Monodopsis subterranea is a source of α-tocomonoenol, and its concentration, in contrast to α-tocopherol, is not affected by nitrogen depletion.

α-Tomonoenols (αT1) are tocochromanols structurally related to tocopherols (T) and tocotrienols (T3), the bioactive members of the vitamin E family. However, limited evidence exists regarding the sources and biosynthesis of tocomonoenols. Nitrogen depletion increases the content of α-tocopherol (αT), the main vitamin E congener, in microalgae, but little is known regarding its effect on other tocochromanols, such as tocomonoenols and tocotrienols. We therefore quantified the concentrations of T, T1, and T3, in freeze-dried biomass from nitrogen-sufficient, and nitrogen-depleted Monodopsis subterranea (Eustigmatophyceae). The identities of isomers of αT1 were confirmed by LC-MS and GC-MS. αT was the predominant tocochromanol (82% of total tocochromanols). αT1 was present in higher quantities than the sum of all T3 (6% vs. 1% of total tocochromanols). 11'-αT1 was the main αT1 isomer. Nitrogen depletion increased αT, but not αT1 or T3 in M. subterranea. In conclusion, nitrogen depletion increased the content of αT, the biologically most active form of vitamin E, in M. subterranea without affecting αT1 and T3 and could potentially be used as a strategy to enhance its nutritional value but not to increase αT1 content, indicating that αT1 accumulation is independent of that of αT in microalgae.

Omega-3 polyunsaturated fatty acids improve intestinal barrier integrity-albeit to a lesser degree than short-chain fatty acids: an exploratory analysis of the randomized controlled LIBRE trial.

PURPOSE: Adherence to the Mediterranean diet is associated with beneficial health effects, including gastrointestinal disorders. Preclinical studies suggest that omega-3 polyunsaturated fatty acids (n-3 PUFAs), found in Mediterranean foods like nuts and fish, improve intestinal barrier integrity. Here, we assessed possible effects of n-3 PUFAs on barrier integrity in a randomized controlled trial. METHODS: We studied 68 women from the open-label LIBRE trial (clinicaltrials.gov: NCT02087592) who followed either a Mediterranean diet (intervention group, IG) or a standard diet (control group, CG). Study visits comprised baseline, month 3, and month 12. Barrier integrity was assessed by plasma lipopolysaccharide binding protein (LBP) and fecal zonulin; fatty acids by gas chromatography with mass spectrometry. Median and interquartile ranges are shown. RESULTS: Adherence to the Mediterranean diet increased the proportion of the n-3 docosahexaenoic acid (DHA) (IG + 1.5% [0.9;2.5, p < 0.001]/ + 0.3% [- 0.1;0.9, p < 0.050] after 3/12 months; CG + 0.9% [0.5;1.6, p < 0.001]/ ± 0%) and decreased plasma LBP (IG - 0.3 µg/ml [- 0.6;0.1, p < 0.010]/ - 0.3 µg/ml [- 1.1; - 0.1, p < 0.001]; CG - 0.2 µg/ml [- 0.8; - 0.1, p < 0.001]/ ± 0 µg/ml) and fecal zonulin levels (IG - 76 ng/mg [- 164; - 12, p < 0.010]/ - 74 ng/mg [- 197;15, p < 0.001]; CG - 59 ng/mg [- 186;15, p < 0.050]/ + 10 ng/mg [- 117;24, p > 0.050]). Plasma DHA and LBP (R2: 0.14-0.42; all p < 0.070), as well as plasma DHA and fecal zonulin (R2: 0.18-0.48; all p < 0.050) were found to be inversely associated in bi- and multivariate analyses. Further multivariate analyses showed that the effect of DHA on barrier integrity was less pronounced than the effect of fecal short-chain fatty acids on barrier integrity. CONCLUSIONS: Our data show that n-3 PUFAs can improve intestinal barrier integrity. TRIAL REGISTRATION NUMBER: The trial was registered prospectively at ClinicalTrials.gov (reference: NCT02087592).

