Non-enzymatic degradation of aliphatic Brassicaceae isothiocyanates during aqueous heat treatment.

Glucosinolate-derived isothiocyanates are valuable for human health as they exert health promoting effects. As thermal food processing could affect their levels in a structure dependent way, the stability and reactivity of 12 Brassicaceae isothiocyanates during aqueous heating at 100 °C and pH 5-8 were investigated. The formation of their corresponding amines and N,N'-dialk(en)yl thioureas was quantified. Further, the potential to form odor active compounds was investigated by HRGC-MS-olfactometry. A strong structure-reactivity relationship was found and shorter side chains and electron withdrawing groups increase the reactivity of isothiocyanates. 3-(Methylsulfonyl)-propyl isothiocyanate was least stable. The main products are the corresponding amines (up to 69% recovery) and formation of N,N'-dialk(en)yl thioureas is only relevant at neutral to basic pH values. Apart from allyl isothiocyanate also 3-(methylthio)propyl isothiocyanate is precursor to many sulfur-containing odor active compounds. Thus, the isothiocyanate-structure affects their levels but also contributes to the flavor of boiled Brassicaceae vegetables.

Comparison of different density functional theory methods for the calculation of vibrational circular dichroism spectra.

The determination of the absolute configuration (AC) of an organic molecule is still a challenging task for which the combination of spectroscopic with quantum-mechanical methods has become a promising approach. In this study, we investigated the accuracy of DFT methods (480 overall combinations of 15 functionals, 16 basis sets, and 2 solvation models) to calculate the VCD spectra of six chiral organic molecules in order to benchmark their capability to facilitate the determination of the AC.

Glucosinolate-derived amine formation in Brassica oleracea vegetables.

Glucosinolates are precursors of bioactive and health-promoting isothiocyanates (ITCs). Upon enzymatic hydrolysis, Brassica vegetables, such as cabbage, also often yield nitriles and epithionitriles as main products next to ITCs. Here, we show that amines can be additional main enzymatic hydrolysis products of glucosinolates in Brassica vegetables. We propose that a plant endogenous ITC hydrolase (ITCase) is responsible for the enzymatic-like conversion of ITCs to amines in cabbage samples. This ITCase seems to have high activity towards alkenyl ITCs like allyl ITC and lower activity towards methylthioalkyl ITCs, and not to converting methylsulfinylalkyl ITCs like sulforaphane. In contrast, during heat treatment of homogenized cabbage material, methylsulfinylalkylamine levels increased by 400 % after 2 h of heating, which is likely due to thermal decomposition of ITCs, whereas alkenyl amine levels did not change due to heat treatment. The results show that amines from glucosinolates are part of the human diet.

Assessing Bioavailability and Bioactivity of 4-Hydroxythiazolidine-2-Thiones, Newly Discovered Glucosinolate Degradation Products Formed During Domestic Boiling of Cabbage.

Glucosinolates are plant secondary metabolites found in cruciferous vegetables (Brassicaceae) that are valued for their potential health benefits. Frequently consumed representatives of these vegetables, for example, are white or red cabbage, which are typically boiled before consumption. Recently, 3-alk(en)yl-4-hydroxythiazolidine-2-thiones were identified as a class of thermal glucosinolate degradation products that are formed during the boiling of cabbage. Since these newly discovered compounds are frequently consumed, this raises questions about their potential uptake and their possible bioactive functions. Therefore, 3-allyl-4-hydroxythiazolidine-2-thione (allyl HTT) and 4-hydroxy-3-(4-(methylsulfinyl) butyl)thiazolidine-2-thione (4-MSOB HTT) as degradation products of the respective glucosinolates sinigrin and glucoraphanin were investigated. After consumption of boiled red cabbage broth, recoveries of consumed amounts of the degradation products in urine collected for 24 h were 18 ± 5% for allyl HTT and 21 ± 4% for 4-MSOB HTT (mean ± SD, n = 3). To investigate the stability of the degradation products during uptake and to elucidate the uptake mechanism, both an in vitro stomach and an in vitro intestinal model were applied. The results indicate that the uptake of allyl HTT and 4-MSOB HTT occurs by passive diffusion. Both compounds show no acute cell toxicity, no antioxidant potential, and no change in NAD(P)H dehydrogenase quinone 1 (NQO1) activity up to 100 µM. However, inhibition of glycogen synthase kinases-3 (GSK-3) in the range of 20% for allyl HTT for the isoform GSK-3ß and 29% for 4-MSOB HTT for the isoform GSK-3α at a concentration of 100 µM was found. Neither health-promoting nor toxic effects of 3-alk(en)yl-4-hydroxythiazolidine-2-thiones were found in the four tested assays carried out in this study, which contrasts with the properties of other glucosinolate degradation products, such as isothiocyanates.

