Effect of production system and inhibitory potential of aroma volatiles on polyphenol oxidase and peroxidase activity in tomatoes.

BACKGROUND: Polyphenol oxidase (PPO) and peroxidase (POD) are key enzymes associated with shelf life and defense responses. Thus, the activity of PPO and POD enzymes is usually assessed to check the quality of food samples and to understand the physiological responses of plants to different stresses. However, the outcomes of PPO and POD activity assessment studies are highly dependent on assay conditions. Hence, in this study, we initially optimized PPO and POD extraction and high-throughput 96-well plates-based enzymatic activity assessment methods to evaluate the inhibitory potential of tomato volatile compounds. Later, we explored the effects of net-house and open-field growing on the PPO and POD activity in tomato fruits of eight cultivars. RESULTS: We found 150 mM of catechol and pH 7.0 were the optimal conditions for the maximum activity for the PPO assay. Conversely, 24 mM guaiacol with 12 mM H2 O2 and pH 6.0 was the best condition for the POD assay. Thermal inactivation studies confirmed that tomato POD is more resistant to heat than PPO. We found that the production systems had a considerable genotype-specific impact on tomato PPO and POD activity. Moreover, amongst the volatiles that were studied, ß-damascenone and d-limonene showed 50% PPO inhibition at 40 and 80 mM, respectively. CONCLUSION: The results of this study can be used to improve the shelf-life of fresh tomato fruit and its products. The findings also underscore the significance of PPO and POD enzymes as physiological trait markers in the tomato crop and fruit quality improvement programs. © 2020 Society of Chemical Industry.

Improved Sample Preparation and Optimized Solvent Extraction for Quantitation of Carotenoids.

Accurate, rapid quantitation of key antioxidants such as carotenoids is important for assessment of food quality. Carotenoids are lipid-soluble pigments that are susceptible to oxidation due to their highly conjugated carbon-carbon double bonds. Therefore, the present work focuses on improving sample preparation to facilitate rapid analysis of carotenoids. The method involves optimized carotenoid extraction followed by direct HPLC analysis without further concentration and redissolution. For extraction, we tested the effect of blending time (1, 3 and 5 min) and 12 different solvent combinations for carotenoid extraction from cantaloupe (Cucumis melo var. cantalupensis) and oranges (Citrus sinensis), two popular fruits that are high in carotenoids. The identification of carotenoids was performed by LC-APCI-QTOF-HR-MS in positive-ionization mode. In melon, 1 min blending time gave significantly higher ß-carotene content with CHCl3: Ace (1:1) solvent. The optimized method was validated with tomato, watermelon, oranges, grapefruit, melon varieties and commercial products such as fruit juices. Among the different melon varieties, Western Shipper had significantly higher ß-carotene (25.1 ± 0.4 µg/g) contents. In oranges, ß-carotene and (all-E)-lycopene contents were 4.4 ± 0.1and 3.8 ± 0.1 µg/g, respectively. The optimized method has fewer unit operations and is reproducible for the quantitation of carotenoids and their isomers. This is the first report on the identification of ζ-carotene isomers, and lycopene isomers from cantaloupe varieties and lycopene from oranges. Graphical Abstract.

CIRCADIAN CLOCK-ASSOCIATED1 Controls Resistance to Aphids by Altering Indole Glucosinolate Production.

CIRCADIAN CLOCK-ASSOCIATED1 (CCA1), a well-known central circadian clock regulator, coordinates plant responses to environmental challenges. Its daily rhythmic expression in Arabidopsis (Arabidopsis thaliana) confers host resistance to the caterpillar Trichoplusia ni However, it is unclear whether CCA1 plays a role in defense against phloem sap-feeding aphids. In this study, we showed that green peach aphid (Myzus persicae) displayed an intrinsic circadian feeding rhythm. Under constant light, wild-type Columbia-0 (Col-0) Arabidopsis plants coentrained with aphids in the same light/dark cycles exhibited greater antixenotic activity than plants preentrained in the opposite cycle from the aphids. Consistently, circadian mutants cca1-1, cca1-11, lhy-21, ztl-1, ztl-4, and lux-2 suffered more severe damage than Col-0 plants when infested by aphids, suggesting that the Arabidopsis circadian clock plays a defensive role. However, the arrhythmic CCA1 overexpression line (CCA1-OX) displayed strong antixenotic and antibiotic activities despite its loss of circadian regulation. Aphids feeding on CCA1-OX plants exhibited lower reproduction and smaller body size and weight than those on Col-0. Apparently, CCA1 regulates both clock-dependent and -independent defense responses. Systematic investigation based on bioinformatics analyses indicated that resistance to aphids in CCA1-OX plants was due primarily to heightened basal indole glucosinolate levels. Interestingly, aphid feeding induced alternatively spliced intron-retaining CCA1a/b transcripts, which are normally expressed at low levels, whereas expression of the major fully spliced CCA1 transcript remained largely unchanged. We hypothesize that posttranscriptional modulation of CCA1 expression upon aphid infestation maximizes the potential of circadian-mediated defense and stress tolerance while ensuring normal plant development.

