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Within the framework of in vitro alternatives for in vivo safety assessment, the kinetic behaviour of a compound can be described by biokinetic models. These models, with emphasis on the physiologically based pharmacokinetic models, need a variety of biological, physicochemical and biochemical parameters. This paper deals with the possibilities for obtaining these data from in vitro studies. Examples are given for parameters on absorption (both dermal and intestinal), distribution and metabolism.

The friction between surfaces in relative motion lubricated by food emulsions has been measured. Different types of surfaces were tested, including metal, glass, rubber, and mucosal surfaces (pig tongue and pig esophagus). We demonstrate that the load-dependent behavior of the coefficient of kinetic friction was different for various types of surfaces used; it decreased with increasing load for the mucosal surfaces, but was constant for glass-metal surfaces. We show that this difference is an effect of the roughness and the deformability of the surfaces. For mucosal surfaces, the friction force did not depend on the oil fraction in the measured range of 10% to 40% (w/w). Furthermore, 3 commercial dairy products were tested between these surfaces. The lubrication properties of the low-fat (1.5% fat content) and high-fat (3.5% fat content) milk were comparable to those of the model emulsions. However, the friction force of the commercial cream (35% fat content) was found to be much lower. © 2006 Institute of Food Technologists.

The paper summarizes and evaluates data on estimates for the amount and amino acid (AA) composition of basal endogenous crude protein (CP) at the terminal ileum of pigs derived from 33 ileal digestibility studies with pigs (30-100 kg BW). Different methods and approaches are used to obtain these estimates: (A) the N-free diet method (16 studies), (B) calculations based on ileal digestibility values of diets with highly digestible protein sources (casein or wheat gluten) (11 studies), (C) the regression method (3 studies), (D) the enzymatic hydrolyzed casein (EHC) method (2 studies) and (E) the N-free diet method with intravenous infusion of AAs (1 study). The mean values for the flow of basal endogenous ileal CP (6.25 × N) for the five respective methods were 10.5, 12.6, 12.0, 17.1 and 12.7 g/kg dry matter (DM) intake. The overall mean value using all observations was 11.8 g/kg DM intake. The mean AA composition of basal endogenous protein (g/16 g N) was estimated as 3.2 for isoleucine, 4.2 for leucine, 3.4 for lysine, 1.0 for methionine, 1.8 for cystine, 2.9 for phenylalanine, 2.6 for tyrosine, 5.1 for threonine, 1.2 for tryptophan, 4.5 for valine, 3.4 for arginine and 1.6 for histidine. The flow of basal endogenous ileal CP and AAs can be considered as an inevitable loss for the pig. Data on the flow of basal endogenous protein and AAs can be used to calculate true or standardized ileal digestibility values or corrected apparent ileal digestibility values of feed ingredients for pigs. The use of more accurate and additive ileal digestibility values of AAs in feed ingredients improves diet formulation for pigs, in which AA supply via the diet is matched with AA requirements. © 2002 Elsevier Science B.V. All rights reserved.

Cross-linking experiments of skimmed bovine milk with bacterial transglutaminase isolated from Streptoverticillium mobaraense showed only some degree of formation of high-molecular-weight casein polymers. Studies on the nature of this phenomenon revealed that bovine milk contains an inhibitor of transglutaminase activity. Removal of the casein and whey proteins from the milk resulted in a protein-poor fraction that still inhibited transglutaminase activity at cross-linking of β-casein and in several activity assays of transglutaminase. The inhibitor was partially purified by column chromatography and appeared to be a heat labile low molecular weight component. Inhibition of transglutaminase activity was observed with microbial transglutaminase, plasma transglutaminase and guinea pig liver transglutaminase. The inhibiting activity was found in bovine, goat, sheep, and human milk, but could not be detected in horse milk.