Potential Modulation of Inflammation and Physical Function by Combined Probiotics, Omega-3 Supplementation and Vitamin D Supplementation in Overweight/Obese Patients with Chronic Low-Grade Inflammation: A Randomized, Placebo-Controlled Trial.

Obesity is characterized by low-grade inflammation and increased gut permeability. Here, we aim to evaluate the effect of a nutritional supplement on these parameters in subjects with overweight and obesity. A double-blinded, randomized clinical trial was conducted in 76 adults with overweight or obesity (BMI 28 to 40) and low-grade inflammation (high-sensitivity C-reactive protein (hs-CRP) between 2 and 10 mg/L). The intervention consisted of a daily intake of a multi-strain probiotic of Lactobacillus and Bifidobacterium, 640 mg of omega-3 fatty acids (n-3 FAs), and 200 IU of vitamin D (n = 37) or placebo (n = 39), administered for 8 weeks. hs-CRP levels did not change post-intervention, other than an unexpected slight increase observed in the treatment group. Interleukin (IL)-6 levels decreased in the treatment group (p = 0.018). The plasma fatty acid (FA) levels of the arachidonic acid (AA)/eicosapentaenoic acid (EPA) ratio and n-6/n-3 ratio (p < 0.001) decreased, and physical function and mobility improved in the treatment group (p = 0.006). The results suggest that hs-CRP may not be the most useful inflammatory marker, but probiotics, n-3 FAs, and vitamin D, as non-pharmaceutical supplements, may exert modest effects on inflammation, plasma FA levels, and physical function in patients with overweight and obesity and associated low-grade inflammation.

Potentially Beneficial Effects on Healthy Aging by Supplementation of the EPA-Rich Microalgae Phaeodactylum tricornutum or Its Supernatant-A Randomized Controlled Pilot Trial in Elderly Individuals.

Dietary supplements that promote healthy aging are mostly warranted in an aging society. Because of age-related risks, anti-inflammatory and anti-oxidative agents such as microalgae are potential candidates for intervention. In a randomized controlled trial, we tested Phaeodactylum tricornutum (PT), a microalgae rich in eicosapentaenoic acid (EPA), carotenoids, vitamins, and ß-glucans, cultured in bioreactors. In this pilot trial, 19 healthy elderly received supplements for two weeks based on either the whole PT (A), the ß-1,3-glucan-rich PT supernatant (SupB), the combination thereof (A+SupB), or a Comparator product (Comp). The primary outcome variable plasma interleukin-6 was reduced after treatment with A+SupB compared to the Comp group (p = 0.04). The mobility parameters 5 s sit-to-stand test (p = 0.04 in the A group) and by trend gait speed (p = 0.08 in the A+SupB diet) were improved compared to Comp. No treatment effects were observed for fatty acids, compared to Comp but omega-6 to -3 fatty acid ratio (p = 0.006) and arachidonic acid/EPA ratio (p = 0.006) were reduced within group A+SupB. Further, the SupB study product reduced faecal zonulin (p = 0.03) compared to the Comp. The data revealed an anti-inflammatory and potentially anti-oxidative effect of particular PT preparations, suggesting that they might be suitable for effects in healthy elderly.

Tocochromanol Profiles in Chlorella sorokiniana, Nannochloropsis limnetica and Tetraselmis suecica Confirm the Presence of 11'-α-Tocomonoenol in Cultured Microalgae Independently of Species and Origin.