Evaluation of bitter melon (Momordica charantia) cultivars grown in Texas and levels of various phytonutrients.

BACKGROUND: In the USA, Momordica charantia is relatively unknown and is usually found in specialty markets. In the present study, cultivation of five bitter melon cultivars grown under field conditions in College Station (TX, USA), was evaluated. Additionally, ascorbic acid, amino acids and phenolic compounds were quantified from various cultivars grown in different years. RESULTS: The yield of the first year of evaluation was comparable to other bitter melon growing regions, ranging from 9371.5 kg ha-1 for the Japanese Spindle cultivar to 20 839.1 kg ha-1 for the Hong Kong Green cultivar. Multivariate analysis suggests a strong correlation between yield and growth degree days, water use efficiency and organic matter, as well as an inverse correlation with the amount or precipitation during the growing season. The highest levels of total ascorbic acid were shown in the Japanese Spindle cultivar (162.97 mg 100 g-1 fresh fruit), whereas the lowest levels were expressed in the Hong Kong Green cultivar (42.69 mg 100 g-1 fresh fruit). The highest levels of total phenolics were consistently found the Indian White cultivar, in the range 10.6-12.5 mg g-1 catechin equivalents. Seven phenolics and organic acids were identified and quantified by liquid chromatography-mass spectrometry and high-performance liquid chromatography, respectively. Additionally, the highest levels of total amino acids were found in the Large Top cultivar. CONCLUSION: The current 3-year field study demonstrates that it is feasible to grow bitter melon commercially in Texas with proper climatic and agronomic conditions. Bitter melon is a rich source for ascorbic acid, amino acids and phenolic compounds, which makes it a valuable food source with respect to improving human health. © 2018 Society of Chemical Industry.

Effect of harvest time on the levels of phytochemicals, free radical-scavenging activity, α-amylase inhibition and bile acid-binding capacity of spinach (Spinacia oleracea).

BACKGROUND: Spinach is a green leafy vegetable that is rich in health-promoting compounds. The present study analyzed the levels of phytochemicals and health-promoting properties of spinach harvested at 20, 30, 40, 50 and 60 days after planting. RESULTS: The time of harvest had a significant effect on nitrate levels, which increased from 1909 ± 70.6 µg g-1 (20 days) to 3668 ± 101.3 µg g-1 (40 days) and then decreased to 974 ± 164 µg g-1 (60 days). Lutein and chlorophylls a and b were found to be maximum at 60 days, whereas ß-carotene was higher at 50 days. Liquid chromatography/high-resolution quadrupole time-of-flight tandem mass spectrometry (LC/HR-QTOF-MS) was used to identify 12 flavonoids, and their tentative fragmentation pathways have been proposed. Spinach harvested at 30 and 60 days exhibited significantly higher 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical-scavenging activities and inhibition of amylase. The levels of total phenolics ranged from 885 ± 35.1 to 1162 ± 112.4 µg g-1 in the samples. In vitro bile acid-binding capacity showed that glycochenodeoxycholate and glycodeoxycholate were bound to maximum levels in all spinach samples. CONCLUSION: The harvest time has a major effect on the levels of phytochemicals and health-beneficial properties, which indicates that consumption of both baby and mature spinach will provide maximum health benefits. © 2017 Society of Chemical Industry.

Homeostatic responses of colonic LGR5+ stem cells following acute in vivo exposure to a genotoxic carcinogen.