The nitrogen balance and growth performance of piglets (12-14 kg initial body weight) were measured to evaluate the nutritive value of meal from Lupinus luteus cultivars 'Amulet' and 'Cybis', Lupinus albus cultivar 'Hetman," Lupinus angustifolius cultivar 'Saturn' and a commercial batch of lupin seeds from Australia (ALS). The inclusion level of lupin seed meal in the barley-based diets ranged from 310 to 410 g kg-1 to provide 120 g of crude protein from each lupin species per kilogram of diet. The average apparent total tract digestibility of dry matter (0.91) and crude protein (0.90) in L. luteus diets was as high as in the control (soybean) diet. The lowest (P < 0.05) digestibility of dry matter (0.86) and crude protein (0.83) was found for the diet containing seed meal from L. albus. Utilization of the apparently digested nitrogen (nitrogen retained as a proportion of nitrogen digested) was highest in the soybean group and both groups with L. angustifolius ('Saturn' and ALS) with values of 0.71, 0.72 and 0.71, respectively. Intermediate values were found for L. luteus ('Amulet', 0.68; 'Cybis', 0.68) and the lowest (0.55) for the L. albus group. The growth performance of pigs given diets with seeds of L. luteus and L. angustifolius was not different from that of pigs given the soybean diet, but pigs given the L. albus diet had a higher (P < 0.05) feed conversion ratio. It was concluded that L. albus cultivar 'Hetman' was less suitable than other varieties as a source of supplementary protein for young growing pigs when included in the diet at the level of 37 g kg-1. Seeds of L. luteus cultivar 'Cybis' and both cultivars of L. angustifolius were used at levels of up to 41 g kg-1 in diets without depression of growth performance as compared with soybean diet.

A procedure for a fast and simple purification of bovine plasma transglutaminase was developed, which resulted in a homogeneous enzyme preparation. Two different procedures were developed for the purification of pig erythrocyte transglutaminase, both of which resulted in partial purification. Both enzymes were used in cross-linking reactions of α-lactalbumin, β-lactoglobulin, bovine serum albumin, casein, hemoglobin, glycinin, and myosin. The substrate specificity was compared to that of bacterial transglutaminase isolated from Streptoverticillium mobaraense. The bacterial transglutaminase caused cross-linking of a wider range of proteins and, thus, exhibited a lower substrate specificity than the blood transglutaminases. In addition, differences exist in the necessity of the addition of reducing agents. These differences allow specific applications of blood and bacterial transglutaminases at protein cross-linking in single or complex protein systems. Chemicals/CAS: Transglutaminases, EC 2.3.2.13

The present study was designed to compare the ototoxic effects of volatile ethyl benzene in guinea pigs and rats. Rats showed deteriorated auditory thresholds in the mid-frequency range, based on electrocochleography, after 550-ppm ethyl benzene (8 h/day, 5 days). Outer hair cell (OHC) loss was found in the corresponding cochlear regions. In contrast, guinea pigs showed no threshold shifts and no OHC loss after exposure to much higher ethyl benzene levels (2500 ppm, 6 h/day, 5 days). Subsequently, a limited study (four rats and four guinea pigs) was performed in an attempt to understand these differences in susceptibility. Ethyl benzene concentration in blood was determined in both species after exposure to 500-ppm ethyl benzene (8 h/day, 3 days). At the end of the first day, blood of the rats contained 23.2±0.8-μg/ml ethyl benzene, whereas the concentration in guinea pig blood was 2.8±0.1 μg/ml. After 3 days, the concentration in both species decreased with respect to the first day, but the ethyl benzene concentration in rat blood was still 4.3 times higher than that in guinea pig blood. Thus, the difference in susceptibility between the species may be related to the ethyl benzene concentration in blood. © 2002 Elsevier Science Inc. All rights reserved. Chemicals/CAS: Benzene Derivatives; ethylbenzene, 100-41-4

In 1989, the Community Bureau of Reference started a research program to improve the quality of vitamin analysis in food. To achieve this task, vitamin methodology was evaluated and tested by interlaboratory studies and the preparation of certified reference materials, which will be used for quality control of vitamin measurements. The main improvements in methodology were achieved by testing and standardizing the extraction condition and enzymatic hydrolysis procedures. Results for each individual material are derived from five replicate determinations using at least two independent methods: Liquid chromatography (HPLC) and microbiological assay for vitamins B1, B2, and B6; and radioprotein binding and microbiological assays for vitamin B12. The certificate of analysis for four reference materials gives mass fraction values for water-soluble vitamins. These certified values were based on the acceptable statistical agreement of results from collaborating laboratories. Certified values with uncertainties (mg/kg dry matter) for each CRM are as follows: 4.63 (0.20) and 4.10 (0.51) for vitamins B1 and B6, respectively, in CRM 121 (wholemeal flour); 6.51 (0.24), 14.54 (0.3), 6.66 (0.43), and 0.034 (0.003) for vitamins B1, B2, B6, and B12, respectively, in CRM 421 (milk powder); 3.07 (0.17) and 4.80 (0.40) for vitamins B1 and B6, respectively, in CRM 485 (lyophilized mixed vegetables), and 8.58 (0.55), 106.8 (2.8), 19.3 (1.5), and 1.12 (0.044) for vitamins B1, B2, B6, and B12, respectively, in CRM 487 (lyophilized pig liver).

