Khem B. Adhikari

Benzoxazinoids: Cereal phytochemicals with putative therapeutic and health-protecting properties.

Benzoxazinoids (BXs) are a group of natural chemical compounds with putative pharmacological and health-protecting properties. BXs were formerly identified in and isolated from selected dicot medicinal plants and young cereal plants. Recently, BXs were found to be present in mature cereal grains and bakery products, such that knowledge about the pharmacological properties of BXs, which until now have unknowingly been consumed through the daily bread and breakfast cereals, has come into new focus. This review discusses published results from in vitro studies and a few human and animal model studies on the health effects and pharmacological responses of various BX compounds. Many of these studies have reported antimicrobial, anticancer, reproductive system stimulatory, central nervous system stimulatory, immunoregulatory, and appetite- and weight-reducing effects of BXs and/or BX derivatives. The health benefits of wholegrain intake may be associated with the solitary and/or overlapping biological effects of fibers, lignans, phenolic acids, alkylresorcinols, BXs, and other bioactive compounds. In the context of BXs as dietary ingredients, further comprehensive investigations are required to understand their biological functions, to elucidate the underlying mechanisms, to explore their potential contribution on the health effects associated with wholegrain consumption, and to examine their potential as functional food ingredients.

Absorption and metabolic fate of bioactive dietary benzoxazinoids in humans

SCOPE Benzoxazinoids, which are natural compounds recently identified in mature whole grain cereals and bakery products, have been suggested to have a range of pharmacological properties and health-protecting effects. There are no published reports concerned with the absorption and metabolism of bioactive benzoxazinoids in humans. METHODS AND RESULTS The absorption, metabolism, and excretion of ten different dietary benzoxazinoids were examined by LC-MS/MS by analyzing plasma and urine from 20 healthy human volunteers after daily intake of 143 μmol of total benzoxazinoids from rye bread and rye buns. The results showed that 2-β-D-glucopyranosyloxy-1,4-benzoxazin-3-one (HBOA-Glc) and its oxidized analog, 2-β-D-glucopyranosyloxy-4-hydroxy-1,4-benzoxazin-3-one (DIBOA-Glc), were the major circulating benzoxazinoids. After consuming a benzoxazinoid diet for 1 week, morning urine contained eight benzoxazinoids with abundant HBOA-Glc (219 nmol × μmol⁻¹ of creatinine). The sulfate and glucuronide conjugates of 2-hydroxy-1,4-benzoxazin-3-one (HBOA) and 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) were detected in plasma and urine, indicating substantial phase II metabolism. Direct absorption of lactam glycosides, the reduction of hydroxamic acid glycosides, glucuronidation, and sulfation were the main mechanisms of the absorption and metabolism of benzoxazinoids. CONCLUSION These results indicate that following ingestion in healthy humans, a range of unmetabolized bioactive dietary benzoxazinoids and their sulfate and glucuronide conjugates appear in circulation and urine.

Plasma and urine concentrations of bioactive dietary benzoxazinoids and their glucuronidated conjugates in rats fed a rye bread-based diet.

Thorough knowledge of the absorption and metabolism of dietary benzoxazinoids is needed to understand their health-promoting effects. In this study, the fates of these bioactive compounds were examined by LC-MS/MS in plasma, urine, and feces after ingesting a daily dose of 4780 ± 68 nmol benzoxazinoids from rye bread using Wistar rats as a model. HBOA-glc (2-β-D-glucopyranosyloxy-1,4-benzoxazin-3-one) was the predominant benzoxazinoid in the plasma (74 ± 27 nmol/L), followed by DIBOA-glc (2-β-D-glucopyranosyloxy-4-hydroxy-1,4-benzoxazin-3-one) and HBOA. The total level of benzoxazinoids in the urine was 1176 ± 66 nmol/d, which corresponds to approximately 25% of the total dietary intake. The urinary benzoxazinoid profile differed from that of plasma with HBOA-glc and DIBOA-glc (647 ± 31 and 466 ± 33 nmol/d, respectively) as the major urinary components. The glucuronide conjugates of HBOA and DIBOA were detected in both the plasma and urine. N-dehydroxylation was found to be a critical step in the absorption of hydroxamic acids. This unprecedented study will trigger future interest in the biological effects of benzoxazinoids in whole grain rye and wheat diets in humans and other animals.

Identification and Quantification of Loline-Type Alkaloids in Endophyte-Infected Grasses by LC-MS/MS

Lolines, fungal metabolites of the grass-endophyte association, were identified and quantified using newly developed LC-MS/MS methods in endophyte-infected grasses belonging to the Lolium and Festuca genera after extraction with three different solvents using two extraction methods. The shaking extraction method with isopropanol/water was superior to the other methods due to its high sensitivity, high accuracy (recovery within or close to the range of 80-120%), and high precision (coefficient of variation of <10%). Seven loline alkaloids were identified and quantified using our newly established LC-MS/MS methods, and N-formylloline was the most abundant (5 mg/g dry matter), followed by N-acetylloline. These LC-MS/MS methods used the shortest sample handling time and the fewest sample preparation steps and proved to be good alternatives to existing GC and GC-MS analytical methods without compromising analytical efficiency. In conclusion, we developed for the first time a highly sensitive quantitative LC-MS/MS analytical method for the accurate and reproducible quantification and a LightSight-assisted LC-QTRAP/MS qualitative method for the tentative identification of loline-type alkaloids in endophyte-infected grasses.

Dietary exposure to benzoxazinoids enhances bacteria-induced monokine responses by peripheral blood mononuclear cells

SCOPE To examine potentially immunomodulating effects of dietary benzoxazinoids (BXs), present in cereal grains. METHODS AND RESULTS Nineteen healthy volunteers were randomly distributed into two groups, who received diets with high or low content of BXs for 3 wk. After a weeks wash-out, the groups switched diets. Peripheral blood mononuclear cells (PBMCs) were stimulated with Porphyromonas gingivalis, Escherichia coli lipopolysaccharide (LPS), or tetanus toxoid (TT). PBMCs from a healthy donor received the same stimuli in presence of serum from each participant receiving BXs. The production of monokines, T-cell cytokines and T-helper cell proliferation were assessed. A 3-wk diet with high BX content enhanced IL-1β responses against LPS and P. gingivalis, as well as TNF-α response against P. gingivalis, after 24 h of stimulation. Moreover, IL-6 was found to be increased after 7 days of stimulation with LPS. No effect was observed on T-cell cytokines or proliferation. BX levels in serum after a single meal did not modify cytokine responses. CONCLUSION High dietary intake of BXs enhances bacteria-induced production of pro-inflammatory monokines by PBMCs, but not T-cell responses; presumably due to intrinsic changes within PBMCs, built up over 3 wk of BX-rich diet, rather than to an immediate effects of BXs contained in serum.