11'-α-Tocomonoenol (11'-αT1) is structurally related to vitamin E and has been quantified in the microalgae Tetraselmis sp. and Nannochloropsis oceanica. However, it is not known whether 11'-αT1 is present in other microalgae independent of species and origin. The aim of this study was to analyze the tocochromanol profiles of Chlorella sorokiniana, Nannochloropsis limnetica, and Tetraselmis suecica and to determine if 11'-αT1 is present in these microalgae. Cultured microalgae were freeze-dried and the presence and identity of α-tocomonoenols were confirmed by LC-MSn (liquid chromatography coupled to mass spectroscopy) and GC-MS (gas chromatography coupled to mass spectroscopy). Tocochromanol profiles were determined by HPLC-FLD (liquid chromatography with fluorescence detection) and fatty acid profiles (as fatty acid methyl esters; FAME) by GC-MS. As confirmed by LC-MSn and GC-MS, 11'-αT1 was the dominant αT1 isomer in cultured microalgae instead of 12'-αT1, the isomer also known as marine-derived tocopherol. αT1 represented less than 1% of total tocochromanols in all analyzed samples and tended to be more abundant in microalgae with higher proportions of polyunsaturated fatty acids. In conclusion, our findings confirm that αT1 is not restricted to terrestrial photosynthetic organisms, but can also accumulate in microalgae of different species, with 11'-αT1-and not the marine-derived tocopherol (12'-αT1)-as the predominant αT1 isomer.

Effect of two postharvest technologies on the micronutrient profile of cashew kernels from Mozambique.

The economics involved in processing cashew nuts (Anacardium occidentale) might alter micronutrient profiles and concentrations. We analyzed and evaluated carotenoids, tocopherols, tocotrienols, minerals, fatty acids, and amino acids in (1) cashew kernels with testa recovered from nuts dried with and without the apple, and (2) testa-free industrial grade baby butts, splits, and white whole kernels using HPLC, ICP-OES, and GC-MS techniques. The results indicated that drying cashews with the respective apple slightly decreased the concentration of some carotenoids and total fatty and amino acids, but increased the concentration of iron, magnesium, and total tocotrienols compared with the conventionally (sun-) dried kernels. We also found high concentrations of carotenoids in the testa-containing kernels. Among the industrially processed kernel, baby butt grade was associated with lower content of ß-carotene, total tocopherols, and tocotrienols, but with significantly higher concentrations in minerals, fatty acids, and amino acids than in white wholes and split grades. Conventional sun drying of cashew nuts revealed results similar to drying with apples regarding micronutrient concentrations. The high micronutrient content of industrial grade BB is reflected in widespread human consumption and better market value.

Oral Bioavailability of Omega-3 Fatty Acids and Carotenoids from the Microalgae Phaeodactylum tricornutum in Healthy Young Adults.

The microalgae Phaeodactylum tricornutum (PT) contains valuable nutrients such as proteins, polyunsaturated omega-3 fatty acids (n-3 PUFA), particularly eicosapentaenoic acid (EPA) and some docosahexaenoic acid (DHA), carotenoids such as fucoxanthin (FX), and beta-glucans, which may confer health benefits. In a randomized intervention trial involving 22 healthy individuals, we administered for two weeks in a crossover manner the whole biomass of PT (5.3 g/day), or fish oil (FO) containing equal amounts of EPA and DHA (together 300 mg/day). In an additional experiment, sea fish at 185 g/week resulting in a similar EPA and DHA intake was administered in nine individuals. We determined the bioavailability of fatty acids and carotenoids and assessed safety parameters. The intake of PT resulted in a similar increase in the n-3 PUFA and EPA content and a decrease in the PUFA n-6:n-3 ratio in plasma. PT intake caused an uptake of FX that is metabolized to fucoxanthinol (FXOH) and amarouciaxanthin A (AxA). No relevant adverse effects occurred following PT consumption. The study shows that PT is a safe and effective source of EPA and FX-and likely other nutrients-and therefore should be considered as a future sustainable food item.

Fatty acid profiles in erythrocyte membranes following the Mediterranean diet - data from a multicenter lifestyle intervention study in women with hereditary breast cancer (LIBRE).

