Dietary flavonoid myricetin inhibits invasion and migration of radioresistant lung cancer cells (A549-IR) by suppressing MMP-2 and MMP-9 expressions through inhibition of the FAK-ERK signaling pathway.

Myricetin is a commonly found dietary flavonoid. In the present study, we investigated the effects of myricetin on migration and invasion of radioresistant lung cancer cells (A549-IR). Transcriptome analysis of A549-IR cells identified several differentially expressed genes (DEGs) in A549-IR cells compared to parental A549 cells. Functional enrichment analysis revealed that most of the DEGs were linked with PI3K-AKT signaling, proteoglycans, focal adhesion, and ECM-receptor interactions. A549-IR cells demonstrated enhanced migratory potential with increased expression of vimentin, snail and slug, and reduced expression of E-cadherin. A549-IR cells exposed to myricetin displayed reduced migration and suppressed MMP-2 and MMP-9 expression. Notably, myricetin inhibited the phosphorylation of focal adhesion kinase (FAK) and altered the F-actin/G-actin ratio in A549-IR cells, without modulation of EMT markers. These findings suggest that myricetin can inhibit migration of A549-IR cells by suppressing MMP-2 and MMP-9 expressions through inhibition of the FAK-ERK signaling pathway.

Supercritical Fluid Extraction of Citrus iyo Hort. ex Tanaka Pericarp Inhibits Growth and Induces Apoptosis Through Abrogation of STAT3 Regulated Gene Products in Human Prostate Cancer Xenograft Mouse Model.

Activation of signal transducer and activator of transcription 3 (STAT3) is well known to play a major role in the cell growth, survival, proliferation, metastasis, and angiogenesis of various cancer cells. Most of the citrus species offer large quantities of phytochemicals that have beneficial effects attributed to their chemical components. Our study was carried out to evaluate the anticancer effects of the pericarp of Iyokan ( Citrus iyo Hort. ex Tanaka), locally known as yeagam in Korea, through modulation of the STAT3 signaling pathway in both tumor cells and a nude mice model. The effect of supercritical extracts of yeagam peel (SEYG) on STAT3 activation, associated protein kinases, STAT3-regulated gene products, cellular proliferation, and apoptosis was examined. The in vivo effect of SEYG on the growth of DU145 human prostate xenograft tumors in athymic nu/nu male mice was also investigated. We found SEYG exerted substantial inhibitory effect on STAT3 activation in human prostate cancer DU145 cells as compared to other tumor cells analyzed. SEYG inhibited proliferation and downregulated the expression of various STAT3-regulated gene products such as bcl-2, bcl-xL, survivin, IAP-1/2, cyclin D1, cyclin E, COX-2, VEGF, and MMP-9. This correlated with an increase in apoptosis as indicated by an increase in the expression of p53 and p21 proteins, the sub-G1 arrest, and caspase-3-induced PARP cleavage. When administered intraperitoneally, SEYG reduced the growth of DU145 human prostate xenograft tumors through downmodulation of STAT3 activation in athymic nu/nu male mice. Overall, these results suggest that SEYG extract has the potential source of STAT3 inhibitors that may have a potential in chemoprevention of human prostate cancer cells.

Anti-inflammatory effects of an ethanolic extract of guava (Psidium guajava L.) leaves in vitro and in vivo.

Plant extracts have been used as a source of medicines for a wide variety of human ailments. Among the numerous traditional medicinal herbs, Psidium guajava L. (Myrtaceae), commonly known as guava, has long been used in folk medicines as a therapeutic agent for the treatment of numerous diseases in East Asian and other countries. The aim of this study was to investigate the anti-inflammatory activity of an ethanolic leaf extract of P. guajava (guava) in vitro and in vivo. Our results demonstrated that guava leaf extract (GLE) significantly inhibited lipopolysaccharide (LPS)-induced production of nitric oxide and prostaglandin E2 in a dose-dependent manner. GLE suppressed the expression and activity of both inducible nitric oxide synthase and cyclooxygenase-2 in part through the downregulation of ERK1/2 activation in RAW264.7 macrophages. Furthermore, GLE exhibited significant anti-inflammatory activity in 2 different animal models-Freund's complete adjuvant-induced hyperalgesia in the rat and LPS-induced endotoxic shock in mice.

Anti-metastatic effect of supercritical extracts from the Citrus hassaku pericarp via inhibition of C-X-C chemokine receptor type 4 (CXCR4) and matrix metalloproteinase-9 (MMP-9).

