Extraction recoveries and stability of diarrhetic shellfish poisoning (DSP) toxins in naturally contaminated samples

During the last few years the occurrence of a high percentage of esters of diarrhetic shellfish poisoning (DSP) toxins has been observed in shellfish from the Portuguese coast. Most of the commercial bivalves contain DSP toxins in ester forms, either acyl derivatives of okadaic acid (OA) or of dinophysistoxin-2 (DTX-2). The stability of these toxins in shellfish tissues and in raw methanol extracts was investigated in two different naturally contaminated species, mussel and carpet shell, over a 4-week period. The results for both species revealed that DSP toxins were more stable in tissue than in raw methanol extracts. Losses of DSP toxins were seen in the first 2 weeks and were more than 30%, but after that a period of stabilization was observed. The decrease was due probably from losses of esters of OA and DTX-2, the free toxins were stable over the period studied. The extraction most commonly used for chemical and biochemical assays relied on methanolic extraction with aqueous 80% methanol. In this work we have tested the extraction solvent on the extractability of DSP toxins from several naturally contaminated species. A single dispersive extraction with methanol, with solvent ratios of 70%, 80%, 90% and 100%, were tested. After alkaline hydrolysis of esterified toxins and clean-up with hexane and dichloromethane, the samples were analysed by liquid chromatography-mass spectrometry (LC-MS). The recovery of DSP toxins increased with increasing percentages of methanol up to 90%. A decrease in recovery with 100% methanol was observed probably due to problems during the liquid–liquid partitioning.     

Effects of ball‐milling treatment on mussel (Mytilus edulis) protein: structure,functional properties and in vitro digestibility

In this study, the effects of ball‐milling treatment on structural properties and functional properties of mussel protein were investigated. In vitro protein digestibility and free amino acid contents were also evaluated. Ball‐milling treatment did not change the primary structure of mussel protein, but caused changes of secondary structure. The tertiary and quaternary structure also changed. After 20 min of ball‐milling treatment, the whiteness and oil‐binding capability of mussel protein significantly improved from 13.67 to 26.62 and 43.76% to 196.00%, while the protein solubility and water‐holding capability of mussel protein significantly decreased from 74.89% to 53.10% and 215.67% to 90.91%. Ball‐milling treatment increased the in vitro digestibility significantly. These results provide theoretical basis for the utilisation of mussel protein in food industry.     

A comparison of the in vitro biotransformation of (–)‐epicatechin and procyanidin B2 by human faecal microbiota

The catabolism by human faecal microbiota of (–)‐epicatechin ( 1 ) (2, 3‐cis stereochemistry) and its dimer pure procyanidin B2 ( 2 ), has been compared using a static in vitro culture model. The catabolites were characterised by LC‐MSⁿ, UV absorption and relative retention time, and quantified relative to standards. No more than ～10% of procyanidin B2 ( 2 ) was converted to epicatechin ( 1 ) by scission of the interflavan bond. Five phenolic acid catabolites (M_r<290) were unique to 2 , and ten phenolic acid catabolites (M_r<290) were common to both substrates. The dominant catabolites (≥24 h incubation) were 5‐(3′‐hydroxy phenyl) valeric acid ( 9 ), 3‐(3′‐hydroxyphenyl) propionic acid ( 10 ) and phenyl acetic acid ( 12 ) (maximum yields 27.4±4.2, 38.2±4.2, 22.7±2.9%, respectively, from 1 and 9.4±1.2, 52.8±2.1, 28.8±1.6%, respectively, from 2 ). Substrate 2 was degraded twice as rapidly as 1 . Evidence is presented for the production of previously unreported catabolites of 2 that retain the flavanol A‐ring and the C4→C8 interflavan bond. It was confirmed that catabolism favoured removal of the 4′‐hydroxyl rather than the 3′‐hydroxyl group and that both β‐oxidation and α‐oxidation occurred.     

