Chitin quantitation (as glucosamine) in food raw materials by HPLC-C18-DAD with off-line derivatization.

This HPLC method is suitable for chitin quantitation (reported as glucosamine) in food raw materials like insects (mealworm larvae, crickets), shrimps, mushrooms and fungi in a research (non-routine) laboratory using a C18 column with HPLC system <600 bar with UV detection capability (at 265 nm). To remove interferences, the sample is defatted (Soxhlet) and deproteinized (by alkali) prior to acid hydrolysis in 6 M HCl. A five-point linear calibration (5-100 µg/mL) is used. The use of fluorescence detection (λex = 260 nm, λem = 350 nm) is also possible with this method [1].•18 min HPLC run time•LOD = 0.05 µg/mL and LOQ = 5 µg/mL.

Chitin quantitation (as glucosamine) in food raw materials by spectrophotometry.

Chitin is a water insoluble nitrogen-containing polysaccharide made from N-acetyl-D-glucosamine containing ß-(1→4)-linkages. In food, chitin is considered as a source of fiber with prebiotic properties to gut microflora. Chitin content varies widely in nature from 1% (yeasts) up to 64% (butterfly cuticles) and is mostly found in filamentous or mushroom forming fungi, insects and crustaceans. This spectrophotometric method is suitable for chitin quantitation (reported as glucosamine) in food raw materials like insects (mealworm larvae, crickets), shrimps, mushrooms and fungi in a research (non-routine) laboratory. To remove interferences, the sample is defatted (Soxhlet) prior to acid hydrolysis in 6 M HCl. The color complex is developed after the addition of Katano's reagent (a mix of 0.05 mol/L sodium metasilicate, 0.6 mol/L sodium molybdate, 30% dimethyl sulfoxide and 1.42 mol/L acetic acid) at 70 °C for 30 min and measured at 750 nm against blank. A five-point linear calibration (5-100 µg/mL) is used. Limit of detection is 3 µg GLCN/mL. The correlation (R2) with an HPLC method for chitin analysis is at least 0.93.•a reliable alternative to an HPLC method•does not require expensive equipment•deproteination by alkali is not necessary for most matrices - saves about 30% of time.

The electron beam irradiation of collagen in the dry and gel states: The effect of the dose and water content from the primary to the quaternary levels.

The aim of our study was to describe the impact of collagen in the gel and dry state to various doses of electron beam radiation (1, 10 and 25 kGy) which are using for food processing and sterilization. The changes in the chemical compositions (water, amino acids, lipids, glycosaminoglycans) were analyzed and the changes in the structure (triple-helix or ß-sheet, the integrity of the collagen) were assessed. Subsequently, the impact of the applied doses on the mechanical properties, stability in the enzymatic environment, swelling and morphology were determined. The irradiated gels evinced enhanced degrees of cross-linking with only partial degradation. Nevertheless, an increase was observed in their stability manifested via a higher degree of resistance to the enzymatic environment, a reduction in swelling and, in terms of the mechanical behaviour, an approximation to the non-linear behavior of native tissues. In contrast, irradiation in the dry state exerted a somewhat negative impact on the observed properties and was manifested mainly via the scission of the collagen molecule and via a lower degree of stability in the aqueous and enzymatic environments. Neither the chemical composition nor the morphology was affected by irradiation.

Properties of Bovine Collagen as Influenced by High-Pressure Processing.

The physical properties and structure of collagen treated with high-pressure technologies have not yet been investigated in detail. The main goal of this work was to determine whether this modern gentle technology significantly changes the properties of collagen. High pressure in the range of 0-400 MPa was used, and the rheological, mechanical, thermal, and structural properties of collagen were measured. The rheological properties measured in the area of linear viscoelasticity do not statistically significantly change due to the influence of pressure or the duration of pressure exposure. In addition, the mechanical properties measured by compression between two plates are not statistically significantly influenced by pressure value or pressure hold time. The thermal properties Ton and ∆H measured by differential calorimetry depend on pressure value and pressure hold time. Results from amino acids and FTIR analyses show that exposure of collagenous gels to high pressure (400 MPa), regardless of applied time (5 and 10 min), caused only minor changes in the primary and secondary structure and preserved collagenous polymeric integrity. SEM analysis did not show changes in collagen fibril ordering orientation over longer distances after applying 400 MPa of pressure for 10 min.