Novel transformation products from glucosinolate-derived thioglucose and isothiocyanates formed during cooking.

Glucosinolates are plant secondary metabolites occurring in Brassicaceae plants. Upon tissue disruption, these compounds can be enzymatically hydrolyzed into isothiocyanates, which are very reactive and can react with nucleophiles during thermal processes such as boiling. Here, a novel type of glucosinolate degradation product was identified resulting from the reaction of thioglucose with the isothiocyanates sulforaphane or allyl isothiocyanate during aqueous heating. The two heterocyclic compounds 4-hydroxy-3-(4-(methylsulfinyl)butyl)thiazolidine-2-thione and 3-allyl-4-hydroxythiazolidine-2-thione were isolated and their structure elucidated by NMR spectroscopy and high-resolution mass spectrometry. Based on a set of chemical experiments, a reaction mechanism was proposed. Finally, the formation of the two 3-alk(en)yl-4-hydroxythiazolidine-2-thiones was quantified in boiled cabbage samples using a standard addition method in which 92 pmol/g and 19 pmol/g fresh weight of the sulforaphane and allyl isothiocyanate derivatives were found, respectively.

Formation of volatile sulfur compounds and S-methyl-l-cysteine sulfoxide in Brassica oleracea vegetables.

Besides glucosinolates, Brassica vegetables accumulate sulfur-containing (+)-S-methyl-l-cysteine sulfoxide (SMCSO, methiin), mainly known from Allium vegetables. Such (+)-S-alk(en)yl-l-cysteine sulfoxides can degrade to volatile organosulfur compounds (VOSCs), which have been linked to health beneficial effects. In the present study, the accumulation of SMCSO and the formation of VOSCs was investigated in Brassica oleracea vegetables. SMCSO content of commercially available white and red cabbages was monitored over a three-month period and linked with the formation of VOSCs. S-Methyl methanethiosulfinate was the main VOSC released from SMCSO. Upon heating, it degraded to dimethyltrisulfide and dimethyldisulfide, which were less abundant in fresh homogenates. SMCSO made up approximately 1% of the dry matter of cabbages and the overall contents were similar in white and red cabbages (3.2-10.2 and 3.9-10.3 µmol/g fresh weight, respectively). Using proteome profiling it was shown that recovery of VOSCs correlated with abundance of two isoforms of cystine lyase.

Total Synthesis of epi-Trichosetin.

The natural 3-decalinoyltetramic acid epi-trichosetin was synthesized in ten steps starting from ( R)-(+)-citronellal using an intramolecular Diels-Alder reaction and a Lacey-Dieckmann cyclization as the key steps. The use of a 2-nitrobenzyl protecting group resulted in an efficient synthetic endgame. The natural product was obtained in 4.1% overall yield.

Crystal structure analysis of a star-shaped triazine compound: a combination of single-crystal three-dimensional electron diffraction and powder X-ray diffraction.

The solid-state structure of star-shaped 2,4,6-tris{(E)-2-[4-(dimethylamino)-phenyl]ethenyl}-1,3,5-triazine is determined from a powder sample by exploiting the respective strengths of single-crystal three-dimensional electron diffraction and powder X-ray diffraction data. The unit-cell parameters were determined from single crystal electron diffraction data. Using this information, the powder X-ray diffraction data were indexed, and the crystal structure was determined from the powder diffraction profile. The compound crystallizes in a noncentrosymmetric space group, P212121. The molecular conformation in the crystal structure was used to calculate the molecular dipole moment of 3.22 Debye, which enables the material to show nonlinear optical effects.

Phytotoxic dioxolanones are potential virulence factors in the infection process of Guignardia bidwellii.

Phytotoxic dioxolanones from Guignardia bidwellii can be described as potential virulence factors which cause the formation of lesions upon an infection by G. bidwellii. The toxin guignardic acid was found in planta of G. bidwellii-infected Vitis vinifera leaves, whereas no phytotoxic dioxolanones were detected in uninfected leaf material. Secondary metabolism analyses of further phytopathogenic fungi from the genus Guignardia led to the observation that all species investigated can produce the phytotoxins known from G. bidwellii. In addition to these studies, it was demonstrated that phenguignardic acid is biosynthetically derived from two molecules of phenylalanine and that phenylalanine is a key precursor in the biosynthesis of the two other phytotoxins - alaguignardic acid and guignardic acid.

