Formation Of Isothiocyanates Research Articles

Enhancing the formation of functional glucosinolate degradation products in fermented broccoli stalk by-product with lactic acid bacteria

Broccoli stalk by-product (BsBP) is rich in glucosinolates (GSLs). Its fermentation process is generally characterized by the degradation of GSLs and formation of bioactive isothiocyanates (ITCs), in which lactic acid bacteria (LAB) play an important role. The GSLs-degrading capacity of 61 LAB strains was investigated in vitro. Lacticaseibacillus paracasei YC5, Pediococcus pentosaceus RBHZ36, and Lactiplantibacillus plantarum ND1, with high potential to transform GSLs into ITCs, were screened. The functional GSL degradation products (total content of sulforaphane, indol-3-carbinol, and ascorbigen) increased 22.0–33.5 % compared to natural fermentation after 24 h when BsBP was fermented by the three screened strains in pure culture. LAB fermentation also helped to increase the quantity of indolic GSL degradation products in BsBP brine, suggesting that LAB fermentation promoted BsBP GSLs transformation into bioactive ITCs. The proposed use of the LAB strains characterized in this study provided a fermented BsBP and brine with high profile of functional GSL degradation products.

Acidification and tissue disruption affect glucosinolate and S-methyl-l-cysteine sulfoxide hydrolysis and formation of amines, isothiocyanates and other organosulfur compounds in red cabbage (Brassica oleracea var. capitata f. rubra)

Cabbages are rich in sulfur-containing metabolites like glucosinolates (GLSs) and S-methyl-l-cysteine sulfoxide (SMCSO). Tissue disruption initiates hydrolysis of these compounds and bioactive volatile hydrolysis products such as isothiocyanates (ITCs), sulfides, and thiosulfinates are formed. However, nitriles, epithionitriles, or amines can also result from GLSs. Here, the influence of hydrolysis time, extent of tissue disruption (chopping vs. homogenization), and addition of lemon juice or vinegar on the outcome of enzymatic hydrolysis of GLSs and SMCSO was investigated in red cabbage. Chopping led to partial hydrolysis of GLSs, whereas homogenization completely degraded GLSs but only had a small effect on SMCSO. Homogenization increased amine formation from alkenyl and methylthioalkyl ITCs, but not from methylsulfinylalkyl ITCs. Acidification inhibited formation of products from SMCSO. Further, it reduced nitrile and epithionitrile formation and stopped amine formation, thereby increasing ITC levels. Therefore, acidification is a valuable mean to enhance ITC levels in fresh Brassica foods.

Degradation of glucosinolates and formation of isothiocyanates, nitriles, amines, and N,N’-dialk(en)yl thioureas during domestic boiling of red cabbage

Red cabbage is a popular vegetable in Central Europe and a rich source of glucosinolates (GLSs). Upon hydrolysis, GLSs form health-promoting isothiocyanates (ITCs), but also nitriles and epithionitriles. Recently, ITCs were shown to undergo further hydrolysis, yielding amines. Here, we analyzed the degradation of GLSs and the formation of ITCs, nitriles, epithionitriles, and amines during domestic-like cooking of red cabbage with addition of vinegar or baking soda. Both additives strongly affected the stability of GLSs and the formation of nitriles during boiling. Primary amines were found as a major degradation product of GLSs. In control and vinegar samples, formation of methylsulfinylalkyl amines increased during boiling. Additionally, for the first time, the formation of several N,N’-dialk(en)yl thioureas during boiling of Brassica vegetables was demonstrated, resulting from the reaction of GLS-derived ITCs and amines, and they were subsequently quantified. As references, five N,N’-dialk(en)yl thioureas were synthesized and characterized by NMR and HRMS.