Trehalose determination in Norway spruce (Picea abies) roots. Analytics matters.

We present concise results of method validation for trehalose quantitation by LC-MS/MS in spruce ectomycorrhizal roots in order to describe spruce health status, mainly in connection to contamination by a pathogenic fungus, Gemmamyces piceae. The procedure is based on Rogatsky et al. (2005) developed for human plasma. We found out that the best extraction yield was achieved with 80% methanol/water (v/v) solution and optimal extraction temperature was set between 50-60°C. In contrast to previous papers, we minimized the activity of trehalase enzyme by putting root samples into liquid N2 immediately after root excavation, followed by freeze-drying in order to stop trehalase activity. Higher content of trehalose was recorded in healthy trees, confirming the idea that ectomycorrhiza plays a significant role in plant-pathogen interactions.

Somatic Cell Count in Goat Milk: An Indirect Quality Indicator.

A high somatic cell count (SCC) impacts dairy quality to a large extent. The goal of this work was to investigate differences in goat milk composition and technological parameters according to SCC cut-off (600, 700, 800, and 1000.103/mL). Thirty-four individual milk samples of White Shorthair goats in a similar stage of lactation were investigated. The first differences in milk quality appeared already at SCC cut-off of 600.103/mL (5.58 LSCS-linear somatic cell score), yet the most striking differences were found for SCC over 1000.103/mL (6.32 LSCS), which was expressed by lowering heat stability (126 vs. 217 s, p = 0.034), increasing protein (3.41 vs. 3.04%, p = 0.009), casein (2.80 vs. 2.44%, p = 0.034) and chloride (164 vs. 147 mg/100 mL, p = 0.004) levels, as well as non-fat dry matter (8.79 vs. 8.45%, p = 0.045). It has been shown that low levels of Staphylococcus spp. bacteria (120-1600 CFU/mL) in the mammary gland correlated with decreased lactose content (4.60 vs. 4.47 g/100 g, p = 0.022). Since our results indicate that even low SCC values may significantly affect the technological properties of goat milk, SCC should therefore be routinely screened and reported to dairy manufacturers to assure the consumer of high end-product quality.

Applicability of low-flow atmospheric pressure chemical ionization and photoionization mass spectrometry with a microfabricated nebulizer for neutral lipids.

RATIONALE: Mass spectrometry with atmospheric pressure chemical ionization (APCI) or photoionization (APPI) is widely used for neutral lipids involved in many fundamental processes in living organisms. Commercial APCI and APPI sources operate at high flow rates compatible with conventional high-performance liquid chromatography (HPLC). However, lipid analysis is often limited by a small amount of sample, which requires low flow rate separations like capillary or micro-HPLC. Therefore, APCI and APPI suitable for microliter-per-minute flow rates need to be developed and applied for neutral lipids. METHODS: A micro-APCI/APPI source with a heated chip nebulizer was assembled and mounted on a Thermo ion trap instrument. The ion source operated in APCI, APPI or dual mode was optimized for low microliter-per-minute sample flow rates. The source performance was investigated for squalene, wax esters, fatty acid methyl esters, triacylglycerols, and cholesterol. RESULTS: The ion source behaved as a mass-flow-sensitive detector. Direct infusion of methyl oleate showed superior analytical figures of merit when compared with high-flow ion sources. A detection limit of 200 pmol/mL and a linear dynamic range spanning three orders of magnitude were measured for micro-APCI. The mass spectra of most lipids differed from high flow rate spectra. Unlike micro-APCI, micro-APPI spectra were complicated by odd-electron species. Dual APCI/APPI mode did not show any benefits for neutral lipids. Applications for lipid samples were demonstrated. CONCLUSIONS: Micro-APCI-MS is a useful detection technique for neutral lipids at microliter-per-minute flow rates. It offers high sensitivity and high quality of spectra in direct infusion mode and promises successful utilization in capillary and micro-HPLC applications.

Free and esterified carotenoids in pigmented wheat, tritordeum and barley grains.