BACKGROUND & AIMS: Evidence-based concepts to prevent breast cancer in women with BRCA1/2 mutations are limited. Adherence to a Mediterranean diet (MedD) has been associated with a lower risk for breast cancer, possibly due to a favorable fatty acid (FA) intake. Here, we studied in an at-risk population the effect of a lifestyle intervention that included the MedD on FA composition in red blood cell membranes (RBCM). METHODS: Data derived from the German multicenter trial LIBRE, from which 68 women were randomized into an intervention group (IG) trained for MedD and increased physical activity for 12 months, and a usual care control group (CG). Adherence to the diet was assessed after 3 and 12 months using the validated Mediterranean Diet Adherence Screener (MEDAS) and a food frequency questionnaire. RBCM FA were analyzed by gas chromatography with mass spectrometry. RESULTS: The MEDAS was increased in both groups after 3 months (IG: P < 0.001; CG: P = 0.004), and remained increased only in the IG after 12 months (P < 0.001). The food frequency questionnaire revealed an increased intake of omega-3 (n-3) FA at month 3 and month 12 in the IG (both P < 0.01), but not in the CG, in which intake of energy, protein and saturated FA decreased. In both groups n-6 FA in the RBCM decreased (P < 0.001), while n-9 FA increased (P < 0.001) and n-3 FA were unchanged. Women with higher consumption of fish had higher amounts of n-3 fatty acids in the RBCM. The MEDAS was inversely correlated with n-6 fatty acids. CONCLUSIONS: The RBCM FA composition was associated with dietetic parameters related to the MedD. Adherence to the MedD resulted in an altered, likely favorable FA composition. Our data suggest selected FA as biomarkers to monitor compliance to a dietetic intervention such as the MedD. CLINICAL TRIAL REGISTRY: The trial is registered at ClinicalTrials.gov (reference: NCT02087592).

The Furan Fatty Acid 9M5 Acts as a Partial Ligand to Peroxisome Proliferator-Activated Receptor gamma and Enhances Adipogenesis in 3T3-L1 Preadipocytes.

A food that has been praised for its beneficial effects on overall health is fish, particularly its polyunsaturated n-3 fatty acids, including docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). However, it has recently been suggested that minor fatty acids such as furan fatty acids are needed in combination with DHA and EPA to exert these positive effects of fish and fish oils. Only recently have furan fatty acids become available in quantities that allow the investigation of their biofunctional properties. In this study, the uptake and effect of the furan fatty acid 9-(3-methyl-5-pentylfuran-2-yl)-nonanoic acid (9M5) as a sole component and in combination with DHA and EPA on adipogenesis were analyzed using the 3T3-L1 cell model. 9M5 is taken up and metabolized into 7M5, 5M5, and 3M5 in 3T3-L1 adipocytes during a 24-h period as shown with gas chromatography with mass spectrometry (GC/MS). Furthermore, 9M5 significantly increased lipid accumulation during the differentiation process of 3T3-L1 preadipocytes into adipocytes. In addition, the combinations of DHA + 9M5 and EPA + DHA + 9M5 also exerted a significant increase compared to control adipocytes. 3T3-L1 cells incubated with 9M5 resulted in an increased protein expression of PPARγ, C/EBPα, FABP4, and adiponectin, although not to the extent that DHA as a sole component or DHA + 9M5 did. Earlier studies have shown that DHA is a natural ligand for PPARγ, thus being a potential alternative to the antidiabetic thiazolidinediones. We show that 9M5 activates a PPARγ-responsive reporter gene and could therefore be a natural ligand for PPARγ.

Quantification of bromophenols in Islay whiskies.

Two single malt whiskies from the Scottish island Islay, i.e., Laphroiag and Lagavulin, are characterized by an iodine-like flavor associated with marine environments. In this study we investigated if this flavor impression could be due to bromophenols which are character impact compounds of marine fish and shrimps. In this study we developed a method suited for the determination of dibromo- and tribromophenols in whisky. Aliquots were O-acetylated, and quantification was carried out with gas chromatography with electron-capture negative ion mass spectrometry (GC/ECNI-MS). Both Islay whiskies contained more than 400 ng/L bromophenols with 2,6-dibromophenol being the most relevant homologue (>300 ng/L, respectively). These concentrations are at least 1 order of magnitude higher than the taste threshold of 2,6-dibromophenol in water. A third Islay whisky, Bowmore, contained ∼100 ng/L bromophenols while seventeen other whiskies from other regions in Scotland as well as from the USA, Ireland, and Germany contained at least 1 order of magnitude less than the two whiskies with the marine taste. Accordingly, bromophenols may contribute to the marine flavor and taste of Laphroaig and Lagavulin.

