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Background: Canola meal has limited utilization in feed and food applications because of the presence of antinutritional factors and a high fiber content. Thus, the present study used 3-day canola seed sprouting followed by hull removal to improve the nutritional quality of canola as a feed and food ingredient to further enhance and diversify the canola market., Results: Seed sprouting and the hull removal process resulted in 63.2% sprouts, 29.3% mix fractions (MF) (hulls, ungerminated seed, and delayed sprouts) and 8.1% mass loss during sprouting. Fresh sprouts and MF were dried, ground and defatted to compare the obtained meals and oils with their counterparts of raw seed. Defatted sprouts (DFSP) resulted in a 46.2% reduction in crude fiber, a 34.3% reduction in acid detergent fiber and a 43.4% reduction in neutral detergent fiber compared to defatted raw seed (DFSE). DFSP provided a 10.1% higher protein content and a 5.9% increase in total amino acid content with higher essential amino acids compared to DFSE. Total carbohydrate was lowered by 5.5%, phytic acid content was lowered by 25.9%, and ash content was lowered by 5.5% in DFSP, whereas total glucosinolate content was higher in DFSP (13.1 μmol g -1 ) than in DFSE (8.8 μmol g -1 ). Sprouts and MF showed an oil content of 38.4% and 9.6%, respectively, compared to raw seed (34.5%)., Conclusion: Sprouting and hull removal of canola seed can potentially provide nutritive meal for food and feed applications. © 2022 Society of Chemical Industry., (© 2022 Society of Chemical Industry.)

The objective was to compare the impact of black lentil (BL) water and delphinidin 3-O-(2-O-β-d-glucopyranosyl-α-l-arabinopyranoside) (D3G)-rich lentil extracts on tumor development, inflammation and immune response in an azoxymethane (AOM)/dextran sodium sulfate (DSS) model. C57BL/6 mice were randomly separated into four groups: healthy control (n = 6), AOM/DSS control (n = 14), AOM/DSS + BL (600 mg/kg body wt, n = 12) and AOM/DSS + D3G (41 mg/kg body wt, equivalent to D3G concentration in BL, n = 12). Mice were given treatments for 11 weeks using a voluntary jelly administration. AOM/DSS + BL presented a lower (P < 0.05) disease activity index, throughout and at the end (2.4) compared with AOM/DSS (6.3). AOM/DSS + BL mice had an average of 7.8 neoplasms versus 12.8 for the AOM/DSS (P < 0.05). Proinflammatory cytokines were downregulated in the colon mucosa: interleukin (IL)-1β (-77.5%, -70.7%) and IL-6 (-44.4%, -44.9%) by AOM/DSS + BL and AOM/DSS + D3G, respectively, compared with AOM/DSS. IL-6 protein expression was decreased by BL in plasma (-72.6%) and gene expression in colon polyps (fold change: -4.0) compared with AOM/DSS. AOM/DSS + D3G non-polyp tissue gene expression clustered with the healthy control tissue with only four genes modified (secreted phosphoprotein 1 and CXC motif chemokine ligands 2, 5 and 10). AOM/DSS + BL downregulated programmed death-ligand 1 protein expression in colon tissue (-54.7%) and gene expression by 2.8-fold compared with the AOM/DSS control. In fecal samples, gallic and protocatechuic acids and epicatechin were found, and concentration of most amino acids was lower and unsaturated fatty acids were higher for AOM/DSS + BL and AOM/DSS + D3G. BL and D3G-rich extracts showed anti-inflammatory and proimmune response effects while BL additionally prevented growth of neoplasia., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The house cricket, Acheta domesticus L. (Orthoptera: Gryllidae) is one of the most important species of industrialized insects in the United States. Within the past five years the market of cricket powder as a food ingredient has been growing with increasing consumer interest on more sustainable sources of food. However, high labor costs of cricket production and high prices of cricket feed formulations result in cricket powder market prices much higher than other protein-rich food ingredients, making cricket powder only competitive within the novelty food market. In this study new diets formulated using by-products were developed using dietary self-selection followed by regression analysis. Crickets selected among seven different combinations of ingredients. Consumption ratios of food ingredients and by-products were used to determine macro and micro-nutrient intake. Regression analysis was used to determine the individual nutrient intake effect on cricket biomass production. Intake of vitamin C, sterol, manganese, and vitamins B1 and B5 had the most significant impact on live biomass production. Four diets were formulated based on this information and compared with a reference (Patton's 13) and a commercial diet. Although, crickets reared on Patton's diet 13 produced the most dry-weight biomass and developed the fastest, diet 4 (consisting of 92% by-products) generated the most profit (with a cost of $0.39 USD per kg) after an economic analysis that did not include the commercial formulation. Dry-weight biomass production was not significantly different among the four new diets and the commercial diet. This study demonstrated the value of dietary self-selection studies in developing oligidic insect diets and in studies of insect nutrition. This is the first such study involving farmed edible crickets and agricultural by-products. Four new cricket diet formulations contain between 62 and 92% agricultural by-products are included., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: The author Aaron T. Dossey as the owner of All Things Bugs LLC is planning to commercialize cricket feeds formulated based on this study, however formulations for commercialization are different than the ones reported in this manuscript. Plans of All Things Bugs to commercialize diets using this study do not alter our adherence to PLOS ONE policies on sharing data and materials.

