We tried to find inhibitors targeting both HIF-1α and DLL4 in tumor which have not yet been developed. STUDY DESIGN: In this study, we examined a natural compound that inhibits sprouting angiogenesis and tumor growth by targeting both HIF-1α and DLL4 under hypoxic conditions. METHODS: After examining cell viability of 70 selected natural compounds, we assessed the effects of compounds on HIF-1α and DLL4 transcriptional activity using a dual-luciferase reporter assay. Western blot analysis, immunofluoresecnt assay and real-time qPCR were performed to identify expression of proteins, such as HIF-1α and DLL4, as well as HIF-1α target genes under hypoxic conditions. In vitro angiogenesis assay and in vivo allograft tumor experiment were performed to investigate inhibition of tumor growth through anti-angiogenic activity. RESULTS: Among these compounds, steppogenin, which is extracted from the root bark of Morus alba l, respectively inhibited the transcriptional activity of HIF-1α under hypoxic conditions in HEK293T cells and vascular endothelial growth factor (VEGF)-induced DLL4 expression in vascular ECs in a dose-dependent manner. In tumor cells and retinal pigment epithelial cells, steppogenin significantly suppressed HIF-1α protein levels under hypoxic conditions as well as VEGF-induced DLL4 expression in ECs. Furthermore, steppogenin suppressed hypoxia-induced vascular EC proliferation and migration as well as VEGF-induced sprouting of EC spheroids. CONCLUSION: These results suggest that the natural compound steppogenin could potentially be used to treat angiogenic diseases, such as those involving solid tumors, because of its dual inhibition of HIF-1α and DLL4.

Label-free affinity screening, design and synthesis of inhibitors targeting the Mycobacterium tuberculosis L-alanine dehydrogenase.

The ability of Mycobacterium tuberculosis (Mtb) to persist in its host may enable an evolutionary advantage for drug resistant variants to emerge. A potential strategy to prevent persistence and gain drug efficacy is to directly target the activity of enzymes that are crucial for persistence. We present a method for expedited discovery and structure-based design of lead compounds by targeting the hypoxia-associated enzyme L-alanine dehydrogenase (AlaDH). Biochemical and structural analyses of AlaDH confirmed binding of nucleoside derivatives and showed a site adjacent to the nucleoside binding pocket that can confer specificity to putative inhibitors. Using a combination of dye-ligand affinity chromatography, enzyme kinetics and protein crystallographic studies, we show the development and validation of drug prototypes. Crystal structures of AlaDH-inhibitor complexes with variations at the N6 position of the adenyl-moiety of the inhibitor provide insight into the molecular basis for the specificity of these compounds. We describe a drug-designing pipeline that aims to block Mtb to proliferate upon re-oxygenation by specifically blocking NAD accessibility to AlaDH. The collective approach to drug discovery was further evaluated through in silico analyses providing additional insight into an efficient drug development strategy that can be further assessed with the incorporation of in vivo studies.

8-Methoxybutin inhibits α-MSH induced melanogenesis and proliferation of skin melanoma by suppression of the transactivation activity of microphthalmia-associated transcription factor.

Microphthalmia-associated transcription factor (MITF) is highly expressed in melanocytes and is the main regulator of melanogenesis and melanocyte cell fate. Although MITF is important for the differentiation and development of melanocytes, it is also considered an oncogene of skin melanoma. Based on these findings, MITF could be an attractive therapeutic target for skin cancer intervention. This study identified 8-methoxybutin as an inhibitor of MITF and investigated the underlying mechanism. 8-Methoxybutin inhibited α-MSH-induced melanogenesis in murine melanoma cells (B16F10) and skin melanoma proliferation by reducing melanogenic gene expression via blockade of the transactivation activity of MITF. In silico docking analysis and pull-down analysis suggested that 8-methoxybutin binds to the DNA-binding domain of MITF and further inhibits its binding to the E-box in the promoter of target genes, including tyrosinase. In addition, 8-methoxybutin suppressed growth of skin melanoma in a xenograft mouse model. These results indicate that 8-methoxybutin has potential as a therapeutic agent for hyperpigmentation disorder and skin cancer. SIGNIFICANCE STATEMENT: 8-Methoxybutin inhibits MITF transactivation activity resulting suppression of melanogenesis and skin melanoma growth.