Within the framework of experiments related to the association between dietary fiber and breast cancer an in vitro test system was used to study the binding of estrogens to various fibers (e.g. cholestyramin, lignin and cellulose) and fiber sources (e.g. wheat bran, cereals, seeds and legumes). Furthermore, the in vivo apparent digestibility of the different fiber sources was tested using a mobile nylon bag technique in intestine-cannulated pigs. Estradiol-17β (E2) bound more strongly to the various fibers than did estrone (E1), estriol or estrone-3-glucuronide. At increasing pH (> 7) binding of both E1 and E2 to wheat bran decreased significantly. Cholestyramine and lignin bound almost all estrogens present in the medium. Linseed (91%), oats (83%), barley chaff (88%) and wheat bran (82%) are other excellent binders of E2. Corn, rye and white wheat flour showed lower binding capacity with a relatively low affinity. Cereals with the highest percentage of lignin in the fiber (> 3%) were also the fiber sources with the lowest apparent digestibility. Estrogens bound with the highest affinity (relative to bovine serum albumin) to these fiber sources. Together with wheat bran and lignin, oats, linseed and soybean seem to be products with good perspectives for in vivo evaluation of the lowering effect of dietary fiber on estrogen exposure of estrogen-sensitive tissues. Chemicals/CAS: Estradiol, 50-28-2; Estrogens; Serum Albumin

Fattening pigs are often infected with campylobacter. To eliminate campylobacter from the pig population, a top-down approach, involving the breeding and reproduction farms, seems appropriate. In order to investigate the effectiveness of a top-down approach, sows' faeces from the following farms were analysed for the presence of campylobacter: one specific pathogen free (SPF) farm, three top-breeding farms with no connection with SPF breeding, and a breeding farm repopulated with SPF sows after a period of vacancy (farm 5). The faeces samples from the SPF farm were free from campylobacter. The three top-breeding farms provided faeces samples which were 98% positive for campylobacter. However, only 22% of the faeces samples from farm 5 were positive for campylobacter. In a period of 20 months, the percentage of sows infected with campylobacter on farm 5 did not significantly increase. Genetic typing with ERIC-PCR and RFLP of campylobacter isolates from one of the top-breeding farms and from farm 5 showed a high diversity of campylobacter types. The results suggest that a campylobacter-free pig population can be established in breeding farms by combining a top-down approach (campylobacter-free top-breeding farms) with a strict regime of hygiene management.

Two experiments were conducted to determine apparent ileal DM and crude-protein (CP) digestibilities in rations fed to pigs. An evaluation was made of Cr2O3 and HCl-insoluble ash as digestive markers. In addition, the effects of body weight (BW) on apparent ileal DM and CP (N x 6.25) digestibilities were studied. In Expt 1, thirteen barrows averaging 35 kg BW were fitted with post-valve T-caecum (PVTC) cannulas to determine the apparent ileal DM and CP digestibilities of a wheat gluten-bran ration (B2) and a soyabean-meal ration (E1). Immediately after morning feeding ileal digesta samples were collected on an hourly basis for a total of 12 h. Subsequently, N and marker contents were determined in the samples. The postprandial patterns of N and Cr passage were more similar than those of N and HCl-insoluble ash. Therefore Cr2O3 is more suitable as a marker than HCl-insoluble ash. The apparent ileal CP digestibility coefficient of ration B2 derived using Cr2O3 as a marker was significantly (P < 0.05) higher by 0.018 compared with the value obtained using HCl-insoluble ash. The corresponding values for ration E2 obtained using Cr2O3 and HCl-insoluble ash were both 0.825. In Expt 2, apparent ileal DM and CP digestibilities were determined in eighteen rations using twelve barrows also fitted with PVTC cannulas (BW from 40 to 100 kg). The protein sources for these rations were from different groups of feedstuffs. In four and three of the rations apparent ileal DM and CP digestibilities respectively were significantly different (P < 0.05) when assessed using the two markers. The digestibility coefficients were not systematically higher or lower for either marker. Absolute differences were < 0.049 on average. Significant effects of live weight on apparent ileal CP digestibilities mere found.Chemicals/CAS: Biological Markers; Chromium Compounds; chromium oxide, 1308-38-9; Dietary Proteins; Nitrogen, 7727-37-9

The effects of lectins in the diet have been mainly studied in rats. An important question is whether results obtained in rats can be extrapolated to larger animals like the pig. Phaseolus vulgaris beans are rich in toxic lectins. Therefore a study was carried out to compare the effects of diets containing 200 g Phaseolus vulgaris beans (raw or toasted)/kg in rats and piglets. Live-weight gain, nitrogen digestibility and N balance were much lower in piglets than in rats fed on diets containing raw beans. Live-weight gain and N balance were slightly negative in the piglets. When toasted beans were given, live-weight gain and N balance values were reduced in piglets but hardly at all in rats. Giving raw beans caused hypertrophy of the pancreas in the rats but in piglets the weight of the pancreas was reduced. Spleen weight was depressed in the piglets but not in the rats. Weight of liver was not affected in either animal species. When toasted beans were given no effects on the weights of pancreas, spleen or liver were found in piglets or rats. It was concluded that the piglet is much more sensitive to antinutritional factors in thge Phaseolus vulgaris bean than the rat.