Effect of long-term storage on glucosinolate and S-methyl-l-cysteine sulfoxide hydrolysis in cabbage (Brassica oleracea var. capitata)

Cabbages are good sources for glucosinolates and S-methyl-l-cysteine sulfoxide (SMCSO), precursors to bioactive volatile hydrolysis products such as isothiocyanates, sulfides and thiosulfinates. Often, white and red cabbages are stored at 0 °C for many months before being sold. Here, we investigated the effect of storage for up to eight months on glucosinolates, SMCSO and the formation of isothiocyanates and derived amines, (epithio)nitriles and volatile organosulfur compounds (VOSCs) in white and red cabbages. Further, the effect of storage on protein expression was evaluated. Overall, glucosinolates and well as SMCSO contents were stable during storage. While in white cabbage glucosinolate hydrolysis was not much affected, in red cabbage storage increased formation of isothiocyanates and methylthioalkylamines, which was linked with reduced epithiospecifier protein 1 abundance. Long-term storage reduced formation of VOSCs from SMCSO which correlated with decline in predicted cystine lyase. Therefore, storage maintains these phytonutrients and can increase formation of health-promoting isothiocyanates.

Metabolism of plant-derived toxins from its insect host increases the success of the entomopathogenic fungus Beauveria bassiana

Beauveria bassiana is a soil fungus that parasitizes a large number of arthropod species, including numerous crop pests, causing white muscardine disease and is therefore used as a biological insecticide. However, some insects, such as the cabbage aphid (Brevicoryne brassicae), defend themselves chemically by sequestering dietary pro-toxins (glucosinolates) from their Brassicales host plants. Glucosinolates are accumulated by cabbage aphids and activated to form toxic isothiocyanates when under attack. While isothiocyanate formation protects aphids against most attackers, B. bassiana is still able to infect the cabbage aphid under natural conditions. We therefore investigated how this fungus is able to circumvent the chemical defense system of the cabbage aphid. Here, we describe how B. bassiana infection activates the cabbage aphid defense system, but the resulting toxins are metabolized by B. bassiana via the mercapturic acid pathway, of which the first step is catalyzed by glutathione-S-transferases of low substrate specificity. This detoxification pathway enhances B. bassiana growth when isothiocyanates are present in natural concentrations, and so appears to be an important factor in fungal parasitization of these chemically defended aphids.

Perubahan Fıtokımıa Kecambah Kembang Kol (Brassica oleraceae L., var. botrytis) sebagai Respons terhadap Cahaya Biru

The aim of this study was to observe the use of blue light on phytochemical changes during cauliflower (Brassica oleraceae L., var. botrytis) germination and the correlation between the parameters measured. The treatment was in the form of giving blue light and dark conditions as a control. Measurement of phytochemical compounds including ascorbic acid, glucosinolates, isothiocyanates and myrosinase enzymes was carried out every day for 5 days of germination. This research method is experimental research. Data obtained from measurements for all parameters are presented in the form of mean ± standard error. Furthermore, all data were analyzed using analysis of variance (ANOVA). To determine differences between groups, Duncan's multiple range test (DMRT) was used at the 95% confidence level. All data analysis used SPSS software version 24.0 (IBM, Chicago, IL, USA). In addition, the measurement of the correlation between parameters is carried out using the Pearson rank correlation coefficient. The results showed that blue light treatment significantly increased all measured phytochemical parameters. A significant positive correlation occurred between myrosinase activity and isothiocyanate formation. The glucosinolate content of cauliflower sprouts remained high during observations in the blue light treatment compared to the dark conditions. Blue light induces isothiocyanate formation and higher myrosinase activity when compared to dark conditions. The conclusion from this study is that blue light significantly increases the phytochemical changes of cauliflower sprouts.
 