The fruit of hassaku (Citrus hassaku Hort. ex Tanaka) is locally known as phalsak in Korea. Recently, the fruit extract has been known to exhibit in vivo preventive effects against UVB-induced pigmentation, antiallergic activity, and enhancement of blood fluidity. However, the exact mechanisms of how supercritical extracts of phalsak peel (SEPS) inhibits tumor metastasis and invasion are still not fully understood. We found that SEPS could downregulate the constitutive expression of both CXCR4 and HER2 in human breast cancer MDA-MB-231 cells as compared with other cells. SEPS also suppressed matrix metalloproteinase-9 (MMP-9) expression and its enzymatic activity under non-cytotoxic concentrations. Neither proteasome inhibition nor lysosomal stabilization had any effect on the SEPS-induced decrease in CXCR4 expression. A detailed study of the underlying molecular mechanisms revealed that the regulation of the downregulation of CXCR4 was at the transcriptional level, as indicated by downregulation of mRNA expression, suppression of NF-κB activity, and inhibition of chromatin immunoprecipitation activity. Suppression of CXCR4 expression by SEPS correlated with the inhibition of CXCL12-stimulated invasion of MDA-MB-231 cells. Overall, our results indicate, for the first time, that SEPS can suppress CXCR4 and MMP-9 expressions through blockade of NF-κB activation and thus has the potential to suppress metastasis of breast cancer.

ß-Caryophyllene oxide inhibits constitutive and inducible STAT3 signaling pathway through induction of the SHP-1 protein tyrosine phosphatase.

Constitutive activation of STAT3 is frequently observed and closely linked with proliferation, survival, invasion, metastasis and angiogenesis in tumor cells. In the present study, we investigated whether ß-caryophyllene oxide (CPO), a sesquiterpene isolated primarily from the essential oils of medicinal plants such as guava (Psidium guajava), and oregano (Origanum vulgare L.), can mediate its effect through interference with the STAT3 activation pathway in cancer cells. The effect of CPO on STAT3 activation, associated protein kinases and phosphatase, STAT3-regulated gene products and apoptosis was investigated using both functional proteomics tumor pathway technology platform and different tumor cell lines. We found that CPO suppressed constitutive STAT3 activation in multiple myeloma (MM), breast and prostate cancer cell lines, with a significant dose- and time-dependent effects observed in MM cells. The suppression was mediated through the inhibition of activation of upstream kinases c-Src and JAK1/2. Also, vanadate treatment reversed CPO-induced down-regulation of STAT3, suggesting the involvement of a tyrosine phosphatase. Indeed, we found that CPO induced the expression of tyrosine phosphatase SHP-1 that correlated with the down-regulation of constitutive STAT3 activation. Interestingly, deletion of SHP-1 gene by siRNA abolished the ability of CPO to inhibit STAT3 activation. The inhibition of STAT3 activation by CPO inhibited proliferation, induced apoptosis and abrogated the invasive potential of tumor cells. Our results suggest for the first time that CPO is a novel blocker of STAT3 signaling cascade and thus has an enormous potential for the treatment of various cancers harboring constitutively activated STAT3.

ß-Caryophyllene oxide potentiates TNFα-induced apoptosis and inhibits invasion through down-modulation of NF-κB-regulated gene products.

We have recently reported that ß-caryophyllene oxide (CPO) can induce apoptosis, suppress tumor growth, and inhibit metastasis through the suppression of signal transducer and activator of transcription 3, PI3K/AKT/mTOR/S6K1 signaling cascades and ROS-mediated MAPKs activation. In the present study, we found that CPO potentiated the apoptosis induced by tumor necrosis factor α (TNFα) and chemotherapeutic agents, suppressed TNFα-induced tumor cell invasion, all of which are known to require NF-κB activation. We found that TNFα stimulated the expression of gene products involved in anti-apoptosis (IAP1, IAP2, Bcl-2, Bcl-xL, and survivin), proliferation (COX-2, cyclin D1, and c-Myc), invasion (MMP 9 and ICAM-1), and angiogenesis (VEGF) and that CPO treatment suppressed their expression. Because these gene products are also regulated by proinflammatory transcription factor NF-κB, we postulated that CPO may mediate its effects by modulating the NF-κB pathway. We found that CPO blocked both inducible and constitutive NF-κB activation in a wide variety of tumor cells. CPO was also found to inhibit the TNFα-induced degradation of IκBα through the inhibition of activation of IκBα kinase and p65 nuclear translocation and phosphorylation. Interestingly, CPO failed to potentiate the apoptotic effect induced by TNFα in p65 (-/-) cells as compared to the wild-type. Thus, overall, our results indicate that the inhibition of NF-κB is one of major mechanisms by which CPO enhances TNFα-induced apoptosis and suppresses invasion.