Perturbations in DNA damage, DNA repair, apoptosis and cell proliferation in the base of the crypt where stem cells reside are associated with colorectal cancer (CRC) initiation and progression. Although the transformation of leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5)(+) cells is an extremely efficient route towards initiating small intestinal adenomas, the role of Lgr5(+) cells in CRC pathogenesis has not been well investigated. Therefore, we further characterized the properties of colonic Lgr5(+) cells compared to differentiated cells in Lgr5-EGFP-IRES-creER(T2) knock-in mice at the initiation stage of carcinogen azoxymethane (AOM)-induced tumorigenesis using a quantitative immunofluorescence microscopy approach. At 12 and 24h post-AOM treatment, colonic Lgr5(+) stem cells (GFP(high)) were preferentially damaged by carcinogen, exhibiting a 4.7-fold induction of apoptosis compared to differentiated (GFP(neg)) cells. Furthermore, with respect to DNA repair, O(6)-methylguanine DNA methyltransferase (MGMT) expression was preferentially induced (by 18.5-fold) in GFP(high) cells at 24h post-AOM treatment compared to GFP(neg) differentiated cells. This corresponded with a 4.3-fold increase in cell proliferation in GFP(high) cells. These data suggest that Lgr5(+) stem cells uniquely respond to alkylation-induced DNA damage by upregulating DNA damage repair, apoptosis and cell proliferation compared to differentiated cells in order to maintain genomic integrity. These findings highlight the mechanisms by which colonic Lgr5(+) stem cells respond to cancer-causing environmental factors.

5-Geranyloxy-7-methoxycoumarin inhibits colon cancer (SW480) cells growth by inducing apoptosis.

For the first time, three coumarins were isolated from the hexane extract of limes (Citrus aurantifolia) and purified by flash chromatography. The structures were identified by NMR (1D, 2D) and mass spectral analyses as 5-geranyloxy-7-methoxycoumarin, limettin, and isopimpinellin. These compounds inhibited human colon cancer (SW-480) cell proliferation, with 5-geranyloxy-7-methoxycoumarin showing the highest inhibition activity (67 %) at 25 µM. Suppression of SW480 cell proliferation by 5-geranyloxy-7-methoxycoumarin was associated with induction of apoptosis, as evidenced by annexin V staining and DNA fragmentation. In addition, 5-geranyloxy-7-methoxycoumarin arrested cells at the G0/G1 phase, and induction of apoptosis was demonstrated through the activation of tumour suppressor gene p53, caspase8/3, regulation of Bcl2, and inhibition of p38 MAPK phosphorylation. These findings suggest that 5-geranyloxy-7-methoxycoumarin has potential as a cancer preventive agent.

Obacunone represses Salmonella pathogenicity islands 1 and 2 in an envZ-dependent fashion.

Obacunone belongs to a class of unique triterpenoids called limonoids, present in Citrus species. Previous studies from our laboratory suggested that obacunone possesses antivirulence activity and demonstrates inhibition of cell-cell signaling in Vibrio harveyi and Escherichia coli O157:H7. The present work sought to determine the effect of obacunone on the food-borne pathogen Salmonella enterica serovar Typhimurium LT2 by using a cDNA microarray. Transcriptomic studies indicated that obacunone represses Salmonella pathogenicity island 1 (SPI1), the maltose transporter, and the hydrogenase operon. Furthermore, phenotypic data for the Caco-2 infection assay and maltose utilization were in agreement with microarray data suggesting repression of SPI1 and maltose transport. Further studies demonstrated that repression of SPI1 was plausibly mediated through hilA. Additionally, obacunone seems to repress SPI2 under SPI2-inducing conditions as well as in Caco-2 infection models. Furthermore, obacunone seems to repress hilA in an EnvZ-dependent fashion. Altogether, the results of the study seems to suggest that obacunone exerts an antivirulence effect on S. Typhimurium and may serve as a lead compound for development of antivirulence strategies for S. Typhimurium.

Inhibition of prostate cancer (LNCaP) cell proliferation by volatile components from Nagami kumquats.