 Keywords: Ascorbic Acid, Glucosinolate, Isothiocyanate, Correlation, Myrosinase

CaCl2–HCl electrolyzed water promotes glucosinolate metabolism in broccoli sprouts via calcium signalling

BackgroundElectrolyzed water, a green chemistry invention, is a novel disinfectant that has been widely used in the food field. Previous studies have found that slightly acidic electrolyzed water not only has a bactericidal effect but also promotes the accumulation of active substances in many types of sprouts. We developed a new type of electrolyzed water, CaCl2–HCl electrolyzed water (CHEW), which can effectively enhance the formation of glucosinolates and isothiocyanates in broccoli sprouts. The molecular mechanism by which CaCl2–HCl electrolyzed water promotes glucosinolate metabolism in broccoli sprouts during growth was investigated in this study.ResultsOn the 4th day, the total glucosinolate content in broccoli sprouts reached 195.50 μmol/g DW under CHEW treatment. On the 8th day, compared to treatment with tap water, the contents of three isothiocyanates (sulforaphane, erucin, and 4-isothiocyanato-1-butene) in broccoli sprouts under CHEW treatment were significantly increased by 45.93, 34.04 and 185.15%, respectively. The results of calcium distribution and qRT‒PCR assessments indicated that CHEW treatment activated calcium signalling in broccoli sprouts and promoted the relative expression of glucosinolate biosynthesis-related genes through calcium sensors [calmodulin (CaM), calmodulin-like protein (CML), and calcium-dependent protein kinase (CDPK)] and the calcium-binding nuclear protein IQD1.ConclusionOur research provides new insights into the molecular mechanism of glucosinolate metabolism and a theoretical basis for cultivating high-quality broccoli sprouts.Graphical

Comprehensive Evaluation of the Bioactive Composition and Neuroprotective and Antimicrobial Properties of Vacuum-Dried Broccoli (Brassica oleracea var. italica) Powder and Its Antioxidants.

In this study, vacuum drying (VD) was employed as an approach to protect the bioactive components of and produce dried broccoli powders with a high biological activity. To achieve these goals, the effects of temperature (at the five levels of 50, 60, 70, 80 and 90 °C) and constant vacuum pressure (10 kPa) were evaluated. The results show that, with the increasing temperature, the drying time decreased. Based on the statistical tests, the Brunauer-Emmett-Teller (BET) model was found to fit well to sorption isotherms, whereas the Midilli and Kucuk model fit well to the drying kinetics. VD has a significant impact on several proximate composition values. As compared with the fresh sample, VD significantly reduced the total phenol, flavonoid and glucosinolate contents. However, it was shown that VD at higher temperatures (80 and 90 °C) contributed to a better antioxidant potential of broccoli powder. In contrast, 50 °C led to a better antimicrobial and neuroprotective effects, presumably due to the formation of isothiocyanate (ITC). Overall, this study demonstrates that VD is a promising technique for the development of extracts from broccoli powders that could be used as natural preservatives or as a neuroprotective agent.

Impact of epithiospecifier proteins on the formation of isothiocyanates, nitriles and epithionitriles from glucosinolate hydrolysis in kohlrabi

LebensmittelchemieVolume 76, Issue S2 p. S2-347-S2-347 Weitere Themen (ATW-W) Impact of epithiospecifier proteins on the formation of isothiocyanates, nitriles and epithionitriles from glucosinolate hydrolysis in kohlrabi K. G. Mbudu, K. G. Mbudu Großbeeren/DSearch for more papers by this authorK. Witzel, K. Witzel Großbeeren/DSearch for more papers by this authorF. Börnke, F. Börnke Großbeeren/D Potsdam/DSearch for more papers by this authorF. S. Hanschen, F. S. Hanschen Großbeeren/D Leibniz Institute of Vegetable and Ornamental Crops (IGZ), Theodor-Echtermeyer-Weg 1, 14979 Großbeeren/DSearch for more papers by this author K. G. Mbudu, K. G. Mbudu Großbeeren/DSearch for more papers by this authorK. Witzel, K. Witzel Großbeeren/DSearch for more papers by this authorF. Börnke, F. Börnke Großbeeren/D Potsdam/DSearch for more papers by this authorF. S. Hanschen, F. S. Hanschen Großbeeren/D Leibniz Institute of Vegetable and Ornamental Crops (IGZ), Theodor-Echtermeyer-Weg 1, 14979 Großbeeren/DSearch for more papers by this author First published: 01 September 2022 https://doi.org/10.1002/lemi.202259292 Funding: F. S. Hanschen and this study is funded by the Leibniz Association (Leibniz-Junior Research Group OPTIGLUP; J16/2017). AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume76, IssueS2September 2022Pages S2-347-S2-347 RelatedInformation

