Isolation of quinone reductase (QR) inducing agents from ginger rhizome and their in vitro anti-inflammatory activity

To investigate the potential activity of ginger rhizome extracts to induce quinone reductase (QR), we performed bioactivity-guided fractionation using a murine hepatoma cell (Hepa 1c1c7) bioassay. Anti-inflammatory effects were then studied utilizing lipopolysaccharide (LPS)-stimulated mouse macrophage (RAW 264.7) cells. An ethyl acetate-partitioned fraction from ethanolic extract, rich in both QR inducing potency and anti-inflammatory activity, was subjected to repeated silica gel column and preparative thin layer chromatography to yield three compounds. The three isolated compounds were [6]-shogaol, 1-dehydro-[6]-gingerdione and hexahydrocurcumin. [6]-Dehydroshogaol, a minor component in ginger rhizome, was chemically synthesized and used for comparison in the subsequent bioassay based on its excellent QR inducing potency. Results showed that [6]-dehydroshogaol had the highest ability to induce QR activity (CD = 9.23 ± 0.22 ??M), followed by 1-dehydro-[6]-gingerdione (CD = 13.24 ± 0.45 ??M), and then hexahydrocurcumin (CD = 68.81 ± 3.90 ??M). Increasing QR activity in induced cells was associated with elevated expression of NQO-1 protein as confirmed by Western blot. [6]-Dehydroshogaol, [6]-shogaol and 1-dehydro-[6]-gingerdione were also potent inhibitors of nitric oxide (NO) synthesis in activated macrophages. Their IC₅₀ values ranged from 5.80 ± 1.27 to 25.06 ± 4.86 ??M. Hexahydrocurcumin exhibited the weakest inhibitory effect (IC₅₀ = 304.76 ± 54.80 ??M). These findings contribute to our theoretical understanding of the potential beneficial effects of consuming ginger as food and/or dietary supplement.     

Antioxidative activities of chromatographic fractions obtained from root,fruit and leaf of Mengkudu (Morinda citrifolia L.)

Crude extracts of root, leaf and fruit of Morinda citrifolia were fractionated on a Sephadex LH-20 column with ethanol as eluate. Based on UV absorption intensity of phenolic compound (725 mm) the Sephadex LH-20 column was able to separate fruit, leaf and root extracts into six, five and five fractions, respectively. The results showed that all the fractions tested exhibited considerably high antioxidative activity in the ferric thiocyanate assay and thiobarbituric acid test and the activities of some of the fractions were as good as those of either tocopherol or BHT. The fractions from different parts of the plants were found to contain different amounts of total phenolic compounds, which, interestingly, do not correspond to the antioxidative activity measured. This is probably due to the presence of different phenolics in the samples, with different antioxidative activities which involves various mechanisms inhibiting the oxidation process. The study suggested that root, leaf and fruit of M. citrifolia might contribute significantly to exogenous antioxidant which is crucial in combating oxidative stress.     

Flow injection spectrophotometry using natural reagent from Morinda citrifolia root for determination of aluminium in tea

A flow injection (FI) spectrophotometric method with using natural reagent extracted from Morinda citrifolia root has been developed for determination of aluminium. The extract contained anthraquinone compounds which could react with Al³⁺ to form reddish complexes which had maximum absorption wavelength at 499.0 nm. The extract could be used as a reagent in FI system without further purification to obtain pure compound. A sensitive method for determination of aluminium in concentration range of 0.1-1.0 mg L⁻¹, with detection limit of 0.05 mg L⁻¹ was achieved. Relative standard deviations of 1.2% and 1.7% were obtained for the determination of 0.1 and 0.6 mg L⁻¹ Al³⁺ (n = 11). Sample throughput of 35 h⁻¹ was achieved with the consumption of 3 mL each of carrier and reagent solutions per injection. The developed method was successfully applied to tea samples, validated by the FAAS standard method. The method is simple, fast, economical and could be classified as a greener analytical method.     

Quantification of coumarin derivatives in Noni (Morinda citrifolia) and their contribution of quenching effect on reactive oxygen species

The quantification of coumarin derivatives such as scopoletin, 7-hydroxycoumarin (7-HC) and 4-hydroxycoumarin (4-HC) in Noni (Morinda citrifolia) was described. The coumarin derivatives were determined by HPLC-UV or -fluorescence detection. More than 95% of peak purity for coumarin derivatives in Noni sample was confirmed by a multi-wavelength fluorescence detector. Amounts of scopoletin and 7-HC in Noni juices (A–H) were ranging 5.1–231 μg/ml and 0.04–0.45 μg/ml, respectively (n = 12). No 4-HC was detected in any Noni samples examined.     