Three New Phthalide Glycosides from the Rhizomes of Cnidium officinale and Their Recovery Effect on Damaged Otic Hair Cells in Zebrafish.

The extract from Cnidium officinale rhizomes was shown in a prior experiment to markedly recover otic hair cells in zebrafish damaged by neomycin. The current study was brought about to identify the principal metabolite. Column chromatography using octadecyl SiO2 and SiO2 was performed to isolate the major metabolites from the active fraction. The chemical structures were resolved on the basis of spectroscopic data, including NMR, IR, MS, and circular dichroism (CD) data. The isolated phthalide glycosides were assessed for their recovery effect on damaged otic hair cells in neomycin-treated zebrafish. Three new phthalide glycosides were isolated, and their chemical structures, including stereochemical characteristics, were determined. Two glycosides (0.1 µM) showed a recovery effect (p < 0.01) on otic hair cells in zebrafish affected by neomycin ototoxicity. Repeated column chromatography led to the isolation of three new phthalide glycosides, named ligusticosides C (1), D (2), and E (3). Ligusticoside C and ligusticoside E recovered damaged otic hair cells in zebrafish.

Whitening and inhibiting NF-κB-mediated inflammation properties of the biotransformed green ginseng berry of new cultivar K1, ginsenoside Rg2 enriched, on B16 and LPS-stimulated RAW 264.7 cells.

BACKGROUND: Main bioactive constituents and pharmacological functions of ripened red ginseng berry (Panax ginseng Meyer) have been frequently reported. Yet, the research gap targeting the beneficial activities of transformed green ginseng berries has not reported elsewhere. METHODS: Ginsenosides of new green berry cultivar K-1 (GK-1) were identified by HPLC-QTOF/MS. Ginsenosides bioconversion in GK-1 by bgp1 enzyme was confirmed with HPLC and TLC. Then, mechanisms of GK-1 and ß-glucosidase (bgp1) biotransformed GK-1 (BGK-1) were determined by Quantitative Reverse Transcription-Polymerase Chain Reaction and Western blot. RESULTS: GK-1 possesses highest ginsenosides especially ginsenoside-Re amongst seven ginseng cultivars including (Chunpoong, Huangsuk, Kumpoong, K-1, Honkaejong, Gopoong, and Yunpoong). Ginseng root's biomass is not affected with the harvest of GK-1 at 3 weeks after flowering period. Then, Re is bio-converted into a promising pharmaceutical effect of Rg2 via bgp1. According to the results of cell assays, BGK-1 shows decrease of tyrosinase and melanin content in α-melanocyte-stimulating hormone challenged-murine melanoma B16 cells. BGK-1 which is comparatively more effective than GK-1 extract shows significant suppression of the nuclear factor (NF)-κB activation and inflammatory target genes, in LPS-stimulated RAW 264.7 cells. CONCLUSION: These results reported effective whitening and anti-inflammatory of BGK-1 as compared to GK-1.

Aurones and Flavonols from Coreopsis lanceolata L. Flowers and Their Anti-Oxidant, Pro-Inflammatory Inhibition Effects, and Recovery Effects on Alloxan-Induced Pancreatic Islets in Zebrafish.