Supplementation of cooked broccoli with exogenous moringa myrosinase enhanced isothiocyanate formation

Brassica vegetables, especially broccoli, have health benefits such as anticancer activity, which are attributed to isothiocyanate (ITC), products of glucosinolate hydrolysis. This study aimed to explore the effect of cooking time and addition of exogenous myrosinase (MYR) from moringa seeds on the yield of ITCs. The results showed that raw broccoli produced a significantly high amount of ITCs, which decreased by almost 40% after microwaving the broccoli for 1 min. Introducing exogenous MYR by adding ground moringa seeds to cooked broccoli caused a notable increase in ITC of 38%. At pH 4.0–6.0, MYR showed optimal activity, and the thermal stability of MYR from moringa seeds was better than that from broccoli. The kinetic parameters indicated that MYR from moringa seeds had a higher affinity to sinigrin than that from broccoli seeds. This study was novel in reporting that adding ground moringa seeds to cooked broccoli enhanced ITC formation.

β-Cyanoalanine synthase protects mites against Arabidopsis defenses.

Glucosinolates are antiherbivory chemical defense compounds in Arabidopsis (Arabidopsis thaliana). Specialist herbivores that feed on brassicaceous plants have evolved various mechanisms aimed at preventing the formation of toxic isothiocyanates. In contrast, generalist herbivores typically detoxify isothiocyanates through glutathione conjugation upon exposure. Here, we examined the response of an extreme generalist herbivore, the two-spotted spider mite Tetranychus urticae (Koch), to indole glucosinolates. Tetranychus urticae is a composite generalist whose individual populations have a restricted host range but have an ability to rapidly adapt to initially unfavorable plant hosts. Through comparative transcriptomic analysis of mite populations that have differential susceptibilities to Arabidopsis defenses, we identified β-cyanoalanine synthase of T. urticae (TuCAS), which encodes an enzyme with dual cysteine and β-cyanoalanine synthase activities. We combined Arabidopsis genetics, chemical complementation and mite reverse genetics to show that TuCAS is required for mite adaptation to Arabidopsis through its β-cyanoalanine synthase activity. Consistent with the β-cyanoalanine synthase role in detoxification of hydrogen cyanide (HCN), we discovered that upon mite herbivory, Arabidopsis plants release HCN. We further demonstrated that indole glucosinolates are sufficient for cyanide formation. Overall, our study uncovered Arabidopsis defenses that rely on indole glucosinolate-dependent cyanide for protection against mite herbivory. In response, Arabidopsis-adapted mites utilize the β-cyanoalanine synthase activity of TuCAS to counter cyanide toxicity, highlighting the mite’s ability to activate resistant traits that enable this extreme polyphagous herbivore to exploit cyanogenic host plants.

Selection of broccoli (Brassica oleracea var. italica) on composition and content of glucosinolates and hydrolysates

Three commercial cultivars and eleven inbred broccoli lines were used to investigate the glucosinolate (GSL) profile, hydrolysate and nitrile contents, and nitrile formation rate. Glucobrassicin (BRA) and glucoraphanin (GRA) were the most dominant GSLs in all lines. The 5405 and 5406 lines exhibited the highest total GSL content (18.39 and 22.97 μmol·g−1 DW, respectively). Altogether, five lines (5306, 5311, 5312, 5404, and 5406) were selected as potential highly functional broccoli lines based on GSL content. Three lines (5405, 5406, and 5311) exhibited relatively higher total hydrolysates than the other lines. Three inbred lines (5404, 5407, and 5410) with low nitrile formation rate showed a high isothiocyanate (ITC) formation (2.29 ± 0.40, 2.16 ± 0.08, 2.24 ± 0.47 μmol·g−1 DW) and low nitrile formation (0.72 ± 0.12, 0.64 ± 0.12, 0.90 ± 0.20 μmol·g−1 DW). Therefore, lines 5404, 5407, and 5410 were selected as functional broccoli breeding lines based on their hydrolysate content and nitrile formation rates. Consequently, hydrolysate and GSL content are more reliable than GSL content only for selecting and developing highly functional varieties.