Background: Some plant essential oils have insecticidal properties against mosquitoes and can be harnessed as ecofriendly tools for mosquito control. We conducted bioassays to determine the toxicity of Italian honeysuckle (Lonicera caprifolium) essential oil and its fractions against larvae of the yellow fever mosquito, Aedes aegypti., Results: Sixteen constituents were identified in honeysuckle essential oil compared to 15, 15, 15, and 11 constituents in fractions A, B, C, and E, respectively. The chemical constituents for fraction D were not determined due to lack of enough fraction sample. The two major constituents identified were patchouli alcohol (29.3%) and 6-acetyl-1,1,2,4,4,7-hexamethyltetralin (20.6%) in whole essential oil, alpha-bulnesene (27.6%) and 6-acetyl-1,1,2,4,4,7-hexamethyltetralin (23.2%) in fraction A, unknown chemical (47.3%) and diethyl phthalate (19.5%) in fraction B, unknown chemical (38.3%) and diethyl phthalate (23.2%) in fraction C, and patchouli alcohol (58.7%) and diethyl phthalate (20.5%) in fraction E. The LC 50 for whole essential oil was 34.4 ppm and significantly higher than 20.6, 19.7, 18.6, and 17.7 ppm for fractions B, C, D, and E, respectively. In contrast, fraction A was inactive. At 50 ppm, all individual constituents tested were less toxic than the whole essential oil with exception of patchouli alcohol, which caused 100% mortality., Conclusion: These findings suggest that patchouli alcohol is one of the chemical constituents responsible for bioactivity of honeysuckle essential oil and some of its fractions. The findings also demonstrate that honey suckle essential oil and its fractions can be exploited as a source of ecofriendly larvicides for mosquito control. Published 2019. This article is a U.S. Government work and is in the public domain in the USA., (Published 2019. This article is a U.S. Government work and is in the public domain in the USA.)