酸化蛋清粉的体外降血脂及抗氧化作用

为探究酸化蛋清粉对油酸诱导Hep G2细胞建立的体外肝细胞脂肪堆积的清除作用,将质量浓度分别为4 mg/mL、2 mg/mL、0.8mg/m L的酸化蛋清粉孵育经1.0mol/L的油酸诱导24h的Hep G2细胞后,用油红O染色观察细胞内脂质沉积情况,测定细胞内TC、TG含量及SOD活性的变化,检测ROS荧光强度,测定·DPPH自由基及·OH自由基的清除率。结果显示酸化蛋清粉浓度为2 mg/mL时具有最显著的降血脂效果,TG和TC的清除率分别为27.27%±0.02%、44.66%±0.02%;SOD活性及ROS荧光检测结果显示:随着酸化蛋清粉浓度的升高,细胞内抗氧化能力呈现上升趋势;·DPPH自由基和·OH自由基的清除能力随酸化蛋清粉浓度升高呈现出显著的随剂量依赖性的上升趋势,当酸化蛋清粉浓度18 mg/m L时,其清除率分别为88.58%±0.04%、87.82%±0.03%。结果证明酸化蛋清粉具有显著的体外降血脂及抗氧化作用。     

In vitro bioavailability of iron and calcium in cereals and derivatives: A review

Cereals are a staple food in both developed and developing countries, and are considered to be the best vehicle for iron and calcium fortification, as an important strategy for combating dietary deficits. Inadequate dietary intake of iron and calcium is related to a number of disease conditions such as anemia, osteoporosis, hypertension, and different cancers. From a nutritional point of view, it is interesting to know not only the amount of minerals consumed, but also their bioavailability. The present study reviews the current knowledge on the in vitro bioavailability of iron and calcium in cereals, placing emphasis on the methodologies used and on the influence of dietary factors and food processing.     

蜂王浆冻干粉对HepG2细胞葡萄糖消耗作用及对糖尿病小鼠降糖作用的实验研究

目的:研究蜂王浆冻干粉体内外降糖作用。方法:采用与人肝细胞表型相似的HepG2细胞,检测24h后培养液中葡萄糖的消耗量,用MTT法监测细胞增殖的情况;以四氧嘧啶造成小鼠糖尿病模型,观察蜂王浆对糖尿病模型小鼠的降糖作用。结论:蜂王浆冻干粉可使高糖状态下HepG2细胞的葡萄糖消耗量增加,对胰岛素刺激的HepG2细胞的葡萄糖消耗量增加有协同作用;能降低四氧嘧啶所致糖尿病小鼠的血糖。     

In vitro and in vivo anti-Staphylococcus aureus activities of a new disinfection system utilizing photolysis of hydrogen peroxide

The present study aimed to evaluate in vitro and in vivo antibacterial activity of hydroxyl radical generation system by photolysis of H(2)O(2), which is a new disinfection system for the treatment of oral infection diseases such as periodontitis developed in our laboratory. Firstly, generation of the hydroxyl radical by the photolysis of H(2)O(2) in which 1 mol l(-1) H(2)O(2) was irradiated with a dual wavelength-light emitting diode (LED) at wavelengths of 400 and 465 nm was confirmed by applying an electron spin resonance-spin trapping technique. Secondly, the bactericidal effect of the system was examined under a similar condition in which Staphylococcus aureus suspended in 1 mol l(-1) H(2)O(2) was irradiated with LED light, resulting in substantial reduction of the colony forming unit (CFU) of the bacteria within a short time as 2 min. Finally, in vivo antibacterial effect of the photolysis of H(2)O(2) on a rat model of S. aureus infection was evaluated by a culture study. Since a significant reduction of recovered CFU of S. aureus was obtained, it is expected that in vitro antibacterial effect attributable to hydroxyl radicals generated by photolysis of H(2)O(2) could be well reflected in in vivo superficial bacterial infection.     