Carotenoids are important phytonutrients responsible for the yellow endosperm color in cereal grains. Five carotenoids, namely lutein, zeaxanthin, antheraxanthin, α- and ß-carotene, were quantified by HPLC-DAD-MS in fourteen genotypes of wheat, barley and tritordeum harvested in Czechia in 2014 and 2015. The highest carotenoid contents were found in yellow-grained tritordeum HT 439 (12.16µg/gDW), followed by blue-grained wheat V1-131-15 (7.46µg/gDW), and yellow-grained wheat TA 4024 (7.04µg/gDW). Comparing carotenoid contents, blue varieties had lower whereas purple ones had the same or higher levels than conventional bread wheat. Lutein was the main carotenoid found in wheat and tritordeum while zeaxanthin dominated in barley. The majority of cereals contained considerable levels of esterified forms (up to 61%) of which lutein esters prevailed. It was assessed that cereal genotype determines the proportion of free and esterified forms. High temperatures and drought during the growing season promoted carotenoid biosynthesis.

Changes in anthocyanidin levels during the maturation of color-fleshed potato (Solanum tuberosum L.) tubers.

Certain potato cultivars are capable of producing anthocyanin pigments in the potato skin and flesh and those pigments have been shown, together with other phytochemicals, to promote good health. Six common anthocyanidins (cyanidin, delphinidin, petunidin, pelargonidin, malvidin and peonidin) were analyzed weekly for 15weeks in red- and purple-fleshed potato cultivars (Red Emma, Königspurpur, Valfi and Blaue de la Mancha) grown in field conditions using a validated LC-(+ESI)MS/MS method. Pelargonidin was the major type detected in red-fleshed cultivars whereas petunidin was the major type detected in the purple ones. Neither cyanidin nor delphinidin were found in any of the cultivars. The anthocyanidin levels observed were as high as 78mg/100g FW during tuber growth; however, fully matured tubers contained only 10-39mg anthocyanidins/100gFW. Anthocyanidin levels were moderately correlated with global solar irradiation (r<0.6252) but not with rainfall or daily temperature.

Bisphenol S negatively affects the meotic maturation of pig oocytes.

Bisphenol A (BPA), a chemical component of plastics, is a widely distributed environmental pollutant and contaminant of water, air, and food that negatively impacts human health. Concerns regarding BPA have led to the use of BPA-free alternatives, one of which is bisphenol S (BPS). However, the effects of BPS are not well characterized, and its specific effects on reproduction and fertility remain unknown. It is therefore necessary to evaluate any effects of BPS on mammalian oocytes. The present study is the first to demonstrate the markedly negative effects of BPS on pig oocyte maturation in vitro, even at doses lower than those humans are exposed to in the environment. Our results demonstrate (1) an effect of BPS on the course of the meiotic cell cycle; (2) the failure of tubulin fibre formation, which controls proper chromosome movement; (3) changes in the supply of maternal mRNA; (4) changes in the protein amounts and distribution of oestrogen receptors α and ß and of aromatase; and (5) disrupted cumulus cell expansion. Thus, these results confirm that BPS is an example of regrettable substitution because this substance exerts similar or even worse negative effects than those of the material it replaced.

Carotenoid profile and retention in yellow-, purple- and red-fleshed potatoes after thermal processing.

This research aimed to investigate the effect of thermal processing on carotenoid profile, quantity and stability in 22 colour-fleshed potato cultivars grown in the Czech Republic. The total of nine carotenoids was analysed by HPLC using a C30 column and PDA detection. The total carotenoid content for all cultivars ranged from 1.44 to 40.13 µg/g DM. Yellow cultivars showed a much higher average total carotenoid content (26.22 µg/g DM) when compared to red/purple-fleshed potatoes (5.69 µg/g DM). Yellow cultivars were dominated by antheraxanthin, whereas neoxanthin was the main carotenoid in red/purple cultivars. Thermal processing significantly impacted all potato cultivars. Boiling decreased the total carotenoids by 92% compared to baking (88%). Lutein was the most stable carotenoid against thermal processing (decreased by 24-43%) followed by ß-carotene (decreased by 78-83%); other carotenoids were degraded nearly completely. Increased formation of (Z)-isomers by thermal processing has not been confirmed.

Endogenously produced hydrogen sulfide is involved in porcine oocyte maturation in vitro.