Total phenolics,ascorbic acid,and antioxidant capacity of noni (Morinda citrifolia L.) juice and powder as affected by illumination during storage

Total phenolics, ascorbic acid, and antioxidant capacity of noni (Morinda citrifolia L.) juice and powder were determined during storage at 24 °C. After 2 weeks of storage, illuminated noni juice lost 32% of total phenolics, 89% of ascorbic acid, and 46–65% of antioxidant capacity—about 8%, 22%, and 9–15% more than unilluminated juice. Both illuminated and unilluminated juice lost 97% of ascorbic acid by 4 weeks. The difference in antioxidant characteristics between illuminated and unilluminated juice became insignificant at 12 weeks. After 12 weeks, illuminated noni powder lost 21% of total phenolics, 17% of ascorbic acid, and 23–36% of antioxidant capacity—about 13%, 4%, and 7–19% more than the unilluminated powder. Noni powder in brown bottles retained antioxidant characteristics significantly greater than that in clear bottles. Protection from light effectively reduced degradation of antioxidant characteristics of noni juice for only 2 weeks but those of noni powder for at least 3 months.     

Development and validation of an RP-HPLC method for the analysis of anthraquinones in noni fruits and leaves

Noni fruits and leaves, which have been used traditionally for thousands of years to improve health, are increasingly attracting the interests of consumers and researchers. A selective and validated HPLC method for the analysis of anthraquinones in noni fruits and leaves has been developed and is reported for the first time. Four anthraquinones, 5,15-dimethylmorindol (5,15-DMM, 1), lucidin (2), and alizarin (3), and rubiadin (4) are examined. The limits of detection of 1–4 were in the range of 1.0 and 20.0 ng. Intra- and inter-day precisions of 1 were determined to be less than 5.3%. The accuracy, expressed as the percent recovery of 1 after spiking at three concentrations ranged from 83.0% to 93.3%. Further, the linear correlation coefficient was >0.999, within the range of concentration investigated. The 5,15-DMM content of noni fruit puree and noni leaf infusion are between 0.186 to 0.202 μg/mL (ppm), and 5.82 to 20.93 ng/mL (ppb), respectively. Lucidin, rubiadin and alizarin were not detected in any of the noni samples. The presence of only trace amounts in the noni fruits and leaves may help to eliminate safety concerns regarding anthraquinone contents.     

Inhibitory effect of anthraquinones isolated from the Noni (Morinda citrifolia) root on animal A-, B- and Y-families of DNA polymerases and human cancer cell proliferation

During the course of our studies to develop new uses for the Noni (Morinda citrifolia) root, 10 anthraquinones, rubiadin (1), rubiadin 1-methyl ether (2), lucidin (3), damnacanthol (4), 1,3-dihydroxy-2-methoxymethylanthraquinone (5), 3-hydroxy-1-methoxy-2-methoxymethylanthraquinone (6), nordamnacanthal (7), damnacanthal (8), sorandidiol (9) and morindone (10), were isolated. Compounds 5, 6, 7, 8 and 10 exhibited remarkable inhibition against the activities of animal pols, and compound 10 was the strongest inhibitor in the anthraquinones investigated. Among mammalian pols, compound 10 inhibited the pol activities of A- (pol γ), B- (pols α, δ and ε) and Y- (pols η, ι and κ) families, but did not influence the activities of X-family pols (pols β, λ and terminal deoxynucleotidyl transferase). The tendency of pol inhibition showed a positive correlation with the suppression of human colon cancer cell HCT116 growth. These results suggested that the Noni root containing anthraquinones may be used as an anticancer functional food.     