Bioavailability of Elemental Iron Powders in Bread Assessed with an In vitro Digestion/Caco-2 Cell Culture Model

ABSTRACT: Iron fortification of staple foods is arguably the most widely used strategy for increasing the iron intake of populations. Although FeSO₄ is a bioavailable form of iron, elemental iron powders are often used to fortify products with a long shelf-life, such as wheat flours, to avoid problems associated with the reactive nature of FeSO₄. Therefore, the objectives of this study were to compare the bioavailabilities of elemental iron powders manufactured with different production methods in wheat flour breads and to determine the effects of added ascorbic acid and baking, using an in vitro digestion/Caco-2 cell culture model. Two types of wheat flour (low-extraction and high-extraction) were fortified with 10 different commercial elemental iron powders and baked into breads. Iron bioavailabilities from the resulting breads, with and without added ascorbic acid, were evaluated using FeSO₄ as the control. Depending on the type of wheat flour, bioavailabilities of several powders were comparable to FeSO₄, but there was no consistent trend as to which production method produced the most bioavailable powder. In general, ascorbic acid enhanced, whereas the baking process reduced iron bioavailability from bread. Our results suggest that some elemental iron powders are potential alternatives to FeSO₄. Human studies are warranted before any of these powders are selected for national fortification programs.     

In vivo and in vitro models of the human colonic flora.

The study of colonic flora composition and metabolism presents considerable methodological problems. Attempts to circumvent these problems have led to the development of numerous in vitro and in vivo models to simulate the human colon and its microbial population. In terms of in vivo models, conventional laboratory animals have many limitations. Data of greater relevance to man can be obtained by using germ-free rodents associated with human colonic bacteria. The applications of such animals to studies of toxicity of chemicals and gastrointestinal infections are discussed. The advantages and disadvantages of the various in vitro systems for studying gut microflora and its metabolic activity (from simple static cultures to the more sophisticated continuous and semicontinuous flow models) are reviewed. The apparatus involved is described together with practical information on media, running conditions, and sampling. The bacteriological and metabolic criteria for establishing the similarity of the models to the in situ colonic flora are also discussed. The final sections of the review are devoted to the major applications (current and future) of the models, including fermentation studies on dietary fiber, metabolism of nutrients and foreign compounds (including carcinogens) in food, and the investigation of colonization resistance.     

In vitro fermentation of polysaccharides of rye,wheat and oat brans and inulin by human faecal bacteria

The in vitro fermentabilities of rye, wheat and oat brans and of a commercial fibre preparation, inulin, were compared. The brans were first digested enzymatically to remove starch and protein. The digested brans and inulin were then fermented with human faecal inoculum. The progress of fermentation was studied by following the consumption of carbohydrates and the production of short‐chain fatty acids and gases. Inulin, a short fructose polymer, was consumed significantly faster than the more complex carbohydrates of cereal brans. Carbohydrates of oat bran (rich in β‐glucan) were consumed at a higher rate than those of rye and wheat brans (rich in arabinoxylan). In all brans, glucose was consumed faster than the other main sugars, arabinose and xylose, and arabinose was degraded only slightly. The total production of short‐chain fatty acids was slightly higher with oat bran than with rye and wheat brans and inulin. In the fermentation of inulin, relatively more butyric acid and less propionic acid were produced than in the fermentation of brans. The decrease in pH was also greater in the case of inulin. Wheat bran led to a slightly slower gas formation than rye and oat brans. Formation of gases was fastest and greatest in the case of inulin. In conclusion, rye, wheat and oat brans were fermented in a rather similar way. Fermentation of the brans was different from that of inulin. Cereal brans might serve as a more balanced source of dietary fibre supplement than gas‐producing, readily fermentable polysaccharides such as inulin. © 2000 Society of Chemical Industry     

Effect of fenpropimorph, prochloraz and tebuconazole on growth and production of T-2 and HT-2 toxins by Fusarium langsethiae in oat-based medium