Mass spectra of methyl esters of brominated fatty acids and their presence in soft drinks and cocktail syrups.

RATIONALE: Brominated vegetable oil (BVO) is frequently used as a solubility transmitter in soft drinks. Being banned in Europe and Japan but permitted in the United States and Canada, there is a need for analytical methods suitable for use in food control. Brominated fatty acids in BVO are usually determined by gas chromatography (GC) after their conversion into the corresponding methyl esters. METHODS: GC with mass spectrometry (MS) was used to record the electron ionization (EI) and negative ion chemical ionization (NICI) mass spectra of relevant brominated fatty acid methyl esters synthesized for this purpose. Brominated fatty acids obtained from transesterified BVO from soft drink and syrup samples were also analyzed. RESULTS: GC/NICI-MS was the most sensitive method for the detection of brominated fatty acids but GC/EI-MS was found to be more suited for quantification due to the formation of more selective fragment ions in the higher mass range. Suitable ions were selected for determination of the methyl esters of brominated fatty acids in the selected ion monitoring (SIM) mode. Artifacts produced by the transesterification of BVO with boron trifluoride were observed and discussed. BVO was also quantified in three syrup samples commercially produced for use in cocktails/long drinks. In one of the syrup samples that tested positive, BVO was not labelled in the ingredient list. Bromination experiments produced evidence that one or more Br2 -18:0 isomers identified as a shoulder peak of threo-9,10-dibromooctadecanoic acid in several soft drink and syrup samples originated from the bromination of partly hydrogenated plant oil. CONCLUSIONS: BVO was determined for the first time in syrup samples. Attention should be paid to the problem of BVO occurring unlabeled in soft drinks and cocktail syrups imported from North America.

Differentiation of refined and virgin edible oils by means of the trans- and cis-phytol isomer distribution.

The differentiation of nonrefined (e.g., cold-pressed) and refined edible oils is an important task in food control because of the higher commercial value of the former. Here, we explored the suitability of the relative abundance of cis-phytol as a marker for authentication of nonrefined edible oils. Phytol, the tetramethyl-branched, monoenoic alcohol, is found widespread in nature as a part of chlorophyll. In chlorophyll, only trans-phytol is found. In this study, we present a method for the analysis of the phytol isomers, considering that traces of cis-phytol (contributing 0.1% to the phytol content) can be determined next to trans-phytol. For this purpose, phytol was gathered with the unsaponifiable matter from the oil, trimethylsilylated, and analyzed by gas chromatography coupled to mass spectrometry. With this method, 27 samples of edible oils (16 refined and 11 nonrefined edible oils) were analyzed for the abundance of cis-phytol relative to trans-phytol. In the nonrefined oils (e.g., olive oil, rapeseed oil, maize oil, and sunflower oil), cis-phytol contributed 0.1% (n = 3) or less (n = 8) to the phytol content. In contrast, the refined olive oils (n = 4) contained a share of 1.3-3% cis-phytol; the refined rapeseed oil (n = 3) contained a share of 0.7-1.0% cis-phytol; and the refined sunflower oil (n = 4) contained a share of 0.3-0.9% cis-phytol. Only one refined pomegranate kernel did not contain cis-phytol. The phytol concentration was not suited to distinguish nonrefined from refined oils. In contrast, our data suggest that the virtual absence of cis-phytol can be used as a marker for nonrefined (e.g., cold-pressed) edible oils.

Thorough analysis of polyhalogenated compounds in ray liver samples off the coast of Rio de Janeiro, Brazil.