Absolute Configuration Determination of 2,3-Dihydro-1H,5H-pyrazolo[1,2-a]pyrazoles Using Chiroptical Methods at Different Wavelengths.

A correlation between the absolute configuration and chiroptical properties of nonracemic 1,6,7-trisubstituted 2,3-dihydro-1H,5H-pyrazolo[1,2-a]pyrazoles was studied. A series of 16 novel representatives were prepared by Cu-catalyzed [3 + 2] cycloadditions of racemic (Z)-2-benzylidene-5-oxopyrazolidin-2-ium-1-ides to tert-butyl (S)-(3-oxopent-4-yn-2-yl)carbamate, and their structures were determined by NMR, VCD, ECD, and X-ray diffraction. A clear correlation between the sign of specific rotation and configuration at position C(1) allows for easy determination of the absolute configuration of 1,6,7-trisubstituted 2,3-dihydro-1H,5H-pyrazolo[1,2-a]pyrazoles by ECD and NMR. While VCD, requiring milligram quantities, allowed the determination of the correct relative and absolute configuration without additional information from other methods, the stereochemical analysis by ECD required knowledge of the relative configuration derived from NMR at a comparable computational level.

Terphenyl Derivatives from Allantophomopsis lycopodina.

Three secondary fungal metabolites 1-3 with a benzo[b]naphtho[2,1-d]furan skeleton were isolated from submerged cultures of the ascomycete Allantophomopsis lycopodina. The NMR-based structure elucidation was challenging due to a low H/C ratio of only 0.64 and 0.68, respectively. NMR measurements in two different solvents and the use of NMR experiments such as HSQC-TOCSY and LR-HSQMBC proved to be helpful in this respect. The proposed structures obtained from the comprehensive analysis of the NMR data were verified by comparison of recorded and computed NMR chemical shifts from quantum chemical calculations of several constitutional isomers and were further analyzed with the aid of the DP4 and DP4+ probabilities.

Separation, isolation and stereochemical assignment of imazalil enantiomers and their quantitation in an in vitro toxicity test.

A simple method for the separation of the enantiomers of the fungicide imazalil was developed. Racemic imazalil was separated into its enantiomers with an enantiomeric purity of 99% using HPLC-UV with an enantioselective column (permethylated cyclodextrin) operated in reversed phase mode (water with 0.2% trimethylamine and 0.08% acetic acid and methanol). The absolute configuration of the separated enantiomers was assigned and unequivocally confirmed by optical rotation as well as by vibrational circular dichroism (VCD) and electronic circular dichroism (ECD) combined with ab-initio calculations. The same enantioselective column was also used to develop an HPLC-MS/MS method for the quantification of imazalil enantiomers. The HPLC-MS/MS method reached limits of quantification (LOQs) of 0.025mg/mL with 5µL injections. This method was used to verify imazalil concentrations and enantiomeric fractions in samples from an in vitro test on effects on human steroidogenesis (H295R steroidogenesis assay). The quantification verified the stability of the enantiomers of imazalil during the in vitro tests.

Total Synthesis of (-)-Hymenosetin.

The 3-decalinoyltetramic acid (-)-hymenosetin and its N-methyl analogue were prepared in 11 and 8 steps, respectively, from (+)-citronellal using an intramolecular Diels-Alder reaction as the key step. This method represents the first example for the synthesis of a 3-decalinoyltetramic acid with a free NH moiety. The stereochemistry of the title compound, an unnatural diastereomer, and of a decalin building block was studied in detail using circular dichroism spectroscopy in the IR and UV/VIS freqeuncy range. This allowed to determine the absolute configuration of the natural product and to plan the synthetic route.

Chromatographically separable rotamers of an unhindered amide.

Surprisingly stable formamide rotamers were encountered in the tetrahydroisoquinoline and morphinan series of alkaloids. We investigated the hindered rotation around the amide bond by dynamic high-performance liquid chromatography (DHPLC) and kinetic measurements of the interconversion of the rotamers which can readily be separated by HPLC as well as TLC. The experimental results of the different methods were compared to each other as well as to results obtained by DFT calculations.

Iodocyclization of o-alkynylbenzamides revisited: formation of isobenzofuran-1(3H)-imines and 1H-isochromen-1-imines instead of lactams.

The iodocyclization of o-alkynylbenzamides with various electrophiles has been reported to yield five- or six-membered lactams by nucleophilic attack of the amide nitrogen onto the triple bond. While the formation of an isobenzofuran-1(3H)-imine with two bulky substituents under Larock conditions was initially attributed to steric hindrance, we found out that cyclization via the amide oxygen is the rule rather than the exception. Thus, the structures of the products reported in the literature need to be revised.