Fusarium langsethiae has been isolated from infected cereals in central and northern Europe where it has been identified in the last decade as the main species involved in the occurrence of high levels of T-2 and HT-2 toxins, mainly in oats. The efficacy of three fungicides (prochloraz, tebuconazole, fenpropimorph) for controlling growth of two strains of F. langsethiae isolated from oats was examined at 0.96 and 0.98 a_w at 15, 20 and 25 °C on oat-based media. The concentrations necessary for 50 and 90% growth inhibition (ED₅₀ and ED₉₀ values) were determined. The effect on the trichothecene type A mycotoxins T-2 and HT-2 was also determined. Without fungicides both strains grew faster at 0.98 than at 0.96 a_w and the influence of temperature on growth rates was 25 > 20 > 15 °C. Prochloraz and tebuconazole were more effective than fenpropimorph against F. langsethiae. Strain, temperature and type of fungicide significantly influenced the ED₅₀ and ED₉₀ values for growth. The concentration ranges under different environmental conditions were: prochloraz (0.03-0.1 and 0.3-1.5), tebuconazole (0.06-0.9 and 1.3-8.2), and fenpropimorph (22-59 and 125-215 mg l⁻¹). Production of T-2 and HT-2 toxins was influenced by temperature, a_w, type of fungicide and dose. Levels of T-2 were usually higher than those of HT-2 under the same conditions. The biosynthesis of T-2 toxin increased after 10 day incubation, but was reduced with decreasing temperature and increasing fungicide dose. At 0.98 a_w T-2 levels increased in cultures containing fenpropimorph while at 0.96 a_w the toxin concentrations increased in response to the other two fungicides. Low doses of prochloraz or tebuconazole enhanced toxin production when compared with untreated cultures for strain 2004-59 at 0.96 a_w and 20-25 °C. HT-2 was hardly detectable in the treatments with prochloraz or tebuconazole at 0.98 a_w. This is the first study on the effect of these anti-fungal compounds on control of growth of F. langsethiae and on production of T-2 and HT-2 toxins in oat-based media.     

In vitro assessment of water resistance of sun care products: a reproducible and optimized in vitro test method

The aim of the study was to develop a simple reproducible and reliable in vitro water resistance (WR) method to assess the sun care products. This paper is the result of a scientific collaboration between seven different international industrial laboratories and testing institutes. The same group has already achieved an in vitro protocol for the sun protection factor (SPF) determination [1]. The in vitro WR of sunscreens was tested by applying the same principle as in vivo, which determines the percentage of retention of sunscreen products by assessing the SPF before and after water immersion. Special care was taken to study the parameters influencing the WR and the possibility to follow the kinetics of sunscreen retention during water immersion. The influence of different water qualities has been tested, and osmosed water (1-3 microS cm(-1)) was chosen for the main ring study. Measurement was carried out after 5, 20 and 40 min of immersion. Histograms of selected products demonstrate the percentage of WR at all measuring times and centres, and the regression coefficient to the in vivo determination was shown and statistical calculations clearly demonstrate the reproducibility of the results between the different evaluation centres. The presented method is a practical, convenient and relevant tool for WR screening of sun care and skin care products. It even has the potential to be the starting point for the replacement of the in vivo method in future.     

罗非鱼皮胶原蛋白水解产物的体外抗氧化活性和体内抗衰老作用

对罗非鱼皮胶原蛋白水解制备水解产物（tilapia skin gelatin hydrolysate,TSGH）,分离得到3 个肽组分（TSGH-1、TSGH-2和TSGH-3）,分析肽组分氨基酸组成,并对TSGH-3的体外抗氧化活性和体内抗衰老能力进行评价。结果表明：TSGH-3的分子质量低,疏水性氨基酸含量高,从自由基清除和金属螯合两个方面证实了TSGH-3具有很好的体外抗氧化活性。TSGH-3能显著增加由D-半乳糖诱导衰老大鼠的血清、肝脏和肾脏组织的总抗氧化能力、超氧化物歧化酶和谷胱甘肽过氧化物酶活性,并且能显著降低丙二醛的含量。此外,TSGH-3的摄入可以保护大鼠的肝脏和肾脏指数。因此,TSGH-3在抗氧化、抗衰老方面具有较好的应用潜力。     

In vitro and in situ disappearance of β‐carotene and lutein from lucerne (Medicago sativa) hay in bovine and caprine ruminal fluids