Hydrogen sulfide, one of three known gasotransmitters, is involved in physiological processes, including reproductive functions. Oocyte maturation and surrounding cumulus cell expansion play an essential role in female reproduction and subsequent embryonic development. Although the positive effects of exogenous hydrogen sulfide on maturing oocytes are well known, the role of endogenous hydrogen sulfide, which is physiologically released by enzymes, has not yet been described in oocytes. In this study, we observed the presence of Cystathionine ß-Synthase (CBS), Cystathionine γ-Lyase (CTH) and 3-Mercaptopyruvate Sulfurtransferase (3-MPST), hydrogen sulfide-releasing enzymes, in porcine oocytes. Endogenous hydrogen sulfide production was detected in immature and matured oocytes as well as its requirement for meiotic maturation. Individual hydrogen sulfide-releasing enzymes seem to be capable of substituting for each other in hydrogen sulfide production. However, meiosis suppression by inhibition of all hydrogen sulfide-releasing enzymes is not irreversible and this effect is a result of M-Phase/Maturation Promoting Factor (MPF) and Mitogen-Activated Protein Kinase (MAPK) activity inhibition. Futhermore, cumulus expansion expressed by hyaluronic acid (HA) production is affected by the inhibition of hydrogen sulfide production. Moreover, quality changes of the expanded cumuli are indicated. These results demonstrate hydrogen sulfide involvement in oocyte maturation as well as cumulus expansion. As such, hydrogen sulfide appears to be an important cell messenger during mammalian oocyte meiosis and adequate cumulus expansion.

Influence of parasitism on trace element contents in tissues of red fox (Vulpes vulpes) and its parasites Mesocestoides spp. (Cestoda) and Toxascaris leonina (Nematoda).

Bioaccumulation of cadmium, chromium, copper, manganese, nickel, lead, and zinc in 56 foxes (Vulpes vulpes) and their parasites Mesocestoides spp. (Cestoda) and Toxascaris leonina (Nematoda) was studied. The levels of heavy metals were determined in the livers and kidneys of the animals depending on parasitism in the following ranges: Pb, 0.029-3.556; Cd, 0.055-9.967; Cr, 0.001-0.304; Cu, 4.15-41.15; Mn, 1.81-19.94; Ni: 0.037-0.831; Zn, 52.0-212.9 microg/g dry weight (dw). Cd in parasites (0.038-3.678 microg/g dw) were comparable with those in the livers of the host and lower than in the kidneys (0.095-6.032 microg/g dw). Contents of Pb, Cr, Cu, Mn, Ni, and Zn in cestodes were predominantly higher than those in the kidney and liver of the host. Median lead levels in Mesocestoides spp. (45.6 microg/g dw) were 52-fold higher than in the kidney and liver of the red fox (Vulpes vulpes) infected by both parasites and median Pb values in T. leonina (8.98 microg/g dw) were 8-fold higher than in the tissues of the parasitized red fox. Bioaccumulation factors of copper, zinc, nickel, and manganese are lower than those of lead and mostly range from 1.9 to 24 for Mesocestoides spp. and from 1.5 to 6 for nematode T. leonina depending on the tissue of host and element. A significant decrease in the content of Pb was found in the kidney of animals infected by T. leonina (0.260 microg/g dw) as well as those infected by Mesocestoides spp. (0.457 microg/g dw) in comparison with the lead content (0.878 microg/g dw) in the kidneys of the nonparasitized red fox. Regardless of a bioaccumulation of copper and manganese in the parasites, a significant increase of the concentrations of Mn and Cu was observed in the host's livers infected predominantly by Mesocestoides spp.

Influence of parasitism on the use of small terrestrial rodents in environmental pollution monitoring.

Bioaccumulation of cadmium, chromium, copper, manganese, nickel, lead and zinc in small terrestrial rodents - voles and their cestode parasite Paranoplocephala dentata was studied. Contents of Pb, Mn, Ni and Zn in the parasite were found to be higher than in the kidney and liver of the parasitized animals. Lead level in the cestode was 37 fold higher than in the liver of the infected rodents. Bioaccumulation factors of zinc, nickel and manganese in the cestode are mostly in the range from 2 to 4.5. Considering the different contents of manganese and zinc in livers of non-parasitized and parasitized rodents, kidney tissue was found to be more reliable than liver as an indicator of environmental pollution by manganese and zinc; the kidneys of parasitized animals showed no significant change in the concentrations of those elements that are accumulated in the cestode.