Development of a moisture-activated antimicrobial film containing ground mustard seeds and its application on meat in active packaging system

An antimicrobial cellulose acetate film containing finely ground mustard seeds was developed. The effect of the film properties on the allyl isothiocyanates (AITC) formation and release into the headspace was investigated. Less porous structure and larger thickness caused slower moisture absorption and consequently a slower AITC formation and release rate. For its application in food, the film was tested on whether the concentration of AITC was sufficient to inhibit bacterial growth in ground beef with different fat contents. The AITC concentration for the low-fat ground beef was lower in the product and higher in the headspace compared to medium fat ground beef. The shelf life was extended by 3.7 and 0.6 days for low fat and medium fat ground beef respectively. Besides a novel way to control the compounds’ release to the packaging system, this study also shows the importance of the food composition for tailoring the effective active packaging. (150 words).

Metabolic Fate of Dietary Glucosinolates and Their Metabolites: A Role for the Microbiome.

Robust evidence shows that phytochemicals from cruciferous vegetables, like broccoli, are associated with numerous health benefits. The anti-cancer properties of these foods are attributed to bioactive isothiocyanates (ITCs) and indoles, phytochemicals generated from biological precursor compounds called glucosinolates. ITCs, and particularly sulforaphane (SFN), are of intense interest as they block the initiation, and suppress the progression of cancer, through genetic and epigenetic mechanisms. The efficacy of these compounds is well-demonstrated in cell culture and animal models, however, high levels of inter-individual variation in absorption and excretion of ITCs is a significant barrier to the use of dietary glucosinolates to prevent and treat disease. The source of inter-individual ITC variation has yet to be fully elucidated and the gut microbiome may play a key role. This review highlights evidence that the gut microbiome influences the metabolic fate and activity of ITCs. Human feeding trials have shown inter-individual variations in gut microbiome composition coincides with variations in ITC absorption and excretion, and some bacteria produce ITCs from glucosinolates. Additionally, consumption of cruciferous vegetables can alter the composition of the gut microbiome and shift the physiochemical environment of the gut lumen, influencing the production of phytochemicals. Microbiome and diet induced changes to ITC metabolism may lead to the decrease of cancer fighting phytochemicals such as SFN and increase the production of biologically inert ones like SFN-nitrile. We conclude by offering perspective on the use of novel “omics” technologies to elucidate the interplay of the gut microbiome and ITC formation.

Effects of Prohydrojasmon on the Number of Infesting Herbivores and Biomass of Field-Grown Japanese Radish Plants.

Prohydrojasmon (PDJ), an analog of jasmonic acid (JA), was found to induce direct and indirect defenses against herbivores in non-infested plants. To test whether PDJ can be used for pest control in crop production, we conducted experiments in pesticide-free Japanese radish fields from October 4 to December 12 in 2015. Twenty-four Japanese radish plants in three plots were treated with a 100 times-diluted commercial formulation (5%) of PDJ (treated plants), and 24 plants in three different plots were treated with water (control plants) until November 29 every week. Throughout the observation period, the number of aphids, leaf-mining fly larvae, vegetable weevils, and thrips was significantly lower on the treated plants than on the control plants. In contrast, the number of lepidopteran larvae was not significantly different between the treated and control plants throughout the study period. Parasitized aphids (mummies) were also observed in both plots. Poisson regression analyses showed that a significantly higher number of mummies was recorded on the treated plants as compared to that on the control plants when the number of aphids increased. This suggested that PDJ application to Japanese radish plants attracted more parasitoid wasps on the treated plants than on the control plants. We also identified eight terpenoids and methyl salicylate as the PDJ-induced plant volatiles in the headspace of the treated plants. Some of these volatiles might be responsible for attracting aphid-parasitoid wasps in the field. However, for other insect pests, we did not find any natural enemies. Interestingly, the genes of the JA and salicylic acid signaling pathways were differentially upregulated in the treated plants. We also observed that the PDJ treatments induced the expression of the genes related to glucosinolate biosynthesis and the subsequent isothiocyanate formation. Additionally, the weights of both the aboveground and belowground parts of the treated plants were significantly lower than those of the respective parts of the control plants. These results indicated that the treatment of Japanese radish plants with a 100 times-diluted commercial formulation of PDJ induced their direct and indirect defenses against several insect pest species to reduce their numbers, and negatively affected their biomass.