A comparison was made of the effects of antinutritional factors present in Phaseolus vulgaris on piglets, rats and chickens. Also the hypothesis of whether the negative effect on weight gain due to the inclusion of raw Phaseolus vulgaris in the diet can be attributed to an insufficient supply of amino acids was tested. Test diets containing 200 g raw Phaseolus beans/kg were balanced for digestible protein and amino acids; in one diet extra casein was incorporated. The main response criteria were live-weight gain and the weight of various organs including the intestine. Live-weight gain in piglets was markedly reduced during feeding 200 g raw Phaseolus vulgaris/kg in the diet, but not in rats and chickens. Addition of casein did not improve the weight gain of the piglets, indicating that a toxic factor was responsible for the reduced weight gain and not an insufficient supply of amino acids. The weights of the spleen and thymus were markedly reduced in the piglets when the diets with raw Phaseolus beans were given, but not in the rats and chickens. Additional supply of casein did not change this effect. Indications were found that when the supply of dietary protein is adequate there is no reduction in pancreas weight with raw Phaseolus beans as was obversed in previous experiments. The weight of the intestine was increased in all three species due to feeding raw Phaseolus vulgaris.

Hemicellulose consists primarily of pentose sugars, joined together in a polysaccharide chain with D-xylose as the most abundant component. Ileal digestibility and urinary excretion of D-xylose and associated effects of this pentose sugar on ileal and faecal digestibility of dry matter (DM), organic matter (OM), gross energy (GE) and nitrogen were studied in pigs. Castrated pigs were prepared with a post-valvular T-caecum cannula to measure ileal digestibility. Faecal digestibility was measured in non-cannulated pigs. D-xylose was given at dietary inclusion levels of 100 and 200 g/kg, and the control sugar, D-glucose, at a rate of 200 g/kg diet. Ileal digestibility of D-xylose as well as that of D-glucose was found to be close to 100%. The presence of D-xylose in the diet decreased ileal digesta pH and increased ileal flow of volatile fatty acids, suggesting the occurrence of microbial degradation of D-xylose in the pig small intestine. In pigs fed on the 100 g D-xylose/kg diet, 44.5% of the D-xylose intake appeared in the urine. This percentage increased significantly to 52.6 when pigs were fed on the 200 g D-xylose/kg diet. Ileal and faecal digestibility of DM, OM, GE and N, as well as N retention, decreased significantly in pigs fed on the 200 g D-xylose/kg diet. Chemicals/CAS: Fatty Acids, Volatile; Xylose

Because of the public concern surrounding the issue of the safety of genetically modified organisms, it is critical to have appropriate methodologies to aid investigators in identifying potential hazards associated with consumption of foods produced with these materials. A regent panel of experts convened by the Food and Agriculture Organization and World Health Organization suggested there is scientific evidence that using data from animal studies will contribute important information regarding the allergenicity of foods derived from biotechnology. This view has given further impetus to the development of suitable animal models for allergenicity assessment. This article is a review of what has been achieved and what still has to be accomplished regarding several different animal models. Progress made in the design and evaluation of models in the rat, the mouse, the dog and in swine is reviewed and discussed.

Quercetin is a dietary polyphenolic compound with potentially beneficial effects on health. Claims that quercetin has biological effects are based mainly on in vitro studies with quercetin aglycone. However, quercetin is rapidly metabolized, and we have little knowledge of its availability to tissues. To assess the long-term tissue distribution of quercetin, 2 groups of rats were given a 0.1 or 1% quercetin diet [∼50 or 500 mg/kg body weight (wt)] for 11 wk. In addition, a 3-d study was done with pigs fed a diet containing 500 mg quercetin/kg body wt. Tissue concentrations of quercetin and quercetin metabolites were analyzed with an optimized extraction method. Quercetin and quercetin metabolites were widely distributed in rat tissues, with the highest concentrations in lungs (3.98 and 15.3 nmol/g tissue for the 0.1 and 1% quercetin diet, respectively) and the lowest in brain, white fat, and spleen. In the short-term pig study, liver (5.87 nmol/g tissue) and kidney (2.51 nmol/g tissue) contained high concentrations of quercetin and quercetin metabolites, whereas brain, heart, and spleen had low concentrations. These studies have for the first time identified target tissues of quercetin, which may help to understand its mechanisms of action in vivo. © 2005 American Society for Nutritional Sciences.