Black beans contain anthocyanins that could be used as colorants in foods with associated health benefits. The objective was to optimize anthocyanins extraction from black bean coats and evaluate their physicochemical stability and antidiabetes potential. Optimal extraction conditions were 24% ethanol, 1:40 solid-to-liquid ratio and 29°C (P<0.0001). Three anthocyanins were identified by MS ions, delphinidin-3-O-glucoside (465.1m/z), petunidin-3-O-glucoside (479.1m/z) and malvidin-3-O-glucoside (493.1m/z). A total of 32mg of anthocyanins were quantified per gram of dry extract. Bean anthocyanins were stable at pH 2.5 and low-temperature 4°C (89.6%), with an extrapolated half-life of 277days. Anthocyanin-rich extracts inhibited α-glucosidase (37.8%), α-amylase (35.6%), dipeptidyl peptidase-IV (34.4%), reactive oxygen species (81.6%), and decreased glucose uptake. Black bean coats are a good source of anthocyanins and other phenolics with the potential to be used as natural-source food colorants with exceptional antidiabetes potential., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Aims: The objective of this study was to determine the optimal pretreatment and fungal strain to reduce glucosinolates (GLS), fibre and residual sugars while increasing the nutritional value of canola meal., Methods and Results: Submerged incubation conditions were used to evaluate four pretreatment methods (extrusion, hot water cook, dilute acid and dilute alkali) and three fungal cultures (Aureobasidium pullulans Y-2311-1, Fusarium venenatum NRRL-26139 and Trichoderma reesei NRRL-3653) in hexane-extracted (HE) and cold-pressed (CP) canola meal., Conclusions: The combination of extrusion pretreatment followed by incubation with T. reesei resulted in the greatest overall improvement to HE canola meal, increasing protein to 51·5%, while reducing NDF, GLS and residual sugars to 18·6%, 17·2 μmol l -1  g -1 and 5% w/w, respectively. Extrusion pretreatment and incubation with F. venenatum performed the best with CP canola meal, resulting in 54·4% protein while reducing NDF, GLS and residual sugars to 11·6%, 6·7 μmol l -1  g -1 and 3·8% w/w respectively., Significance and Impact of the Study: The work is significant in that it provides a method of reducing GLS (up to 98%) and neutral detergent fibre (up to 65%) while increasing the protein content (up to 45%) of canola meal. This novel pretreatment and submerged incubation process could be used to produce a canola product with higher nutritional value for livestock consumption., (© 2017 The Society for Applied Microbiology.)

Anthocyanin-rich powders and aqueous extracts, with high antioxidant activities, were obtained from black bean seed coats and applied to colour a sport beverage. Idaho and Otomi bean coats were extracted in water-citric acid 2% (1/50, w/v), stirring for 4h at 40°C. Anthocyanins from Idaho and Otomi extracts (1.83mg and 1.02mg C3G/g, respectively) were applied to a commercially available sport beverage, with and without 2% β-cyclodextrin (βCD) under light and darkness conditions for 10days, and stored at 4°C and 25°C for 6weeks. At different light and storage conditions, anthocyanin degradation fitted a first-order reaction model. All bean coat anthocyanins combined with βCD showed extended half-life (up to 13months), higher D-values (up to 43months) and fewer differences in colourimetric properties (lightness, chroma and hue angle) under darkness and 4°C conditions. These black bean coat aqueous extracts and powders might represent natural alternatives to synthetic colorants, ecologically extracted, and with a high antioxidant potential., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Ethnopharmacological Relevance: Calea urticifolia leaves are traditionally used as a remedy to treat gastric ulcers, diabetes, and inflammation by the Xi'uy ancient native community of San Luis Potosi, Mexico., Aim of the Study: The aim was to assess the effects of the aqueous extract of the Mexican plant C. urticifolia as anti-inflammatory and anti-oxidant using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages and to provide evidence on the phenolic compounds., Materials and Methods: RAW 264.7 macrophages were stimulated with 1µg/mL of LPS and treated with 10, 25 50, 75 y 100µg/mL of Calea urticifolia lyophilized aqueous extract (CuAqE). Nitric oxide (NO) release, tumor necrosis factor alpha, prostaglandin E2 production, inducible nitric oxide synthase (iNOS), cyclooxygenase-2, nuclear factor-κB (NF-κB) p65, NF-κB p50 expression and reactive oxygen species (ROS) were measured; other pro-inflammatory proteins were measured with membrane antibody array. Phenolic compounds were analyzed by LC-ESI-MS., Results: Inflammation was inhibited by suppressing iNOS/NO pathway through inhibiting nucleus translocation of NF-κB p65 and p50 sub-units. ROS production was significantly (P<0.05) inhibited in a dose-dependent manner in LPS-stimulated macrophages. Moreover, the expression of inflammatory markers was suppressed (34.5-88.3%) by CuAqE. A mix of caffeoylquinic acid derivatives and flavonoid-glycosides were found in CuAqE., Conclusion: Phenolic compounds in CuAqE such as caffeoylquinic acid derivatives and flavonoid glycosides could be responsible for inhibiting LPS-induced inflammation and oxidative stress by iNOS/NO pathway through suppressing NF-κB signaling pathway and by inhibition of ROS production in RAW 264.7 macrophages. Therefore, these results support the traditional knowledge of C. urticifolia tea such as an anti-inflammatory and antioxidant agent., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Flavonoid and limonoid glycosides influence taste properties as well as marketability of Citrus fruit and products, particularly grapefruit. In this work, nine grapefruit putative natural product glucosyltransferases (PGTs) were resolved by either using degenerate primers against the semiconserved PSPG box motif, SMART-RACE RT-PCR, and primer walking to full-length coding regions; screening a directionally cloned young grapefruit leaf EST library; designing primers against sequences from other Citrus species; or identifying PGTs from Citrus contigs in the harvEST database. The PGT proteins associated with the identified full-length coding regions were recombinantly expressed in Escherichia coli and/or Pichia pastoris and then tested for activity with a suite of substrates including flavonoid, simple phenolic, coumarin, and/or limonoid compounds. A number of these compounds were eliminated from the predicted and/or potential substrate pool for the identified PGTs. Enzyme activity was detected in some instances with quercetin and catechol glucosyltransferase activities having been identified.