INTRODUCTION: Five liver samples of two different ray species (Gymnura altavela and Zapteryx brevirostris) off the coast of Rio de Janeiro, Brazil, were analyzed for their pollution with anthropogenic and naturally occurring organohalogen compounds. MATERIAL AND METHODS: The samples were extracted with accelerated solvent extraction, and after a clean-up procedure, organohalogen compounds were separated by a modified group separation on activated silica. Subsequent analyses were done by targeted and non-targeted gas chromatography-mass spectrometry in the electron capture negative ion mode. RESULTS AND DISCUSSION: "Classic" organohalogen compounds such as polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), and technical 1,1,1-trichloro-2,2-di(4-chlorophenyl)ethane (DDT) were detected and quantified. PCBs generally exceeded the parts per million level and represented up to 90% of the total contamination of the ray livers. High concentrations were also detected for p,p'-DDE. Non-targeted full scan investigations lead to the detection of an abundant trichlorinated compound which was identified as a new DDT metabolite in biota. Different PBDE congeners and several halogenated natural products were quantified as well. In addition, polychlorinated terphenyls were identified and analyzed in the two species. Moreover, both ray species showed different fatty acid patterns and stable carbon isotope signatures. CONCLUSIONS: The two ray species showed high concentrations of organohalogen compounds in their liver tissue. Varied δ (13)C values by up to 3.1‰ indicated that the two ray species were living in different habitats.

Identification of a fungi-derived terrestrial halogenated natural product in wild boar (Sus scrofa).

In this study, we identified and quantitated a tetrachlorinated compound found at high concentrations in some samples of the meat of free-ranging wild boar (Sus scrofa) from Southern Germany. Mass spectrometric analysis indicated that the compound was a tetrachloromethoxyphenol isomer, and the subsequently synthesized tetrachloro-p-methoxyphenol was identical with the unknown compound in wild boar. Tetrachloro-p-methoxyphenol is a known secondary metabolite of basidiomycetous fungi, which in turn are regular feed items of the wild boar. It is extremely likely that the wild boar have accumulated tetrachloro-p-methoxyphenol by exploiting basidiomycetes. The highest concentration in the samples (n = 22) was ~1 mg/kg lipids tetrachloro-p-methoxyphenol. This concentration was higher than that of polychlorinated biphenyls (PCBs) and dichlorodiphenyltrichloroethane (DDT) in any of the samples. Some samples did not contain tetrachloro-p-methoxyphenol, which indicates varied preferences in fungi by wild boars. Our data suggest that during their entire evolution, humans have been in contact with the natural product tetrachloro-p-methoxyphenol by consuming wild boars.

Establishing a chromium-reactor design for measuring delta2H values of solid polyhalogenated compounds using direct elemental analysis and stable isotope ratio mass spectrometry.

2H/1H isotope ratios of polyhalogenated compounds were determined by elemental analysis and isotope ratio mass spectrometry (EA-IRMS). Initial measurements with standard EA-IRMS equipment, which used high-temperature pyrolysis to convert the organic compounds into hydrogen, did not achieve significant signals for polychlorinated pesticides and related compounds, presumably due to the formation of HCl instead of hydrogen. To reverse this problematic reaction, a chromium reactor was incorporated into the element analyzer system, which scavenged Cl, forming chromium chloride and releasing hydrogen again in the form of H2. The optimized system therefore allowed the delta2H values of polyhalogenated compounds to be determined. A quality assurance program was developed based on several parameters. (i) Each compound was analyzed using a sequence of five injections, where the first measurement was discarded. (ii) Recovery of H (when calculated relative to acetanilide) had to be >90% for all replicates in a sequence. (iii) All delta-values within a sequence had to vary by less than 10/1000. (iv) Results had to be reproducible on another day with a different sample scheme. Once this reproducibility had been established, variabilities in the delta2H values of organohalogen standards were investigated using the technique. The highest delta2H value of +75/1000 was found for o,p'-DDD, whereas the strongest depletion in deuterium was found for Melipax (-181/1000). The most important results for comparable compounds were as follows. DDT-related compounds gave delta2H values of between +59 and +75/1000 (technical DDT, o,p'- and p,p'-DDD) or in the range of approximately -1/1000, indicative of the different sources/methods of producing this compound. Four HCH isomers from the same supplier showed relatively similar hydrogen isotope distributions, whereas two lindane (gamma-HCH) standards from other sources had 39/1000 less deuterium. This difference is likely due to different purification steps during the isolation of pure lindane from the technical HCH mixture. An even greater difference was observed between the delta2H values of Toxaphene (US product dating from 1978) and Melipax (product from the former East Germany, dating from 1979), which gave delta2H values of -101/1000 and -181/1000, respectively, meaning that both products were easily distinguished via delta2H-IRMS. Fractioning of hydrogen isotopes in the atmospheric water cycle was suggested as one reason for the different values. In this theory, the water (which had different delta2H values depending on where it was taken from) was incorporated during the biosynthesis of camphene, which is the natural product used to produce both products. These results indicate that hydrogen isotope-specific analysis can be a valuable tool for tracing the origins of a compound in certain cases.