Capillarisin inhibits iNOS, COX-2 expression, and proinflammatory cytokines in LPS-induced RAW 264.7 macrophages via the suppression of ERK, JNK, and NF-κB activation.

The aerial parts of Artemisia capillaris (Compositae) have been used in traditional Korean medicine as a cholagogic, antipyretic, anti-inflammatory, and diuretic purposes. In our previous study, ethanolic extracts of the plant demonstrated a marked anti-inflammatory effect in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells (J. Korean Soc. Appl. Biol. Chem., 2010, 53, 275-282). In the present study, capillarisin (CPS), a flavone, main constituent of A. capillaris, was examined for its anti-inflammatory activity in the cells. We found that CPS highly suppressed LPS-induced nitric oxide (NO) without exerting cytotoxic effects on RAW 264.7 cells. CPS inhibited the expression of LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein and their mRNA in a dose-dependent manner. Also, tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1ß, and prostaglandin E(2) (PGE(2)) secretion were decreased by CPS in LPS-stimulated macrophages. As a result, CPS inhibited proinflammatory cytokines, iNOS, and COX-2, which is attributed to the suppression of LPS-induced ERK, JNK, and nuclear factor-κB (NF-κB) activation. Therefore, we demonstrate here that CPS potentially inhibits the biomarkers related to inflammation through the abrogation of ERK, JNK, and NF-κB p65 activation, and it may be a potential therapeutic candidate for the treatment of inflammatory diseases.

Functional regulation of Slug/Snail2 is dependent on GSK-3ß-mediated phosphorylation.

Snail family proteins regulate transcription of molecules for cell-cell adhesion during epithelial-mesenchymal transition (EMT). Based on putative glycogen synthase kinase 3ß (GSK-3ß) phosphorylation sites within the Slug/Snail2, we explored the significance of GSK-3ß-mediated phosphorylation in Slug/Snail2 expression during EMT. Mutation of the putative GSK-3ß phosphorylation sites (S92/96A or S100/104A) enhanced the Slug/Snail2-mediated EMT properties of E-cadherin repression and vimentin induction, compared with wild-type Slug/Snail2. S92/96A mutation inhibited degradation of Slug/Snail2 and S100/104A mutation extended nuclear stabilization. Inhibition of GSK-3ß activity caused similar effects, as did the phosphorylation mutations. Thus, our study suggests that GSK-3ß-mediated phosphorylation of Slug/Snail2 controls its turnover and localization during EMT.

A hexane fraction of guava Leaves (Psidium guajava L.) induces anticancer activity by suppressing AKT/mammalian target of rapamycin/ribosomal p70 S6 kinase in human prostate cancer cells.

This study was carried out to evaluate the anticancer effects of guava leaf extracts and its fractions. The chemical compositions of the active extracts were also determined. In the present study, we set out to determine whether the anticancer effects of guava leaves are linked with their ability to suppress constitutive AKT/mammalian target of rapamycin (mTOR)/ribosomal p70 S6 kinase (S6K1) and mitogen-activated protein kinase (MAPK) activation pathways in human prostate cancer cells. We found that guava leaf hexane fraction (GHF) was the most potent inducer of cytotoxic and apoptotic effects in PC-3 cells. The molecular mechanism or mechanisms of GHF apoptotic potential were correlated with the suppression of AKT/mTOR/S6K1 and MAPK signaling pathways. This effect of GHF correlated with down-regulation of various proteins that mediate cell proliferation, cell survival, metastasis, and angiogenesis. Analysis of GHF by gas chromatography and gas chromatography-mass spectrometry tentatively identified 60 compounds, including ß-eudesmol (11.98%), α-copaene (7.97%), phytol (7.95%), α-patchoulene (3.76%), ß-caryophyllene oxide (CPO) (3.63%), caryophylla-3(15),7(14)-dien-6-ol (2.68%), (E)-methyl isoeugenol (1.90%), α-terpineol (1.76%), and octadecane (1.23%). Besides GHF, CPO, but not phytol, also inhibited the AKT/mTOR/S6K1 signaling pathway and induced apoptosis in prostate cancer cells. Overall, these findings suggest that guava leaves can interfere with multiple signaling cascades linked with tumorigenesis and provide a source of potential therapeutic compounds for both the prevention and treatment of cancer.