Pomacea maculata (formerly P. insularum), an apple snail native to South America, was discovered in Louisiana in 2008. These snails strip vegetation, reproduce at tremendous rates, and have reduced rice production and caused ecosystem changes in Asia. In this pilot study snails were exposed to two molluscicides, a tea (Camellia sinensis) seed derivative (TSD) or niclosamide monohydrate (Pestanal(®), 2',5-dichloro-4'-nitrosalicylanilide, CAS #73360-56-2). Mortality was recorded after exposure to high or low concentrations (0.03 and 0.015 g/L for TSD, 1.3 and 0.13 mg/L for niclosamide). The TSD induced 100 % mortality at both concentrations. Niclosamide caused 100 % and 17 % mortality at high and low concentrations respectively. These molluscicides were also tested on potential biocontrol agents, the red swamp crayfish (Procambarus clarkii) and redear sunfish (Lepomis microlophus). No crayfish mortalities occurred at either concentration for either chemical, but sunfish experienced 100 % mortality with TSD (0.03 g/L), and 21 % mortality with niclosamide (0.13 mg/L).

Previous studies have shown that a mutant of Arabidopsis that lacks the Toc159 receptor is impaired in chloroplast biogenesis. The mutant is referred as plastid protein import 2 or ppi2 and has an albino phenotype due to its inability to import the photosynthetic proteins. In this study, we measured fatty acid composition and transcript levels of plastid-localized fatty acid desaturases in the wild type and ppi2 mutant. The objective was to evaluate whether the Toc159 receptor was critical in the import of lipid-synthesizing enzymes. The ppi2 mutant accumulated decreased levels of oleic acid (18:1) and α-linolenic acid (18:3). The mutant accumulated drastically reduced amounts of the chloroplast lipid monogalactosyldiacylglycerol (MGDG), which contains more than 80% of 18:3. The expression of genes that encode stearoyl-ACP desaturase and MGD1 synthase were down-regulated in the ppi2 mutant, and this corresponded to decreased levels of 18:1 and MGDG, respectively. We conclude that in the ppi2 mutant the impaired synthesis of MGDG resulted in decreased amounts of 18:3. The mutant however, had a 30-fold increase in fad5 transcript levels; this increase was mirrored by a 16- to 50-fold accumulation of hexadecatrienoic acid (16:3), a fatty acid found exclusively in MGDG. Taken together, these data suggest that the Toc159 receptor is required in the import of stearoyl-ACP desaturase and MGD1 synthase into the chloroplasts. Since the expression of fad5 gene was up-regulated in the ppi2 mutant, we propose that fad5 desaturase is imported into plastids through the atToc132/atToc120 protein import pathway., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