Baseline isotopic data of polyhalogenated compounds.

The delta2H- and delta13C-values of polyhalogenated compounds were determined by EA-IRMS. Most of the compounds were related to the chloropesticides DDT and its metabolites, hexachlorocyclohexanes, and toxaphene, as well as several polybrominated compounds such as bromophenols and -anisoles. delta2H-values ranged between -235 per thousand and +75 per thousand whereas delta13C-values were found in the range -22 per thousand to -38 per thousand. No correlation between delta2H- and delta13C-values could be identified. Comparative analysis clarified that bromophenols and the corresponding bromoanisoles may vary in their isotopic distribution. 2H NMR was used to quantify abundances of 2H isotopomers. Quantification of isotopomers of 2,4-dibromophenol and 2,4-dibromoanisole proved that both compounds from different suppliers do not originate from the same source. Differences in the delta2H-values of two toxaphene products were further investigated by the synthesis of products of different degree of chlorination from camphene. It was shown that the delta13C-values remained mostly unaltered as was expected since no carbon is lost in this procedure. However, the reaction products became enriched in 2H with increasing degree of chlorination. Different delta2H-values of the starting material will also impact the delta2H-values of the chlorination products.

Enantioseparation of chiral organochlorines on permethylated beta- and gamma-cyclodextrin, as well as 1:1 mixtures of them.

Gas chromatography columns coated with 10% permethylated beta- and gamma-cyclodextrin in 85% dimethyl-15% diphenyl polysiloxane (beta- and gamma-PMCD, respectively) and 1:1 mixures are prepared and tested with regard to the enantioseparation of chiral chloropesticides. On the columns with the individual O-methylated cyclodextrins (O-tCDs), the enantiomers of aaeeee-hexachlorocyclohexane (alpha-HCH), e-aeee-1,3,4,5,6-pentachlorocyclohex-1-ene (beta-PCCH), and e-aaee-1,3,4,5,6-pentachlorocyclohex-1-ene (gamma-PCCH), cis- and trans-chlordane, and cis-heptachlor epoxide are separated on both columns, with the exception of the latter being separated only on beta-PMCD. On the column coated with 5% beta- and 5% gamma-PMCD, the resulting separation factor (a) is virtually 1/2 of the arithmetric mean of the elution-dependent separation factors on the individual O-tCDs. In case of reversed elution order on beta- and gamma-PMCD, the enantiomers are not resolved on the mixed columns as is the case with cis-chlordane. Likewise, the lower resolution of the gamma-PCCH enantiomers on the mixed columns prove the reversed elution order on beta- and gamma-PMCD without having enantioenriched standards available. On the column coated with 5% beta- and 5% gamma-PMCD, similar retention times to those observed on both 10% beta-PMCD and 10% gamma-PMCD are obtained. On the column coated with 10% beta- and 10% gamma-PMCD, significantly longer retention times are obtained compared with the columns that contain a total of 10% chiral stationary phase (CSP). This indicates that a relevant part of the interaction of the analytes with the chiral selector is non-enantioselective and, thus, only delays the elution of both enantiomers. Moreover, these non-enantioselective interactions prevent a direct comparison of CSPs with different amounts of the chiral selector. However, this is possible by using mixed phases of two CSPs with similar properties. Using this system, it is demonstrated that for the organochlorine compounds studied, no higher separation factor is observed on the mixed CSPs than on the individual O-tCD with the higher separation factor. Estimations allow a prediction that enantioseparations of organohalogen compounds can be achieved on columns coated with as little as 1% of the CSP.