The in vitro and in vivo effects of yerba mate (Ilex paraguariensis) extract on adipogenesis

The aim of this study was to evaluate the effects of yerba mate extract and its principal bioactive compounds on adipogenesis. The anti-adipogenic effects of yerba mate, chlorogenic acid, quercetin and rutin were evaluated in 3T3-L1 cells using a PCR array. The results obtained in vitro were validated in vivo in a high-fat diet-induced model of obesity. The in vitro and in vivo results demonstrated that yerba mate extract down-regulated the expression of genes that regulate adipogenesis, such as Creb-1and C/EBPα, and the extract up-regulated the expression of genes related to the inhibition of adipogenesis, including Dlk1, Gata2, Gata3, Klf2, Lrp5, Pparγ₂, Sfrp1, Tcf7l2, Wnt10b, and Wnt3a. In summary, it was demonstrated that yerba mate and its bioactive compounds regulate the expression of genes related to in vitro adipogenesis. Furthermore, yerba mate might regulate adipogenesis through the Wnt pathway.     

The in vitro and in vivo antioxidant properties of seabuckthorn (Hippophae rhamnoides L.) seed oil

The antioxidant capacity of seabuckthorn (Hippophae rhamnoides L.) seed oil was investigated with a number of established in vitro assays and in an in vivo study of carbon tetrachloride (CCl₄)-induced oxidative stress in mice. The results showed that DPPH radical scavenging activity, ferrous ion chelating activity, reducing power and inhibition of lipid peroxidation activity all increased with increasing concentrations of seabuckthorn seed oil. Moreover, the EC₅₀ values of seabuckthorn seed oil from the hydrogen peroxide, superoxide radical, hydroxyl radical scavenging assays were 2.63, 2.16 and 0.77 mg/ml, respectively. In the in vivo study, seabuckthorn seed oil inhibited the toxicity of CCl₄, as seen from the significantly increased activities of the antioxidant enzymes superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase. The GSH content in the liver was also increased, whereas hepatic malondialdehyde was reduced. Taken together, these results clearly indicate that seabuckthorn seed oil has significant potential as a natural antioxidant agent.     

Isolation,purification and characterisation of lunasin from defatted soybean flour and in vitro evaluation of its anti-inflammatory activity

Lunasin is a chemopreventive peptide present in soybean and other plant sources. The high cost involved in obtaining synthetic lunasin limits its application in chemopreventive and nutritional interventions. The objective of this study was to isolate, purify and characterise lunasin from defatted soybean flour and determine its in vitro anti-inflammatory activity using RAW 264.7 macrophages. Isolation and purification was achieved by ion-exchange chromatography, ultrafiltration and size exclusion chromatography. The identity of lunasin was established by Western blot, HPLC, MALDI-TOF and LC/MS-MS. The results showed that lunasin eluted from a DEAE anion exchange column at 0.15 M NaCl, and after 1.5 void volumes from size exclusion chromatography. Fractions from both chromatographic techniques consistently showed three peptides with positive immunoreactivity against lunasin mouse monoclonal antibody corresponding to 5, 8 and 14 kDa. LC/MS-MS analysis of the three immunoreactive peptides showed that 5 and 14 kDa bands contained the lunasin epitope, RGDDDDDD DDD while 8 kDa band showed high homology with 2 S soy albumin, a lunasin precursor.     

Effects of simulated digestion in vitro on cell wall polysaccharides from kiwifruit (Actinidia spp.)

Cell wall polysaccharides are resistant to digestion and absorption in the human small intestine and are considered to be delivered to the colon in a chemically unaltered state. In this paper, pulp from green and gold kiwifruit was subjected to in vitro upper-intestinal tract digestion and the chemical and physical changes to cell wall polysaccharides (dietary fibre) were investigated. Yields of insoluble fibre decreased slightly with simulated digestion while soluble fibre yields increased. Constituent sugar and glycosyl linkage analysis of the soluble and insoluble fibre fractions revealed that the chemical composition and structure of the non-starch polysaccharides remained largely unchanged. However, the degree of methylesterification of galacturonic acid residues present in the pectin-rich soluble fibre fractions of both fruit decreased with treatment; size-exclusion chromatography detected changes in the molecular weight profiles of these fractions. These changes may affect the physicochemical properties and fermentability of kiwifruit dietary fibre in the large intestine.     