The in vivo bioavailability of the flavone-C-glycosides has been little studied compared to their O-glycoside analogues, which are both more common in nature and considered more easily hydrolyzed than C-glycosides, by both enterocytes and gut microbiota. In this study, we used vitexin-2-O-xyloside (VOX), an apigenin-8-C-glucoside-2-O-xyloside, purified from seeds of Swiss chard (Beta vulgaris cicla), to investigate VOX absorption into portal blood compared to its aglycone, apigenin. We used a rat model in which we ligated the ileo- and colo-caecal junctions, then administered apigenin or VOX directly into the caecum. Blood samples were drawn from the portal vein at timed intervals over 40 min. The kinetic profile of appearance in portal blood of the compounds and their metabolites was evaluated by HPLC-ESI-MS. Apigenin was found in portal blood both as the aglycone and as an apigenin-glucuronide derivative. The VOX was found unchanged and as a reduced monoglycoside, which underwent glucuronidation. By collecting the bile, we confirmed that the liver received unchanged VOX, which was returned to the gut by enterohepatic recirculation for reabsorption from the ileum. The amount of apigenin and VOX remaining in the caecum accounted for ∼15% and ∼26%, respectively. These data show for the first time that the C-glycoside VOX is absorbed unchanged and undergoes enterohepatic recirculation in addition to hydrolysis to the monoglycoside, reduction and conjugation to form a bioavailable glucuronide.

Soybean contains constituents that have antinutritional and bioactive properties. Enzymatic hydrolysis and germination can enhance the biological activity of these compounds in soybean. The objective of this study was to investigate the effect of germination, Alcalase (protease) hydrolysis, and their combination on the concentrations of antinutritional and bioactive compounds in Brazilian soybean cultivar BRS 133. A combination of germination and Alcalase hydrolysis resulted in the degradation of Bowman-Birk inhibitor (BBI), Kunitz trypsin inhibitor (KTI), and lunasin by 96.9, 97.8, and 38.4%. Lectin was not affected by any of the processing treatments when compared to nongerminated and nonhydrolyzed soy protein extract. Total isoflavones (ISF) and total saponins (SAP) increased by 16.2 and 28.7%, respectively, after 18 h of germination, while Alcalase hydrolysis led to the reduction of these compounds. A significant correlation was found between concentrations of BBI and KTI, BBI and lunasin, BBI and ISF, KTI and lunasin, KTI and ISF, KTI and SAP, lunasin and ISF, and ISF and SAP. Germination and Alcalase hydrolysis interacted in reducing BBI, ISF, and SAP. This study presents a process of preparing soy flour ingredients with lower concentrations of antinutritional factors and with biologically active constituents, important for the promotion of health associated with soybean consumption. In conclusion, 18 h of germination and 3 h of Alcalase hydrolysis is recommended for elimination of protease inhibitors, while bioactives are maintained by at least 50% of their original concentrations.

Camelina sativa is an oilseed plant rich in n-3 and n-6 fatty acids and extruding the seeds results in high protein meal (*40%) containing high levels of n-3 fatty acids. In this study, we examined the effects of feeding extruded defatted camelina meal to commercial laying hens, measuring egg production, quality, and fatty acid composition. Lohmann White Leghorn hens (29 weeks old) were randomly allocated to three dietary treatment groups (n = 25 per group) and data was collected over a 12 week production period. All the treatment groups were fed a corn soy based experimental diet containing 0% (control), 5, or 10% extruded camelina meal. We found no significant differences in percent hen-day egg production and feed consumed per dozen eggs. Egg shell strength was significantly higher in both camelina groups compared to the controls. Egg total n-3 fatty acid content increased 1.9- and 2.7-fold in 5 and 10% camelina groups respectively relative to the control. A similar increase in DHA content also occurred. Further camelina meal did not alter glucosinolate levels and no detectable glucosinolates or metabolic product isothiocyanates were found in the eggs from either the 5 or 10% camelina groups. These results indicate that camelina meal is a viable dietary source of n-3 fatty acids for poultry and its dietary inclusion results in eggs enriched with n-3 fatty acids.