(1) Background: Many flavonoids have been reported to exhibit pharmacological activity; a preparatory study confirmed that Coreopsis lanceolata flowers (CLFs) contained high flavonoid structure content; (2) Methods: CLFs were extracted in aqueous methanol (MeOH:H2O = 4:1) and fractionated into acetic ester (EtOAc), normal butanol (n-BuOH), and H2O fractions. Repeated column chromatographies for two fractions led to the isolation of two aurones and two flavonols; (3) Results: Four flavonoids were identified based on a variety of spectroscopic data analyses to be leptosidin (1), leptosin (2), isoquercetin (3), and astragalin (4), respectively. This is the first report for isolation of 2-4 from CLFs. High-performance liquid chromatography (HPLC) analysis determined the content levels of compounds 1-4 in the MeOH extract to be 2.8 ± 0.3 mg/g (1), 17.9 ± 0.9 mg/g (2), 3.0 ± 0.2 mg/g (3), and 10.9 ± 0.9 mg/g (4), respectively. All isolated compounds showed radical scavenging activities and recovery activities in Caco-2, RAW264.7, PC-12, and HepG2 cells against reactive oxygen species. MeOH extract, EtOAc fraction, and 1-3 suppressed NO formation in LPS-stimulated RAW 264.7 cells and decreased iNOS and COX-2 expression. Furthermore, all compounds recovered the pancreatic islets damaged by alloxan treatment in zebrafish; (4) Conclusions: The outcome proposes 1-4 to serve as components of CLFs in standardizing anti-oxidant, pro-inflammatory inhibition, and potential anti-diabetic agents.

Noble 3,4-Seco-triterpenoid Glycosides from the Fruits of Acanthopanax sessiliflorus and Their Anti-Neuroinflammatory Effects.

Acanthopanax sessiliflorus (Araliaceae) have been reported to exhibit many pharmacological activities. Our preliminary study suggested that A. sessiliflorus fruits include many bioactive 3,4-seco-triterpenoids. A. sessiliflorus fruits were extracted in aqueous EtOH and fractionated into EtOAc, n-BuOH, and H2O fractions. Repeated column chromatographies for the organic fractions led to the isolation of 3,4-seco-triterpenoid glycosides, including new compounds. Ultra-high-performance liquid chromatography (UPLC) mass spectrometry (MS) systems were used for quantitation and quantification. BV2 and RAW264.7 cells were induced by LPS, and the levels of pro-inflammatory cytokines and mediators and their underlying mechanisms were measured by ELISA and Western blotting. NMR, IR, and HR-MS analyses revealed the chemical structures of the nine noble 3,4-seco-triterpenoid glycosides, acanthosessilioside G-O, and two known ones. The amounts of the compounds were 0.01-2.806 mg/g, respectively. Acanthosessilioside K, L, and M were the most effective in inhibiting NO, PGE2, TNF-α, IL-1ß, and IL-6 production and reducing iNOS and COX-2 expression. In addition, it had inhibitory effects on the LPS-induced p38 and ERK MAPK phosphorylation in both BV2 and RAW264.7 cells. Nine noble 3,4-seco-triterpenoid glycosides were isolated from A. sessiliflorus fruits, and acanthosessilioside K, L, and M showed high anti-inflammatory and anti-neuroinflammatory effects.

Glycosyltransformation of ginsenoside Rh2 into two novel ginsenosides using recombinant glycosyltransferase from Lactobacillus rhamnosus and its in vitro applications.