Antiproliferative effects of Dangyuja (Citrus grandis Osbeck) leaves through suppression of constitutive signal transducer and activator of transcription 3 activation in human prostate carcinoma DU145 cells.

Although Dangyuja (Citrus grandis Osbeck) exhibits anti-inflammatory and anticancer activities, its molecular targets and pathways, especially in human prostate cancer cells, are not fully understood. In this study, the antiproliferative effect of Dangyuja leaves through the signal transducer and activator of transcription (STAT) 3 signaling pathway was investigated in human prostate carcinoma DU145 cells. The solvent fractions (n-hexane, chloroform, ethyl acetate, and n-butanol) were obtained from a crude extract (80% methanol extract) of Dangyuja leaves. We first found that the chloroform fraction of Dangyuja leaves (DCF) was the most cytotoxic against DU145 cells. DCF inhibited constitutive STAT3 activation through blocking upstream Janus-like kinase 2 and c-Src. Consistent with STAT3 inactivation, DCF down-regulated the expression of STAT3 target genes, including bcl-2, bcl-xl, and cyclin D1; this correlated with the suppression of proliferation, the accumulation of cell cycle at the sub-G(1) phase, and the induction of apoptosis. Furthermore, DCF exerted a relatively minor effect on the growth of human prostate noncancerous RWPE-1 cells. Nobiletin, a major active constituent of DCF, could induce apoptosis via the suppression of constitutive STAT3 activation. Overall, our results indicate that the anti-inflammatory and anticancer activities previously assigned to DCF may be mediated partially through the suppression of the STAT3 signaling.

The butanol fraction of guava (Psidium cattleianum Sabine) leaf extract suppresses MMP-2 and MMP-9 expression and activity through the suppression of the ERK1/2 MAPK signaling pathway.

The leaf extract of guava (Psidium cattleianum Sabine) has traditionally been used for the treatment of diarrhea and diabetes in East Asia and other countries. Recently, the leaf extract has been employed in the therapy of cancer, bacterial infections, and inflammation in experimental models. However, the exact mechanisms of how guava leaf extract inhibits tumor metastasis and invasion are still unknown. In the present study, we investigated in detail the molecular mechanism(s) responsible for the potential antimetastatic and antiinvasive effects of the butanol fraction of guava leaf extract (GBF). Interestingly, we observed for the first time that GBF suppressed both matrix metalloproteinases (MMP)-9 and MMP-2 expression and activity in part through the downregulation of the ERK1/2 activation in lung cancer cells. Also, importantly, the major components of the GBF were identified as d-glucuronic acid, quercetin 3-glucuronide, loganin, and xanthyletin by LC-ESI-MS/MS. Collectively, our data indicate that the guava leaf could reduce the metastasis of lung cancer cells and therefore suggest that it could be advantageously used to control the metastatic process.

Pepsin-solubilised collagen (PSC) from Red Sea cucumber (Stichopus japonicus) regulates cell cycle and the fibronectin synthesis in HaCaT cell migration.

Pepsin-solubilised collagen (PSC) from Red Sea cucumber (Stichopus japonicus) was studied with respect to its wound-healing effects on a human keratinocyte (HaCaT) cell line. Disaggregated collagen fibres were treated with 0.1M NaOH for 24h and digested with pepsin for 72h to reach maximum yield of 26.6%. The results of an in vitro wound-healing test showed that migration of HaCaT cells was 1.5-fold faster on PSC-coated plates than on untreated plates. The migration rate of sea cucumber PSC was similar to that of rat PSC, but five times higher than that of bovine gelatin. HaCaT cells grown on PSC-coated plates revealed increased fibronectin synthesis (6-fold and 3-fold compared to gelatin and rat PSC, respectively). Additionally, sea cucumber PSCs induced HaCaT cell proliferation by decreasing the G1 phase by 5% and maintaining a larger population (8%) of cells in mitosis. Collagen from Red Sea cucumber might be useful as an alternative to mammalian collagen in the nutraceutical and pharmaceutical industries.

ß-Caryophyllene oxide inhibits growth and induces apoptosis through the suppression of PI3K/AKT/mTOR/S6K1 pathways and ROS-mediated MAPKs activation.