Distillers dried grains (DDG) have potential to be a nutritionally important source of protein, oil and phenolic antioxidants. DDG was subjected to high-shear and jet-cooking, with or without alkaline pH adjustment and autoclaving. Soluble and insoluble fractions were analyzed for protein, oil and ash. Extracts were analyzed for phenolic acids and antioxidant activity. Protein contents were significantly elevated in the insoluble fractions after treatment and the oil content was drastically increased in the insoluble fraction after high-shear and jet-cooking without pH adjustment. Alkaline pH adjustment resulted in a soluble fraction that was highest in phenolic acids, but not antioxidant activity. The highest antioxidant activity was found in the 50% ethanol extract from DDG that had been subjected to high-shear and jet-cooking. These results suggest that high-shear and jet-cooking may be useful processing treatments to increase the value of DDG by producing fractions high in protein, oil and extractable phenolic acids with high antioxidant activity. The DDG fractions and extracts described herein may be useful as food and nutraceutical ingredients, and, if used for these applications, will increase the value of DDG and ease economic burdens on ethanol producers, allowing them to compete in the bio-fuel marketplace.

Obesity is a worldwide health concern and a well recognized predictor of premature mortality associated with a state of chronic inflammation. The objective was to evaluate the effect of soy protein hydrolysates (SPH) produced from different soybean genotypes by alcalase (SAH) or simulated gastrointestinal digestion (SGIH) on lipid accumulation in 3T3-L1 adipocytes. The anti-inflammatory effect of SPH produced by alcalase on LPS-induced macrophage RAW 264.7 cell line was also investigated. SAH (100 microM) derived from soybean enriched in beta-conglycinin (BC) (up to 47% total protein) decreased lipid accumulation (33-37% inhibition) through downregulation of gene expression of lipoprotein lipase (LPL) and fatty acid synthase (FAS). SGIH (100 microM) inhibited lipid accumulation to a lesser extent (8-14% inhibition) through inhibition of LPL gene expression. SAH (5 microM) decreased the production of nitric oxide (NO) (18-35%) and prostaglandin E(2) (PGE(2)) (47-71%) and the expression of inducible nitric oxide synthase (iNOS) (31-53%) and cycloxygenase-2 (COX-2) (30-52%). This is the first investigation showing that soy hydrolysates inhibit LPS-induced iNOS/NO and COX-2/PGE(2 )pathways in macrophages. Soybeans enriched in BCs can provide hydrolysates that limit fat accumulation in fat cells and inflammatory pathways in vitro and therefore warrant further studies as a healthful food.

Spearmint has one major monoterpene, (-)-carvone, that constitutes up to 90% of all the monoterpenes present. Likewise, the major phenylpropanoid-rosmarinic acid-in spearmint accounts for up to 70% of the phenylpropanoids produced from the plant. These two compounds are each produced by separate distinct biosynthetic pathways which provide an excellent opportunity to study the influence of a wide number of environmental and chemical conditions on secondary metabolism and plant growth. The techniques presented in this chapter employ 1 g of fresh weight material for each secondary metabolite analyses. Analysis of single compounds obtained from the two distinct metabolic pathways simplifies the interpretation of the metabolic results allowing for direct correlations of culture factors on secondary metabolism.