Fresh Nagami kumquats (Fortunella margarita) were subjected to hydrodistillation using a Clevenger-type apparatus to obtain volatile oil. The chemical composition of the volatile oil was analyzed by GC-MS using Rtx-5 Sil MS and DB Wax columns. A total of 25 volatile compounds were identified by mass spectra, retention index, and comparison with known standards. The major identified compounds are d-limonene (41.64 %), ß-myrecene (16.54 %), linalyl propionate (9.55 %), and germacrene-D (5.93 %) from the Rtx-5 Sil MS column; d-limonene and ß-myrecene were also separated as major compounds on the DB wax column. The oil is rich in hydrocarbons (77.41 %) consisting of 60.05 % monoterpenes and 17.36 % sesquiterpenes. Interestingly, oxygenated hydrocarbons (17.6 %) were also found in kumquat volatile oil. Certain volatile compounds were also confirmed by positive chemical ionization and NMR spectra. Further, the volatile oil demonstrated good DPPH radical scavenging activity and antioxidant capacity. Kumquat volatile oil at 200 ppm concentration exhibited 55 %, 61 %, and 63.4 % inhibition of human prostate cancer (LNCaP) cell proliferation at 24, 48, and 72 h, respectively, by cell count assays. Significant increases in expression of bax/bcl2 and p53 proteins confirmed that volatile oil induces apoptosis. In addition, inhibition of inflammatory markers such as NF-κB and Cox-2 was observed. The cleavage of caspase-8 in the LNCaP cells treated with volatile oil demonstrated that apoptosis occurred through an extrinsic pathway. This is the first report of the identification and possible mechanisms of in vitro antiproliferative effects of kumquat volatile components on human prostate cancer (LNCaP) cells.

Simultaneous separation and identification of limonoids from citrus using liquid chromatography-collision-induced dissociation mass spectra.

Limonoids are considered as potential cancer chemopreventive agents and are widely distributed in the Citrus genus as aglycones and glucosides. In the present study, reversed-phase HPLC coupled with CID mass spectra was developed for the simultaneous separation and identification of aglycones and glucosides of limonoids from citrus. Five aglycones such as limonin, deacetyl nomilin, ichangin, isolimonoic acid and nomilin were identified by positive ion CID MS/MS, whereas five glucosides, viz. limonin glucoside, isoobacunoic acid glucoside, obacunone glucoside, deacetyl nomilinic acid glucoside and nomilinic acid glucoside were analyzed by negative ion CID mass spectra. The developed method was successfully applied to complex citrus samples for the separation and identification of aglycones and glucosides. Citrus seeds were extracted with methanol and partially purified and analyzed by LC-CID mass spectra. The separation was achieved by C-18 column; eight limonoids were identified by comparing the retention times and mass spectral fragmentation. To the best of our knowledge, this is the first report on the identification of citrus limonoids using CID technique.

Production system influences tomato phenolics and indoleamines in a cultivar-specific manner.

Tomato (Solanum lycopersicum) fruit is a rich source of health-promoting compounds, and epidemiological studies show that tomato consumption may reduce the risk of chronic diseases. This study compared the effect of genotype, production system, and their interaction on eight tomato varieties grown in the open-field (OF) or net-house (NH), a structure completely covered with a 50-mesh screen to reduce pest and wind damage, in South Texas. The NH structure reduced solar radiation up to ~30% and decreased wind speed by 6.44 km/h compared with conditions measured in the OF. We simultaneously analyzed 16 phenolics and indoleamines using ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight high-resolution mass spectrometry (UHPLC/ESI-HR-QTOFMS). The chemometric analysis showed a distinct difference between NH- and OF-grown tomatoes irrespective of the variety. The melatonin and serotonin contents showed a cultivar-specific effect of the production system. Likewise, the effect of cultivation systems on levels of phenolic acids and flavonoids varied based on tomato cultivar. Among the studied phenolic acids, significantly enhanced levels of sinapic acid were observed in OF-grown tomatoes. Similarly, we detected a considerable genotypic effect on gallic acid, p-coumaric acid, ferulic acid, sinapic acid, and naringin. The interaction of cultivar and production system substantially affected gallic acid, protocatechuic acid, sinapic acid, and apigenin. However, further studies need to be performed to explore the environment-specific effects on the total composition. In summary, our results indicate that the production system plays an important role in tomato composition beyond the natural genetic variation among cultivars.

1H Nuclear Magnetic Resonance and Liquid Chromatography Coupled with Mass Spectrometry-Based Metabolomics Reveal Enhancement of Growth-Promoting Metabolites in Onion Seedlings Treated with Green-Synthesized Nanomaterials.