Two experiments were conducted to determine the rumen fluid disappearance rates (kd) of β‐carotene, lutein, total carotene and total xanthophyll from lucerne (Medicago sativa) hay, in two ruminant species: Brahman steers (fat‐pigmenting) and Granadine goats (non‐pigmenting). Within species, the in vitro and the in situ methods were compared. The concentration of carotenoid compounds was determined by spectrophotometry and high performance liquid chromatography. The in vitro disappearance trends were linear for all compounds (P<0.01). β‐carotene kd were 0.13 and 0.37; lutein, 0.20 and 0.25; total carotene, 0.20 and 0.62 and total xanthophyll, 0.30 and 0.77 h ⁻¹ for steers and goats, respectively. The in situ disappearance trends were quadratic (P<0.01). Dry matter kd were 1.9 and 1.5% h⁻¹; cellular content, 2.0 and 2.3; β‐carotene, 2.5 and 1.2; lutein, 2.5 and 1.5; total carotene, 2.2 and 1.0 and total xanthophyll, 2.1 and 1.1% h⁻¹ for steers and goats, respectively. The large disappearance rates of carotenoids observed in the in situ method vs the virtual absence of disappearance in the in vitro method in both species, can be related to the dry matter and cellular content kd. These results suggest that carotenoids disappear probably by joining the cellular content and not by their direct destruction or by attack from the ruminal microorganisms, and the ruminal disappearance is independent of the species studied. © 1999 Society of Chemical Industry     

In vitro digestion and characterisation of 2S albumin and digestion‐resistant peptides in pecan

The 2S albumins are one of the major protein families involved in severe food allergic reactions to nuts, seeds and legumes, thus potentially making these proteins clinically relevant for allergic sensitisation and potential diagnostic markers. In this study, we sought to purify native 2S albumin protein from pecan to further characterise this putative allergen. The purified 2S albumin, Car i 1, from pecan was found to be resistant to digestion by pepsin in simulated gastric fluid (SGF) and comparatively stable to proteolysis by trypsin and pancreatin in simulated intestinal fluid (SIF). Digestion of purified Car i 1 in SGF and SIF resulted in formation of different digestion‐resistant peptides that were capable of binding IgE antibodies from allergic individuals. Digestion stability of Car i 1 and formation of digestion‐resistant antigenic peptides may explain why it is a potent sensitising protein in pecan for susceptible individuals. The observation that digestion‐resistant peptides are able to bind IgE implies that pecan can trigger systemic allergic reactions even after digestion in the stomach and small intestine.     

Oxidation of Polyphenols and the Effect on In vitro Iron Accessibility in a Model Food System

ABSTRACT: Tannic acid and green tea extract were added to a model food system of wheat bread, before and after incubation with polyphenol oxidase (mushroom tyrosinase), and the effect on In vitro iron accessibility was studied. Tannic acid and green tea extract had a marked inhibitory effect on the In vitro accessibility of iron when added to the model food system. Incubation of the polyphenols with tyrosinase before addition to the model food system significantly increased the accessibility of iron. The results from the study therefore suggest that oxidation of polyphenols may be a promising way to increase the bioavailability of iron in polyphenol containing foods.     

In vitro and in vivo assessment of the effect of Laurus novocanariensis oil and essential oil in human skin

Laurus novocanariensis is an endemic plant from the Madeira Island forest that derives a fatty oil, with a strong spicy odour, from its berries that has been used for centuries in traditional medicine to treat skin ailments. This work aimed to investigate the effect of the application of both the oil and its essential oil on normal skin, to assess their safety and potential benefits. Diffusion studies with Franz cells using human epidermal membranes were conducted. The steady‐state fluxes of two model molecules through untreated skin were compared with those obtained after a 2‐h pre‐treatment with either the oil or the essential oil. Additionally, eleven volunteers participated in the in vivo study that was conducted on the forearm and involved daily application of the oil for 5 days. Measurements were performed every day in the treated site with bioengineering methods that measure erythema, irritation and loss of barrier function. Slightly higher steady‐state fluxes were observed for both the lipophilic and the hydrophilic molecule when the epidermal membranes were pre‐treated. Nevertheless, such differences had no statistical significance, which seems to confirm that neither the oil nor the essential oil impaired the epidermal barrier. Results collected with the Chromameter, the Laser Doppler Flowmeter and the visual scoring are in agreement with those established in the in vitro study. They indicate that the repeated application of the oil did not cause erythema, because the results observed in the first day of the study were maintained throughout the week. Application of the oil did not affect the skin barrier function, because the transepidermal water loss remained constant throughout the study. The stratum corneum hydration was slightly reduced on days 4 and 5. This work shows that both the oil and the essential oil were well tolerated by the skin and did not cause significant barrier impairment or irritation.