BACKGROUND: Ginsenoside Rh2 is well known for many pharmacological activities, such as anticancer, antidiabetes, antiinflammatory, and antiobesity properties. Glycosyltransferases (GTs) are ubiquitous enzymes present in nature and are widely used for the synthesis of oligosaccharides, polysaccharides, glycoconjugates, and novel derivatives. We aimed to synthesize new ginsenosides from Rh2 using the recombinant GT enzyme and investigate its cytotoxicity with diverse cell lines. METHODS: We have used a GT gene with 1,224-bp gene sequence cloned from Lactobacillus rhamnosus (LRGT) and then expressed in Escherichia coli BL21 (DE3). The recombinant GT protein was purified and demonstrated to transform Rh2 into two novel ginsenosides, and they were characterized by nuclear magnetic resonance (NMR) techniques and evaluated by 3-(4, 5-dimethylthiazol-2-yl)-2-5-diphenyltetrazolium bromide assay. RESULTS: Two novel ginsenosides with an additional glucopyranosyl (6→1) and two additional glucopyranosyl (6→1) linked with the C-3 position of the substrate Rh2 were synthesized, respectively. Cell viability assay in the lung cancer (A549) cell line showed that glucosyl ginsenoside Rh2 inhibited cell viability more potently than ginsenoside Rg3 and Rh2 at a concentration of 10 µM. Furthermore, glucosyl ginsenoside Rh2 did not exhibit any cytotoxic effect in murine macrophage cells (RAW264.7), mouse embryo fibroblasts cells (3T3-L1), and skin cells (B16BL6) at a concentration of 10 µM compared with ginsenoside Rh2 and Rg3. CONCLUSION: This is the first report on the synthesis of two novel ginsenosides, namely, glucosyl ginsenoside Rh2 and diglucosyl ginsenoside Rh2 from Rh2 by using recombinant GT isolated from L. rhamnosus. Moreover, diglucosyl ginsenoside Rh2 might be a new candidate for treatment of inflammation, obesity, and skin whiting, and especially for anticancer.

Volatile Profiles of Five Variants of Abeliophyllum distichum Flowers Using Headspace Solid-Phase Microextraction Gas Chromatography-Mass Spectrometry (HS-SPME-GC-MS) Analysis.

Abeliophyllum distichum (Oleaceae), which is the only species in the monotypic genus and is grown only on the Korean peninsula, has a high scarcity value. Its five variants (white, pink, round, blue, and ivory) have different morphological characteristics in terms of the color of petals and sepals or shape of the fruits. Despite its high value, there has been no study on variant classification except in terms of their morphological characteristics. Thus, we performed a volatile component analysis of A. distichum flowers and multivariate data analyses to reveal the relationship between fragments emitted from five variants of A. distichum flowers with their morphological characteristics. As a result, 66 volatile components of this plant were identified by headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS), showing unique patterns for each set of morphological characteristics, especially the color of the petals. These results suggest that morphological characteristics of each variant are related to the volatile composition.

Correction to: A brief history and spectroscopic analysis of soy isoflavones.

[This corrects the article DOI: 10.1007/s10068-020-00815-6.].

A brief history and spectroscopic analysis of soy isoflavones.

The production of soybean continues to increase worldwide. People are showing more interest in the beneficial health effects of soybeans than before. However, the origin and history of soybeans are still being discussed among many researchers. Chromatographic methods enable the desirable separation of a variety of isoflavones from soybeans. The structures of isolated soy isoflavones have been successfully identified in tandem with spectroscopic analytical instruments and technologies such as liquid chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. The theoretical background behind spectroscopy may help improve the understanding for the analysis of isoflavones in soybeans and soy-derived foods. This review covers the origin of the English name of soybean and its scientific name, Glycine max (L.) Merrill, based on the evidence reported to date. Moreover, the reports of soy isoflavones discovered over a period of about 100 years have been briefly reviewed.

Using Amylosucrase for the Controlled Synthesis of Novel Isoquercitrin Glycosides with Different Glycosidic Linkages.

Many attempts have been made to obtain natural products with certain glycosidic linkages for improvement of their chemo-physical characteristics. Amylosucrase from Deinococcus geothermalis (DGAS; EC.4.2.1.4) is able to transglycosylate natural products. A model compound, isoquercitrin (IQ; quercetin-3-O-glucoside), was employed for producing new IQ glucosides (IQ-Gs). Treatment of IQ with DGAS produced monoglucoside (IQ-G1'), diglucosides (IQ-G2' and IQ-G2″), and triglucoside (IQ-G3). Structural analysis by mass and nuclear magnetic resonance spectrometry revealed that three of the four IQ-Gs were unreported new compounds possessing α-1,2-, α-1,4-, and/or α-1,6-glucosidic linkages at the 3-O-glucosyl moiety of IQ. IQ-G2' and IQ-G3 were dominantly produced at pH 5.0 and 7.2 and 1500 and 100 mM sucrose, respectively (yields of total IQ-Gs: 50-97%). Kinetic studies indicated that the production rate was dependent on buffer/pH and sucrose concentration. The diverse transglycosylations were verified with a molecular docking simulation. This study sheds light on methods for simple glycodiversification of natural products using DGAS, which can synthesize diversely branched glycosides by modulating reaction conditions.