Effect of soaking and heating stimulation on the formation of isothiocyanates in Eruca sativa Mill. seeds and sprouts

The present work investigated the effects of soaking followed by heating on the metabolism of glucosinolates (GLs) in rocket seeds and sprouts. Soaking for 1 h increased myrosinases (MYR) activity of seeds. Heating at 70°C for 10 min after soaking resulted in the highest isothiocyanates (ITCs) formation in seeds. Both immerse-heating and steam-heating decreased the total GLs content in seeds and 2-day old sprouts. However, steam-heating showed minor effect on total GLs content when compared with immerse-heating. Both methods showed a decreased MYR activity in seeds and sprouts. Nevertheless, heating had a positive impact on ITCs formation. Under immerse-heating, ITCs formation was the highest in seeds and sprouts at 70°C heating. However, under steam-heating, ITCs formation in seeds and sprouts was higher at 70 and 60°C, respectively. The results indicated that steam-heating treatment was effective for improving ITCs formation in rocket seeds and 2-day old sprouts.

Multiresponse kinetic modelling of the formation, release, and degradation of allyl isothiocyanate from ground mustard seeds to improve active packaging

This study aims to describe allyl isothiocyanates (AITC) formation from enzymatic sinigrin hydrolysis in ground mustard seeds and its release and degradation in the headspace using multiresponse kinetics modelling. The mechanistic modelling of the steps involved in the packaging system consists of a set of ordinary differential equations established from bio (chemical) reaction models combined with mass transfer models. The estimated parameters consist of the accessible sinigrin fraction, rate constants of sinigrin hydrolysis, AITC degradation in the particles and headspace, and its mass transfer coefficient. The model provides a good fit to experimental results and confirms the proposed mechanism of the AITC formation, degradation, and release inside the packaging system. Fat content has significant effects on AITC formation and release rate constants, while particle sizes significantly affect accessible sinigrin in the particles. These results give an understanding of AITC's controlled release by manipulating the mustard properties to optimize antimicrobial packaging designs. • A multiresponse model can describe AITC formation, release, and degradation. • Fat content has significant effects on rate constants of AITC formation and release. • Particle sizes have significant effects on accessible sinigrin in the particles. • AITC release can be controlled by manipulating the mustard-seeds properties. • The developed model can be used to optimize the design of antimicrobial packaging.

Domestic boiling and salad preparation habits affect glucosinolate degradation in red cabbage (Brassica oleracea var. capitata f. rubra)

Red cabbage contains glucosinolates, precursors to health-promoting isothiocyanates. However, raw cabbage often releases mainly epithionitriles and nitriles from glucosinolates. To increase isothiocyanate formation, the effect of acid usage in the preparation of red cabbage was evaluated. Moreover, the effects of the chosen boiling method (acidic boiled red cabbage versus neutral boiled blue cabbage) on glucosinolate degradation were investigated using UHPLC-DAD-ToF-MS and GC–MS. The addition of vinegar significantly increased isothiocyanate formation of cabbage salad from 0.09 to 0.21 µmol/g fresh weight, while lemon juice only slightly increased isothiocyanate formation. Acidic boiled red cabbage degraded glucosinolates and increased nitrile formation, while in neutral boiled blue cabbage, glucosinolates were stable. However, shortly boiled blue cabbage (5 min) had the highest isothiocyanate levels (0.08 µmol/g fresh weight). Thus, for a diet rich in isothiocyanates it is recommended to acidify raw cabbage salads and prepare shortly boiled blue cabbage instead of red cabbage.