Seed priming is a promising approach to improve germination, emergence, and seedling growth by triggering pre-germinative metabolism and enhancing seedling vigor. Recently, nanopriming gained importance in seed improvement as a result of the small size and unique physicochemical characteristics of nanomaterials. In the present study, silver and gold nanoparticles were synthesized using onion extracts as the reducing agent. Similarly, the agro-food industrial byproducts citrus seed oil and curcumin-removed turmeric oleoresin were used for the preparation of nanoemulsions. For seed priming, these green-synthesized nanomaterials were incubated with seeds of two onion (Allium cepa L.) cultivars (Legend and 50147) for 72 h, and then the plants were grown in a greenhouse for 3 weeks. Seed priming with these nanomaterials increased seed germination and seedling emergence. One-dimensional 1H nuclear magnetic resonance and liquid chromatography coupled with mass spectrometry metabolomics studies showed that different nanopriming treatments distinctly altered the metabolome of onion seedlings. Seed priming treatments significantly inhibited plant hormones and growth regulators, such as abscisic acid and cis-(+)-12-oxo-phytodienoic acid, and enhanced germination stimulators, such as γ-aminobutyric acid and zeatin, in onion seeds and seedlings. Therefore, these priming treatments have positive impact on improving seed performance and plant growth.

Multivariate Analysis of Amino Acids and Health Beneficial Properties of Cantaloupe Varieties Grown in Six Locations in the United States.

Cantaloupe is a good dietary source of amino acids, including γ-aminobutyric acid (GABA), glutamine, and citrulline. However, the levels of these amino acids vary among different cantaloupe varieties grown in different locations. Understanding the variation in amino acid contents provides fundamentally important information for quality control and improving melon varieties. To examine this variation, we measured the amino acid contents in cantaloupes grown in six locations in the United States (Texas, Georgia, North Carolina, California, Indiana, and Arizona). Principal component analyses were applied to analyze the effect of growing location on the amino acid profiles in different varieties. The GABA content ranged from 1006.14 ± 64.77 to 3187.12 ± 64.96 µg/g and citrulline ranged from 92.65 ± 9.52 to 464.75 ± 34.97 µg/g depending on the variety and location. Total phenolic contents, α-amylase inhibition, and antioxidant activities were also measured. Tuscan type Da Vinci had significantly higher phenolic contents in Arizona (381.99 ± 16.21 µg/g) but had the lowest level when grown in California (224.56 ± 14.62 µg/g). Our analyses showed significant differences in amino acid levels, phenolics contents, and antioxidant activity in the cantaloupe varieties based on the growing location. These findings underline the importance of considering growing location in the selection and improvement of cantaloupe varieties.

Effects of genotype and production system on quality of tomato fruits and in vitro bile acids binding capacity.

Tomato is an important source of health-promoting constituents, and researchers have focused on enhancing the content. In the present study, the influence of net-house (NH) and open-field (OF) growing conditions on physicochemical traits of tomatoes from eight different cultivars were evaluated. The tomato fruit qualities, such as color, total soluble solids (TSS), total acidity (TA), and pH were measured. Furthermore, ultra-performance liquid chromatography coupled to electrospray ionization high-resolution quadrupole time-of-flight mass spectrometry (UPLC/ESI-HR-QTOFMS) was used for identification and quantification of health-promoting compounds such as ascorbic acid, ß-carotene, lycopene, and its isomers. In addition, in vitro bile acid binding capacity of all tomato samples was analyzed along with soluble and insoluble dietary fiber analysis as biofunctional properties. The results suggest that production systems influenced tomato fruit quality and biofunctional characteristics in a variety-specific manner. Notably, TA and all-trans-ß-carotene values were considerably influenced by production systems, and their levels were higher in the NH- and OF-grown tomatoes, respectively. Our findings underline the importance of the rational choice of genotype and production system to obtain high-quality tomatoes with enhanced desired traits for breeders and consumers. PRACTICAL APPLICATION: Tomato is one of the nutritional high-valued horticultural crops. The present study aimed to assess the impact of production systems, such as net-house and open-field conditions, on physicochemical traits and biofunctional properties, in vitro bile acid binding capacity of eight tomato varieties. This study supplies a good reference for the rational selection of genotype and production system to obtain high-quality tomatoes with improved desired traits for breeders and consumers.

Dietary curcumin and limonin suppress CD4+ T-cell proliferation and interleukin-2 production in mice.