Amylosucrase from Deinococcus geothermalis can be modulated under different reaction conditions to produce novel quercetin 4'-O-α-d-isomaltoside.

Amylosucrase (ASase, EC.4.2.1.4) is well-known for its distinguishable property of transglycosylation of many flavonoids and phenolics. Quercetin has diverse biological functions, however, its use is limited due to poor solubility and bioavailability. ASase derived from Deinococcus geothermalis (DGAS) showed conditional preference for producing unusual quercetin glucosides (QGs). DGAS produced a variety of QGs including quercetin monoglucosides (QG1), diglucosides (QG2 and QG2'), and triglucoside from quercetin and sucrose. The newly synthesized QG2' was recognized as a novel quercetin isomaltoside with an α-1,6 linkage branched at the -OH of C4' in quercetin by mass and nuclear magnetic resonance spectra. With a higher conversion yield from quercetin to QGs (60-92%), the optimum conditions for producing QG2' were examined under various pH and sucrose concentrations by response surface methodology. QG2' was predominantly produced under acidic conditions (pH 5.0) and at high sucrose concentrations (1000-1500 mM). In contrast, QG1 was generated as an intermediate of consecutive glycosylation. Kinetic evaluations indicated that considerable differences of transglycosylation velocities were caused by the pH and buffer salts of the reaction, which had a 3.9-fold higher overall performance (kcat/K'm) of generating QG2' at pH 5 compared to at pH 7. A rationale of unusual transglycosylations was demonstrated with a molecular docking simulation. Taken together, our study demonstrated that ASase can be used to synthesize unusually branched flavonoid glycosides from flavonol aglycones with clear patterns by modulating reaction conditions.

Syringoleosides A-H, Secoiridoids from Syringa dilatata Flowers and Their Inhibition of NO Production in LPS-Induced RAW 264.7 Cells.

Repeated column chromatography of Syringa dilatata flowers, a native shrub to Korea, led to the isolation of eight new oleoside-type secoiridoids, syringoleosides A-H (1-8), as well as five known secoiridoids (9-13). The new chemical structures were identified through spectroscopic data analysis, as well as the application of chemical methods. Compounds 1, 2, 6, 7, 11, and 13 showed suppression effects on NO production in LPS-induced RAW 264.7 cells, with IC50 values ranging from 32.5 ± 9.8 to 65.7 ± 11.0 µM, and no visible toxicity. The content of the major secoiridoids in S. dilatata flowers, compounds 1, 4, 5, 8, 9, 12, and 13, were determined through HPLC analysis.

Critical enzymes for biosynthesis of cucurbitacin derivatives in watermelon and their biological significance.

Various cucurbitacins have been isolated, and their structures have been elucidated. Owing to their economic potential and importance as active pharmacological compounds, their cytotoxicity in various cancer cells has been assessed. Here, we mined several candidate genes with potential involvement in cucurbitacin biosynthesis in watermelon (Citrullus lanatus) and performed in vitro enzymatic assays and instrumental analyses using various substrates to identify cucurbitacin functions and products. Enzymatic activities of two acetyltransferases (ACTs) and one UDP-glucosyltransferase (UGT) against cucurbitacins were confirmed, resulting in the synthesis of novel cucurbitacins in vivo and/or in vitro to our knowledge. As ACTs and UGT are involved in the dynamic conversion of cucurbitacins by catalyzing acetylation and glucosylation at moieties in the cucurbitacins skeleton, these findings improve our knowledge on how these genes contribute to the diversity of cucurbitacins.