Both PI3K/AKT/mTOR/S6K1 and mitogen activated protein kinase (MAPK) signaling cascades play an important role in cell proliferation, survival, angiogenesis, and metastasis of tumor cells. In the present report, we investigated the effects of ß-caryophyllene oxide (CPO), a sesquiterpene isolated from essential oils of medicinal plants such as guava (Psidium guajava), oregano (Origanum vulgare L.), cinnamon (Cinnamomum spp.) clove (Eugenia caryophyllata), and black pepper (Piper nigrum L.) on the PI3K/AKT/mTOR/S6K1 and MAPK activation pathways in human prostate and breast cancer cells. We found that CPO not only inhibited the constitutive activation of PI3K/AKT/mTOR/S6K1 signaling cascade; but also caused the activation of ERK, JNK, and p38 MAPK in tumor cells. CPO induced increased reactive oxygen species (ROS) generation from mitochondria, which is associated with the induction of apoptosis as characterized by positive Annexin V binding and TUNEL staining, loss of mitochondrial membrane potential, release of cytochrome c, activation of caspase-3, and cleavage of PARP. Inhibition of ROS generation by N-acetylcysteine (NAC) significantly prevented CPO-induced apoptosis. Subsequently, CPO also down-regulated the expression of various downstream gene products that mediate cell proliferation (cyclin D1), survival (bcl-2, bcl-xL, survivin, IAP-1, and IAP-2), metastasis (COX-2), angiogenesis (VEGF), and increased the expression of p53 and p21. Interestingly, we also observed that CPO can significantly potentiate the apoptotic effects of various pharmacological PI3K/AKT inhibitors when employed in combination in tumor cells. Overall, these findings suggest that CPO can interfere with multiple signaling cascades involved in tumorigenesis and used as a potential therapeutic candidate for both the prevention and treatment of cancer.

Metabolic profiling and predicting the free radical scavenging activity of guava (Psidium guajava L.) leaves according to harvest time by 1H-nuclear magnetic resonance spectroscopy.

Guava leaves were classified and the free radical scavenging activity (FRSA) evaluated according to different harvest times by using the (1)H-NMR-based metabolomic technique. A principal component analysis (PCA) of (1)H-NMR data from the guava leaves provided clear clusters according to the harvesting time. A partial least squares (PLS) analysis indicated a correlation between the metabolic profile and FRSA. FRSA levels of the guava leaves harvested during May and August were high, and those leaves contained higher amounts of 3-hydroxybutyric acid, acetic acid, glutamic acid, asparagine, citric acid, malonic acid, trans-aconitic acid, ascorbic acid, maleic acid, cis-aconitic acid, epicatechin, protocatechuic acid, and xanthine than the leaves harvested during October and December. Epicatechin and protocatechuic acid among those compounds seem to have enhanced FRSA of the guava leaf samples harvested in May and August. A PLS regression model was established to predict guava leaf FRSA at different harvesting times by using a (1)H-NMR data set. The predictability of the PLS model was then tested by internal and external validation. The results of this study indicate that (1)H-NMR-based metabolomic data could usefully characterize guava leaves according to their time of harvesting.

Antiviral activity of methylelaiophylin, an alpha-glucosidase inhibitor.

Methylelaiophylin isolated from Streptomyces melanosporofaciens was selected as an alpha-glucosidase inhibitor with an IC50 value of 10 micrometer. It showed mixedtype inhibition of alpha-glucosidase with a Ki value of 5.94 micrometer. In addition, methylelaiophylin inhibited the intracellular trafficking of hemagglutinin-neuramidase (HN), a glycoprotein of Newcastle disease virus (NDV), in baby hamster kidney (BHK) cells. Methylelaiophylin inhibited the cell surface expression of NDV-HN glycoprotein without significantly affecting HN glycoprotein synthesis in NDV-infected BHK cells.

Proteasome inhibition causes epithelial-mesenchymal transition upon TM4SF5 expression.

Transmembrane 4 L six family member 5 (TM4SF5) is highly expressed in hepatocarcinoma and causes epithelial-mesenchymal transition (EMT) of hepatocytes. We found that TM4SF5-expressing cells showed lower mRNA levels but maintained normal protein levels in certain gene cases, indicating that TM4SF5 mediates stabilization of proteins. In this study, we explored whether regulation of proteasome activity and TM4SF5 expression led to EMT. We observed that TM4SF5 expression caused inhibition of proteasome activity and proteasome subunit expression, causing morphological changes and loss of cell-cell contacts. shRNA against TM4SF5 recovered proteasome expression, with leading to blockade of proteasome inactivation and EMT. Altogether, TM4SF5 expression appeared to cause loss of cell-cell adhesions via proteasome suppression and thereby proteasome inhibition, leading to repression of cell-cell adhesion molecules, such as E-cadherin.