Phytochemicals may reduce chronic inflammation and cancer risk in part by modulating T-cell nuclear factor-kappaB (NF-kappaB) activation. Therefore, we examined the effects of curcumin (Cur) and limonin (Lim) feeding on NF-kappaB-dependent CD4(+) T-cell proliferation. DO11.10 transgenic mice (n = 5-7) were fed diets containing 1% Cur or 0.02% Lim combined with either (n-6) PUFA [5% corn oil (CO)] or (n-3) PUFA [4% fish oil+1% corn oil (FO)] for 2 wk, followed by splenic CD4(+) T-cell isolation and stimulation with ovalbumin peptide 323-339 (OVA) and antigen-presenting cells from mice fed a conventional nonpurified rodent diet. Both Cur and Lim diets suppressed (P < 0.05) NF-kappaB p65 nuclear translocation in activated CD4(+) T-cells. In contrast, activator protein-1 (c-Jun) and nuclear factor of activated T-cells c1 were not affected compared with the CO control diet (no Cur or Lim). CD4(+) T-cell proliferation in response to either mitogenic anti-CD3/28 monoclonal antibodies (mAb) or antigenic stimulation by OVA was also suppressed (P < 0.05) by Cur as assessed by carboxyfluorescein succinimidyl ester staining. In contrast, interleukin-2 production was not directly associated with NF-kappaB status. Interestingly, dietary combination with FO enhanced the suppressive effects (P < 0.05) of Cur or Lim with respect to CD4(+) T-cell proliferation in response to anti-CD3/28 mAb. These results suggest that combination chemotherapy (FO+Cur or Lim) may favorably modulate CD4(+) T-cell-mediated inflammation.

Leaf Disc Assays for Rapid Measurement of Antioxidant Activity.

Antioxidant levels are key parameters for studies of food quality, stress responses, and plant health. Herein, we have demonstrated that excised leaf disc has both radical scavenging activity and reducing power, and used this concept to develop 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and potassium permanganate reduction (PPR) leaf disc assays. Reaction time and reagent concentration for these assays were optimized using leaves from spinach, kale, collards, mustard, and watermelon. Further, these assays were validated for linearity and intra-assay precision. Ultra-high performance liquid chromatography coupled to an electrospray quadrupole time-of-flight mass spectrometer (UPLC/ESI-HR-QTOFMS) was used for phytochemical profiling and studying relative abundances of certain phenolic compounds in various leaf discs suspended and cell-free extracts. The mass spectral analysis showed that leaf disc suspended methanolic extracts had almost same phytochemical profiles to those of cell-free extracts. The DPPH leaf disc assay demonstrated better radical scavenging potential than the conventional cell-free extract method. By contrast, the observed antioxidant activity values in ABTS and PPR leaf disc assays were lower than those of conventional cell-free extract-based methods. In conclusion, the developed leaf disc assays are simple and rapid for the qualitative and comparative assessment of the antioxidant potential of leaf samples, as well as can be a good alternative to conventional cell-free extract based methods.

Review on Bile Acids: Effects of the Gut Microbiome, Interactions with Dietary Fiber, and Alterations in the Bioaccessibility of Bioactive Compounds.

Bile acids are cholesterol-derived steroid molecules that serve various metabolic functions, particularly in the digestion of lipids. Gut microbes produce unconjugated and secondary bile acids through deconjugation and dehydroxylation reactions, respectively. Alterations in the gut microbiota have profound effects on bile acid metabolism, which can result in the development of gastrointestinal and metabolic diseases. Emerging research shows that diets rich in dietary fiber have substantial effects on the microbiota and human health. Plant-based foods are primary sources of bioactive compounds and dietary fiber, which are metabolized by microbes to produce different metabolites. However, the bioaccessibility of these compounds are not well-defined. In this review, we discuss the interaction of bile acids with dietary fiber, the gut microbiota, and their role in the bioaccessibility of bioactive compounds. To understand the possible mechanism by which bile acids bind fiber, molecular docking was performed between different dietary fiber and bile salts.

Establishment of a multicomponent dietary bioactive human equivalent dose to delete damaged Lgr5+ stem cells using a mouse colon tumor initiation model.