Effects of binary solvent extraction system,extraction time and extraction temperature on phenolic antioxidants and antioxidant capacity from mengkudu (Morinda citrifolia)

An investigation into the effects of ethanol concentration (0–100%, v/v), extraction time (20–120 min) and extraction temperature (25–65 °C) on the extraction of phenolic antioxidants from mengkudu (Morinda citrifolia) was performed using a single-factor experiment. Total phenolic content (TPC) and total flavonoid content (TFC) assays were used for determination of phenolic compounds. Antioxidant capacity was evaluated by measuring the scavenging effect on 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 2,2′-diphenyl-1-picrylhydrazyl (DPPH) radicals. Experimental results showed that extraction conditions had significant effect on extraction of phenolic compounds and antioxidant capacities. The optimised conditions were 40% ethanol for 80 min at 65 °C, with values of 919.95 mg GAE/100 g DW for TPC, 472.73 mg CE/100 g DW for TFC, 791.71 μmol TEAC/100 g DW for ABTS and 1928.5 μmol TEAC/100 g DW for DPPH. TPC was significantly correlated with DPPH under the effects of ethanol concentration (r = 0.932) and extraction time (r = −0.938).     

The effects of Caulerpa microphysa enzyme-digested extracts on ACE-inhibitory activity and in vitro anti-tumour properties

The aim of this study was to determine the ACE inhibitory activity and its anti-cancer properties of Caulerpa microphysa extracts. C. microphysa samples were digested with Flavourzyme, Alcalase, and pepsin. The ACE inhibitory activity of enzyme-digested C. microphysa decreased in the order of digestion with pepsin > Flavourzyme > Alcalase; that is, pepsin-extracted samples had significantly higher activity than the other enzyme extractions. To test its anti-tumour effects in vitro C. microphysa pepsin-digested extracts were applied to BALB/c mice with transplanted myelomonocytic leukaemia (WEHI-3) and Human promyelocytic leukaemia (HL-60) cell lines. The growth of both cell lines was inhibited, and extracts induced DNA damage, evaluated with a comet assay. The data demonstrate that C. microphysa pepsin-digested extract had the ability to anti-tumour effects. Further application as a health food is worthy of investigation.     

Isolation and structure elucidation of phenolic compounds from Cyclopia subternata Vogel (honeybush) intact plant and in vitro cultures

In the presented work, an insight was made into the polyphenolic composition of intact plant material and in vitro cultures of indigenous South African plant Cyclopia subternata Vogel (honeybush). Ethyl acetate fractions of methanol extracts were separated by means of gravity column chromatography and/or semipreparative HPLC on two serially connected monolithic RP-18 columns. The structures of the isolated compounds were determined by means of 1D and 2D NMR techniques and additionally confirmed by LC-DAD-ESI-MS. Apart from the previously described honeybush components, that is mangiferin (1), scolymoside (2), hesperidin (3) and narirutin (4), three additional compounds: iriflophenone 3-C-β-glucoside (benzophenone) (5), phloretin 3′,5′-di-C-β-glucoside (dihydrochalcone) (6), and isorhoifolin (flavone) (7) were identified for the first time in the herb of C. subternata. Additionally, three isoflavone glucosides, namely calycosin 7-O-β-glucoside (8), rothindin (9) and ononin (10), which had not been previously reported in Cyclopia plants, were identified in the callus of the above species. As far as the authors are concerned, this is the first report on the presence of benzophenone and dihydrochalcone derivatives in Cyclopia genus.     

In vitro and in vivo protein hydrolysis of beans (Phaseolus vulgaris) genetically modified to express different phaseolin types

Experiments were conducted to study whether phaseolin type could influence proteolysis susceptibility and nutritional value of total bean protein. The DOR-390 bean cultivar was genetically modified to express different phaseolin types (S, T or I). Beans were soaked and autoclaved. A sequential hydrolysis was carried out in vitro with pepsin and pancreatin. Differences in the degree of protein hydrolysis among bean lines started at 30 min and remained until 240 min, with the S bean proteins presenting lower values (P < 0.05). Subsequently, rats were fed with diets containing beans expressing different phaseolin types as the only source of protein for N digestibility and nutritional value determination. No differences (P > 0.05) in ileal protein digestibility and rat growth were observed. In conclusion, the differences in in vitro hydrolysis between bean lines expressing different phaseolin types had no consequences on growth and N retention in rats.     