Distribution and biosynthesis of 20-hydroxyecdysone in plants of Achyranthes japonica Nakai.

There is increasing interest in phytoecdysteroids (PEs) because of their potential role in plant defense against insects. To understand the mechanism regulating their levels in plants, the fluctuation, distribution, and biosynthesis of PE 20-hydroxyecdysone (20E) examined in Achyranthes japonica. The total amount of 20E per individual plant initially remained at a constant level, and increased markedly after the first leaf pair (LP) stage, while the concentration of 20E in a given plant decreased rapidly during vegetative growth. In addition, the incorporation of [2-(14)C]-mevalonic acid into 20E did not differ significantly depending on plant organs and developmental stages, suggesting that biosynthesis of 20E is not restricted to particular organs or growth stages.

Nonactin hinders intracellular glycosylation in virus-infected baby hamster kidney cells.

Potent antiviral agents hinder virus-infected cell machinery, leading to rescue from viral damage. In this study, we aimed to identify selective intracellular glycosylation inhibitor(s) that do not suppress glycoprotein synthesis. Our results showed that nonactin is a potent inhibitor of intracellular glycosylation. First, we examined the effects of nonactin on syncytium formation and cytopathic activity in virus-infected baby hamster kidney cells. Nonactin effectively inhibited syncytium formation in a concentration-dependent manner, and infectious virus production was markedly reduced. However, glycoprotein synthesis was not affected. In the presence of 5 µg/ml nonactin, we observed the intracellular accumulation of vesicular stomatitis virus-G protein as well as syncytium formation, but no significant effects on Newcastle disease virus-hemagglutinin-neuramidase glycoprotein synthesis. Our results collectively indicate that nonactin potentially inhibits glycosylation by acting as a suppressor of intracellular glycosylation trafficking.

Antibacterial activity of a disaccharide isolated from Streptomyces sp. strain JJ45 against Xanthomonas sp.

Of the 316 actinomycetes strains isolated from various habitats, Streptomyces sp. strain JJ45 showed the strongest antibiotic activity against the plant pathogenic bacteria Xanthomonas campestris pv. campestris and was thus chosen for further study. The 16S rRNA gene sequence (1500 bp) and rpoB gene partial sequence (306 bp) of Streptomyces strains JJ45A and JJ45B were determined. The respective strain JJ45B sequences exhibited 96.8% identity with the Streptococcus gelaticus 16S rRNA gene sequence and 98.4% identity with the Streptococcus vinaceus ATCC 27478 rpoB partial sequence. The fermentation broth of the JJ45B strain was extracted to find an inhibitor of bacterial growth. The distilled water extract showed the highest activity against pathogenic bacteria. The active molecule was isolated by column chromatography on polyacrylamide or silica gel, thin-layer chromatography, and HPLC. It showed growth inhibition activity only toward phytopathogenic Xanthomonas sp. The structure of the compound was identified as alpha-l-sorbofuranose (3-->2)-beta-D-altrofuranose based on the interpretation of the nuclear magnetic resonance spectra.

Classification and prediction of free-radical scavenging activities of dangyuja (Citrus grandis Osbeck) fruit extracts using (1)H NMR spectroscopy and multivariate statistical analysis.

Different parts of dangyuja (Citrus grandis Osbeck) fruits at different maturation stages were classified using a (1)H NMR-based metabolomic technique. Principal components analysis allowed the clear separation of fractions extracted with 50% methanol of different parts of dangyuja fruits at different maturation stages by combining principal components PC1 and PC2, which together accounted for 80.4% of the variance. A loading-plot analysis revealed that sucrose, glucose, oxaloacetic acid and citric acid were dominant in mature flesh, while naringin, tyramine, proline and alanine were dominant in immature fruit samples. Projections to latent structures using a partial least squares (PLS) model were used to predict the free-radical scavenging activities (FRSA) of dangyuja fruit extracts based on their (1)H NMR spectra. The present study suggests the usefulness of combining (1)H NMR spectroscopy with multivariate statistical analysis for discriminating dangyuja fruit samples, and predicting the FRSA of different parts of dangyuja fruit samples at different stages of maturation.