HPLC isolation and NMR structure elucidation of the most prominent octabromo isomer in technical octabromo diphenyl ether.

The major octabromo isomer of technical octabromo diphenyl ether mixture (technical octaBDE) DE-79 was isolated by RP-HPLC. Three serially coupled columns (each 250 mm long) enabled a good separation of the target compound from other congeners using 100% ACN as eluent. Approximately 100 microg of the target compound was isolated with a purity of >90% and investigated by MS for confirmation of the degree of bromination. 1H-NMR and 2-D 1H-13C correlation NMR spectra unequivocally clarified that the octaBDE in question is 2,2',3,3',4,4',6,6'-octabromodiphenyl ether (BDE 197). Based on annual production rates of technical BDE products (1999/2001), approximately 380 tons of BDE 197 were annually produced which, on the long term, may enter the environment. Compared with other individual BDE congeners, BDE 197 has the seventh highest application rate. Reductive debromination of BDE 197 can lead to four hepta-, 15 hexa-, 23 penta-, and 28 tetra-BDE isomers, respectively. This variety includes all known major BDEs of environmental concern (BDE 47, 85, 99, 100, 153, 154, and 183). The identification of BDE 197 in technical octaBDE DE-79 strongly suggests that research on the environmental fate of BDEs should include this key-BDE congener.

CD38 low IgG-secreting cells are precursors of various CD38 high-expressing plasma cell populations.

Despite the important role immunoglobulin G (IgG)-secreting plasma cells play in memory immune responses, the differentiation and homeostasis of these cells are not completely understood. Here, we studied the differentiation of human IgG-secreting cells ex vivo and in vitro, identifying these cells by the cellular affinity matrix technology. Several subpopulations of IgG-secreting cells were identified among the cells isolated from tonsils and bone marrow, particularly differing in the expression levels of CD9, CD19, and CD38. CD38 low IgG-secreting cells were present exclusively in the tonsils. A major fraction of these cells appeared to be early plasma cell precursors, as upon activation of B cells in vitro, IgG secretion preceded up-regulation of CD38, and on tonsillar sections, IgG-containing, CD38 low cells with a plasmacytoid phenotype were found in follicles, where plasma cell differentiation starts. A unitary phenotype of migratory peripheral blood IgG-secreting cells suggests that all bone marrow plasma cell populations share a common precursor cell. These data are compatible with a multistep model for plasma cell differentiation and imply that a common CD38 low IgG-secreting precursor gives rise to a diverse plasma cell compartment.

Plasma cell survival is mediated by synergistic effects of cytokines and adhesion-dependent signals.

Recent results suggest that plasma cell longevity is not an intrinsic capacity, but depends on yet unknown factors produced in their environment. In this study, we show that the cytokines IL-5, IL-6, TNF-alpha, and stromal cell-derived factor-1alpha as well as signaling via CD44 support the survival of isolated bone marrow plasma cells. The cytokines IL-7 and stem cell factor, crucially important for early B cell development, do not mediate plasma cell survival, indicating that plasma cells and early B cells have different survival requirements. As shown in IL-6-deficient mice, IL-6 is required for a normal induction, but not for the maintenance of plasma cell responses in vivo, indicating that the effects of individual survival factors are redundant. Optimal survival of isolated plasma cells requires stimulation by a combination of factors acting synergistically. These results strongly support the concept that plasma cell survival depends on niches in which a combination of specific signals, including IL-5, IL-6, stromal cell-derived factor-1alpha, TNF-alpha, and ligands for CD44, provides an environment required to mediate plasma cell longevity.