Multicomponent therapy has gained interest for its potential to synergize and subsequently lower the effective dose of each constituent required to reduce colon cancer risk. We have previously showed that rapidly cycling Lgr5 stem cells are exquisitely sensitive to extrinsic dietary factors that modulate colon cancer risk. In the present study, we quantified the dose-dependent synergistic properties of dietary n-3 polyunsaturated fatty acids (PUFA) and curcumin (Cur) to promote targeted apoptotic deletion of damaged colonic Lgr5 stem cells. For this purpose, both heterogeneous bulk colonocytes and Lgr5 stem cells were isolated from Lgr5-EGFP-IRES-CreER knock-in mice injected with azoxymethane (AOM). Isolated cells were analyzed for DNA damage (γH2AX), apoptosis (cleaved caspase-3), and targeted apoptosis (both γH2AX and cleaved caspase-3) at 12 h post-AOM injection. Comparison of the percentage of targeted apoptosis in Lgr5 stem cells (GFP) across a broad bioactive dose-range revealed an ED50 of 16.0 mg/day n-3 PUFA + 15.9 mg/day Cur. This corresponded to a human equivalent dose of 3.0 g n-3 PUFA + 3.0 g Cur. In summary, our results provide evidence that a low dose (n-3 PUFA + Cur) combination diet reduces AOM-induced DNA damage in Lgr5 stem cells and enhances targeted apoptosis of DNA-damaged cells, implying that a lower human equivalent dose can be utilized in future human clinical trials.

Simultaneous extraction of bioactive limonoid aglycones and glucoside from Citrus aurantium L. using hydrotropy.

Citrus limonoids were demonstrated to possess potential biological activities in reducing the risk of certain diseases. Limonoids are present in citrus fruits in the form of aglycones and glucosides. At present, limonoid aglycones and limonoid glucosides are extracted in multiple steps using different solvents. In order to understand their potential bioactivity, it may be beneficial to isolate and purify these compounds using environment friendly methods. A new method of extraction and purification of limonoids was established using a hydrotrope polystyrene adsorbent resin. Extraction of aglycones and glucosides was achieved in a single step, using an aqueous solution of sodium cumene sulphonate (Na-CuS). Sour orange (Citrus aurantium L.) seed powder was extracted with 2 M Na-CuS solution at 45 degrees C for 6 h. The filtered extract was diluted with water and loaded on an SP 700 adsorbent column. The column was washed with distilled water to remove the hydrotrope and then eluted using water and methanol in different compositions to obtain three compounds. The structures of the isolated compounds were confirmed by NMR spectroscopy as deacetyl nomilinic acid glucoside (DNAG), deacetyl nomilin (DAN) and limonin (LIM).

Red Mexican grapefruit: a novel source for bioactive limonoids and their antioxidant activity.

Citrus limonoids have shown to inhibit the growth of cancer in colon, lung, mouth, stomach and breast in animal and cell culture studies. For the first time in the present study, an attempt has been made to isolate antioxidant fractions and five limonoids from red Mexican grapefruit seeds. Defatted seed powder was successively extracted with hexane, ethyl acetate (EtOAc), acetone, methanol (MeOH) and MeOH/water and the extracts were concentrated under vacuum. Radical scavenging activity of 1,1-diphenyl-2-picrylhydrazyl (DPPH) and total phenolic content were also measured for comparison with the antioxidant capacity in the phosphomolybdenum method for the above extracts. Acetone and MeOH extracts, respectively, showed the highest (85.7%) and lowest (53.3%) radical scavenging activity, at 500 ppm. The total phenolic contents were found to be highest in the acetone extract (15.94%) followed by the MeOH extract (5.92%), ethyl acetate extract (5.54%) and water extract (5.26%). Antioxidant capacity of the extracts as equivalents to ascorbic acid (micromol/g of the extract) was in the order, EtOAc extract > acetone extract > water extract > methanol extract. Furthermore, the EtOAC and acetone extracts were loaded onto silica gel columns to obtain four limonoid aglycons. MeOH fraction was loaded onto a dowex-50 and sepabeads resin column to obtain a limonoid glucoside. The purity of the isolated five compounds was analyzed by HPLC using a C18 column and UV detection at 210 nm. Finally, the structures of the compounds were identified as obacunone, nomilin, limonin, deacetylnomilin (DAN) and limonin-17-beta-D-glucopyranoside (LG) using 1H and 13C NMR studies.