Nutritional and nutraceutical comparison of Jamaican Psidium cattleianum (strawberry guava) and Psidium guajava (common guava) fruits

Psidium cattleianum (strawberry guava) is one of many underutilised edible fruits that grow wild in Jamaica, and could potentially be commercially exploited to yield health and economic benefits. In this study, the total phenolics, proximate contents, and antioxidant, anti-inflammatory, and antimicrobial activities of P. cattleianum and P. guajava (common guava), a well-known species, were compared. Strawberry guavas were found to be superior to common guavas in antioxidant and antimicrobial activities, total phenolics and vitamin C content. They also possessed relatively high fibre content (24.9%). The hexane and ethyl acetate extracts of strawberry guavas showed cyclooxygenase-2 enzyme inhibitory activities of 18.3% and 26.5%, respectively (250 μg/mL), indicating anti-inflammatory activity. The EtOAc and MeOH extracts of P. guajava showed 56.4% (COX-2) and 44.1% (COX-1) inhibitory activity, respectively. Additionally, nine compounds were isolated from strawberry guava fruits, some of which demonstrated anti-inflammatory activity. These results indicate that strawberry guavas are beneficial for health.     

Inhibition of aldose reductase and anti-cataract action of trans-anethole isolated from Foeniculum vulgare Mill. fruits

Foeniculum vulgare fruits are routinely consumed for their carminative and mouth freshening effect. The plant was evaluated for aldose reductase inhibition and anti-diabetic action. Bioguided fractionation using silica gel column chromatography, HPLC, and GC-MS analysis revealed trans-anethole as the bioactive constituent possessing potent aldose reductase inhibitory action, with an IC₅₀ value of 3.8 μg/ml. Prolonged treatment with the pet ether fraction of the F. vulgare distillate demonstrated improvement in blood glucose, lipid profile, glycated haemoglobin and other parameters in streptozotocin-induced diabetic rats. Trans-anethole could effectively show anti-cataract activity through the increase in soluble lens protein, reduced glutathione, catalase and SOD activity on in vitro incubation of the eye lens with 55 mM glucose. Trans-anethole demonstrated noncompetitive to mixed type of inhibition of lens aldose reductase using Lineweaver Burk plot.     

Neuronal cell protective and antioxidant effects of phenolics obtained from Zanthoxylum piperitum leaf using in vitro model system

In order to obtain basic data for the utilisation of Zanthoxylum piperitum leaf as a functional substance in the food industry, the antioxidative and neuronal cell protective effects of silica-gel open column chromatographic fractions, obtained from Z. piperitum leaf, were examined. ABTS and FRAP assays indicated that the ZP4 fraction (eluted with methanol/chloroform, 1:4, v/v) of Z. piperitum leaf was a more potent radical-scavenger and reducing agent than the other five fractions. The ZP4 fraction also presented protective effects against H₂O₂-induced neurotoxicity in a dose-dependent manner. Therefore, our study verified that the ZP4 fraction has strong antioxidative and neuronal protective effects which are correlated with its high level of phenolics, particularly quercitrin, afzelin, and hyperoside. These phenolics of Z. piperitum leaf can be utilised as effective and safe functional food substances, i.e., natural antioxidants, and may reduce the risk of neurodegenerative disorders.     

Preparation,identification and their antitumor activities in vitro of polysaccharides from Chlorella pyrenoidosa

Ultrafiltration membranes of different pore size were applied to fractionate Chlorella pyrenoidosa polysaccharides (CPPS) and the main fraction could be separated by a membrane with nominal molecular weight cut-off (NMWCO) of 30 kDa. Ultrafiltration parameters of 40 °C 14.0 psi were optimized for obtaining the main fraction. The resulting sample was further purified by anion-exchange chromatography and size exclusion chromatography, and two distinctive polysaccharides, CPPS Ia and IIa were recovered. CPPS IIa had infrared spectral characteristic of polysaccharides similar to CPPS Ia, and the symmetrical stretching peak at 1408–1382 cm⁻¹ was an indication of the presence of carboxyl groups. The peak molecular weights were 69658 Da and 109406 Da, for CPPS Ia and CPPS IIa, respectively. Both CPPS Ia and IIa were composed of rhamnose, mannose, glucose, galactose and an unknown monosaccharide. Galactose (relative mass 46.5%) was the predominant monosaccharide of CPPS Ia and in CPPS IIa, rhamnose (37.8%) was predominant. CPPS Ia and IIa presented significantly higher antitumor activity against A549 in vitro than did a blank control, in a dose-dependent manner. Both fractions might be useful for developing natural safe antitumor drugs from C. pyrenoidosa resources.