The influence of autoclaved fungal materials such as culture filtrate, freeze-dried mycelium (FDM), mycelium suspension, and spore suspension (SS) on the growth, morphogenesis, and carvone production of spearmint (Mentha spicata L.) plants was studied. Fungal materials were either applied as a drench or spray on the plants. Spearmint plants (cv. "294099") drenched with SS (1 x 10(8) spores/ml) of Trichoderma reesei showed no significant differences in leaf numbers, root numbers, or shoot numbers compared with nontreated controls. However, significantly higher fresh weights and carvone levels were observed in plants drenched with T. reesei SS compared with the untreated controls. Fungal materials derived from Aspergillus sp., Fusarium graminearum, F. sporotrichoides, Penicillium sp., P. acculeatum, Rhizopus oryzae, and T. reesei were sprayed on spearmint foliage. F. graminearum, F. sporotrichoides, or R. oryzae elicited no enhanced growth, morphogenesis, or secondary metabolism responses. The best growth and morphogenesis responses were obtained employing Aspergillus sp., Penicillium sp., or T. reesei foliar sprays. For example, spearmint cv. "557807" plants sprayed with 100 mg/l FDM T. reesei isolate NRRL 11460 C30 stimulated higher fresh weights (75%), shoot numbers (39%), leaf numbers (57%), and root numbers (108%) compared with untreated plants. This effect was not dose-dependent because similar growth and morphogenesis responses were obtained by testing 10, 100, or 1000 mg/l FDM concentrations. Carvone levels in fungal-treated foliar-sprayed plants were comparable to nontreated controls. However, total carvone levels per plant were higher in fungal-treated plants because of their increased fresh weight.

The impact of triterpenoid saponins isolated from soybeans on suppression of colon cancer cell proliferation was evaluated. Experiments were conducted to determine the effects of a purified soybean B-group saponin extract on cell proliferation, cell-cycle distribution and programmed cell death in cultures of human HCT-15 colon adenocarcinoma cells. Treatment of cells with the soyasaponins at concentrations of 25-500 p.p.m. significantly reduced viable cell numbers after 24 and 48 h of exposure. Treatment of cells with 25 and 100 p.p.m. of saponins also resulted in a transient accumulation of cells in the S-phase of the cell cycle that was associated with a significant reduction of cyclin-dependant kinase-2 (CDK-2) activity. More striking was that, when examined by transmission electron microscopy, soyasaponin-treated cells exhibited an approximately 4.5-fold increase in cell morphologies characteristic of Type II non-apoptotic programmed cell death (PCD) including numerous autophagic vacuoles, changes that collectively suggest autophagic cell death. In addition, the protein levels of microtubule-associated protein light chain 3 (LC-3), a specific marker of macroautophagy, increased substantially following soyasaponin treatment. Taken together these results thus indicate that soybean saponins, at physiologically relevant doses, can suppress HCT-15 colon cancer cell proliferation through S-phase cell-cycle delay, and can induce macroautophagy, the hallmark of Type II PCD. These findings suggest that B-group soyasaponins may be another colon-cancer suppressive component of soy that warrants further examination as a potential chemopreventive phytochemical.

Although metabolites of furanocoumarins have been characterized in a wide range of organisms, to date they have been identified in only a single insect species, Papilio polyxenes. Depressaria pastinacella, the parsnip webworm, like P. polyxenes a specialist on Apiaceae, routinely consumes plant tissues higher in furanocoumarin content than does P. polyxenes and is capable of faster cytochrome P-450-mediated detoxification of these compounds. In this study, we characterized metabolites of xanthotoxin, a linear furanocoumarin, and sphondin, an angular furanocoumarin, in midguts and frass of parsnip webworms. Two metabolites were isolated and identified from webworms fed artificial diet containing xanthotoxin. LC-ESI-MS analysis resulted in the determination of a MW of 266 for the compound in the frass and one of the compounds in the midgut; 1H NMR confirmed its structure as 6-(7-hydroxy-8-methoxycoumaryl)-hydroxyacetic acid (HCHA). The second compound from the midgut had a MW of 252 and was identified by 1H NMR and 13C NMR analysis as 6-(7-hydroxy-8-methoxycoumaryl)-hydroxyethanol) (HMCH). Whereas HCHA has been found in frass of Papilio polyxenes fed xanthotoxin, HMCH has not been reported previously in insects. Although the first step of metabolism of xanthotoxin in webworms as well as P. polyxenes is likely the formation of an epoxide on the furan ring, angular furanocoumarin metabolism in webworms appears to differ. The principal metabolite of sphondin was identified as demethylated sphondin (6-hydroxy-2H-furo[2,3-h]-1-benzopyran-2-one) by LC-ESI-MS and confirmed by 1H NMR and 13C NMR analyses. That webworms produce metabolites of xanthotoxin in common not only with other Lepidoptera (e.g., HCHA) but with other vertebrates (e.g., HMCH) suggests a remarkable conservatism in the metabolic capabilities of cytochrome P-450s and raises the possibility that insects may share other detoxification reactions with vertebrates with respect to toxins in foodplants.