A Metabolic Pathway for Activation of Dietary Glucosinolates by a Human Gut Symbiont

Consumption of glucosinolates, pro-drug-like metabolites abundant in Brassica vegetables, has been associated with decreased risk of certain cancers. Gut microbiota have the ability to metabolize glucosinolates, generating chemopreventive isothiocyanates. Here, we identify a genetic and biochemical basis for activation of glucosinolates to isothiocyanates by Bacteroides thetaiotaomicron, a prominent gut commensal species. Using a genome-wide transposon insertion screen, we identified an operon required for glucosinolate metabolism in B.thetaiotaomicron. Expression of BT2159-BT2156 in a non-metabolizing relative, Bacteroides fragilis, resulted in gain of glucosinolate metabolism. We show that isothiocyanate formation requires the action of BT2158 and either BT2156 or BT2157 invitro. Monocolonization of mice with mutant BtΔ2157 showed reduced isothiocyanate production in the gastrointestinal tract. These data provide insight into the mechanisms by which a common gut bacterium processes an important dietary nutrient.

Formulation of Broccoli Sprout Powder in Gastro-Resistant Capsules Protects against the Acidic pH of the Stomach In Vitro but Does Not Increase Isothiocyanate Bioavailability In Vivo.

Broccoli sprout powder is a rich source of glucosinolates, which are hydrolysed to isothiocyanates in the presence of the enzyme myrosinase. We showed that in vitro incubation of broccoli sprout powder extract with isolated lymphocytes resulted in the upregulation of transcription factor Nrf2, however, there was no increase in Nrf2 protein levels in lymphocytes isolated 3 h following the ingestion of broccoli sprout powder by healthy volunteers. This highlights the general issue that potential health benefits of food-derived compounds can be compromised by limitations in bioavailability. In vitro experiments showed that the generation of isothiocyanates was reduced when the powder was first exposed to the low pH (1.2) of the stomach and then transferred to the higher pH (6.8) of the intestine. The loss of activity due to pre-exposure to the low stomach pH indicates that formulating the broccoli sprout powder in gastro-resistant formulations should increase that amount of isothiocyanate generated in the intestine for absorption. Gelatin capsules were hand-coated with either Eudragit® L100 or Eudragit® L100-55 and were assessed for their gastro-resistant properties using paracetamol as a model active for dissolution studies. Disintegration and dissolution studies showed that Eudragit® L100-55 coated capsules and DRcapsTM (Capsugel®) failed the United States Pharmacopeia (USP) requirements for gastro-resistant capsules, whereas the Eudragit® L100 coated capsules passed. Five healthy participants were administered 1 g of broccoli sprout powder, ingested either with water or encapsulated in uncoated or gastro-resistant capsules. Urinary excretion of isothiocyanate metabolites over the 24 h period post ingestion was assessed by HPLC. Broccoli sprout powder and uncoated gelatin-encapsulated powder showed comparable excretion of isothiocyanate metabolites (18.4 ± 2.3 and 23.9 ± 2.7 µmol, respectively). The enteric coated capsules provided a significantly longer Tmax than the uncoated gelatin capsules (15.4 ± 2.3 versus 3.7 ± 0.7 h, respectively), indicating protection from disintegration in the stomach, however, the excretion of isothiocyanate metabolites was significantly decreased compared with uncoated capsules (i.e., 8.5 ± 1.1 µmol). The lower in vivo formation or absorption of isothiocyanates observed for the gastro-resistant capsules may be due to participant variation in intestinal pH or transit times, resulting in inappropriate pH conditions or insufficient time for the complete disintegration and dissolution of the capsules.