In previous studies, we demonstrated that tyrosine hydroxylase and neurofilament proteins are regulated by chronic morphine and chronic cocaine treatments in the ventral tegmental area in Sprague-Dawley rats and that the inbred Lewis and Fischer 344 rat strains, under drug-naive conditions, show different levels of these proteins specifically in this brain region. In the current study, we compared Lewis and Fischer rats with respect to levels of adenylate cyclase, cyclic AMP-dependent protein kinase and G-proteins in the nucleus accumbens (NAc) and locus coeruleus (LC), brain regions in Sprague-Dawley rats where these proteins are regulated by chronic exposure to morphine or to cocaine. We found that levels of adenylate cyclase and cyclic AMP-dependent protein kinase activity are higher in the NAc and LC of Lewis rats compared to Fischer rats, whereas levels of Gi alpha and G beta were lower. These strain differences were not seen in several other brain regions analyzed and no strain differences were detected in levels of other G-protein subunits. Lewis and Fischer rats also differed in the ability of chronic morphine to regulate adenylate cyclase and cyclic AMP-dependent protein kinase in the NAc and LC. In the NAc, chronic morphine increased levels of the two enzymes in the Fischer strain only, whereas in the LC chronic morphine increased levels of the enzymes in both strains, with more robust effects seen in the Lewis rat. To understand possible physiological consequences of these strain differences in the cyclic AMP pathway, we studied LC neuronal activity under basal and chronic morphine-treated conditions. LC neurons of Lewis rats showed higher spontaneous firing rates in brain slices in vitro than those of Fischer rats and also showed greater morphine-induced increases in responsiveness to bath-applied 8-bromo-cyclic AMP. These electrophysiological findings are generally consistent with the biochemical observations. Moreover, Lewis and Fischer rats displayed very different opiate withdrawal syndromes, with different types of behaviors elicited upon precipitation of opiate withdrawal with the opiate receptor antagonist, naltrexone. The possible relationship between these behavioral findings and the biochemical and electrophysiological data is discussed. These studies provide further support for the possibility that Lewis and Fischer rat strains provide a useful model system in which some of the genetic factors that contribute to drug-related behaviors can be investigated.

Operant responses involving a cued discrimination are sensitively disrupted by neuroleptic drugs that block dopamine (DA) receptors in the brain; however, it is not clear which DA receptor subtypes may be involved in these effects. The role of D1 or D2 DA receptor antagonists on the execution of a conditioned reaction-time (RT) motor task was investigated in the present study. Rats were trained to release a lever after the presentation of a visual cue within a RT limit to be reinforced by a food pellet. The D1 receptor antagonist SCH-23390, at doses that significantly decrease the behavioral effects of cocaine, did not impair performance at any dose (5, 10, or 20 micrograms/kg) injected subcutaneously. In contrast, a selective D2 receptor antagonist raclopride (50, 100, or 200 micrograms/kg) induced a dose-dependent increase in the number of incorrect responses (release of the lever over the RT limit) associated with an increase in the RT. The results suggest that the dopaminergic nigrostriatal system, which has previously been shown to be specifically involved in this RT task (Amalric and Koob 1987), appears to be a sensitive site for sensorimotor integration, and that the execution of the conditioned RT motor task may depend preferentially on the activation of the dopaminergic D2 receptors in this system.