In vitro and in vivo skin absorption of the pesticide propoxur (2-isopropoxyphenyl N-methyl carbamate, commercially Baygon(TM) and Unden(TM); log Po/w 1.56, MW 209.2) was investigated. In vivo studies were performed in rats and human volunteers, applying the test compound to the dorsal skin and the volar aspect of the forearm, respectively. In vitro experiments were carried out in static diffusion cells using viable full-thickness skin membranes (rat and human), non-viable epidermal membranes (rat and human) and a perfused-pig-ear model. Percutaneous penetration of propoxur in human volunteers was measured by analysis of its metabolite (2-isopropoxyphenol) in blood and urine; in all other studies radiolabeled propoxur ([ring-U-14C]propoxur) was used. In order to allow for direct comparison, experimental conditions were standardized with respect to dose (150 μg propoxur per cm2), vehicle (60% aqueous ethanol) and exposure time (4 h). In human volunteers, it was found that approximately 6% of the applied dose was excreted via the urine after 24 h, while the potential absorbed dose (amount applied minus amount washed off) was 23 μg/cm2. In rats these values were 21% and 88 μg/cm2, respectively. Data obtained in vitro were almost always higher than those obtained in human volunteers. The most accurate in vitro prediction of the human in vivo percutaneous absorption of propoxur was obtained on the basis of the potential absorbed dose. The absorbed dose and the maximal flux in viable full-thickness skin membranes correlated reasonably well with the human in vivo situation (maximal overestimation by a factor of 3). Epidermal membranes overestimated the human in vivo data up to a factor of 8, but the species-differences observed in vivo were reflected correctly in this model. The data generated in the perfused-pig-ear model were generally intermediate between viable skin membranes and epidermal membranes. Chemicals/CAS: Insecticides; Propoxur, 114-26-1

The present study was designed to evaluate the differential effects of dietary glucose, lactose and starch on small-intestinal morphology, organ weights, pH of chyme and haptoglobin levels in blood plasma of weaned piglets. It was hypothesised that lactose consumption would ameliorate the weaning-induced decrease in gut integrity. A total of forty-two barrows were used. Piglets were weaned at 27 (SD 0.8) d of age and weighed 8.0 (SD 0.51) kg. On the day before weaning (day -1) all pigs were blocked according to body weight and randomly assigned to seven groups (n 6 per group). The groups differed in diet and day of dissection. On the day of weaning, dissection was performed on one group of six piglets. The remaining groups were fed one of three experimental diets in which glucose, lactose or starch had been iso-energetically exchanged, supplying 24 % dietary energy. The piglets received a liquid diet (air-dry meal:water of 1:2, w/w). The piglets were given access to a maximum of dietary energy in order to prevent confounding between feed intake and villus architecture. The piglets were dissected and sampled on days 0, 3, or 10 post-weaning. The results show,that the carbohydrate source did not affect growth performance, organ weights, villus architecture, pH of chyme and plasma haptoglobin level. The weaning transition resulted in decreased villus height and increased haptoglobin levels. In the contents of the caecum and large intestine, the pH decreased after weaning. It is concluded that at least under conditions of similar feed intake and low infectious pressure, dietary lactose does not ameliorate the weaning-induced compromise of small-intestinal integrity when compared with either glucose or starch. Chemicals/CAS: glucose, 50-99-7, 84778-64-3; haptoglobin, 9087-69-8; lactose, 10039-26-6, 16984-38-6, 63-42-3, 64044-51-5; starch, 9005-25-8, 9005-84-9; water, 7732-18-5; Dietary Carbohydrates; Glucose, 50-99-7; Haptoglobins; Lactose, 63-42-3; Starch, 9005-25-8

Barlican, a β-glucanase enzyme obtained from Trichoderma reesei, was produced by a fermentation process and subjected to a series of toxicological tests to document its safety for use as a feed additive. The enzyme product was examined for general oral toxicity, inhalation toxicity, irritation to eye and skin, skin sensitization and mutagenic potential. An extensive literature search on the production organism was also conducted. Furthermore, safety for target species was assessed in a 28-day oral toxicity study with broilers. A strong skin-sensitizing potential of the β-glucanase enzyme was detected, but no other evidence of oral or inhalation toxicity, mutagenic potential, eye or skin irritancy was found. Feeding of the β-glucanase enzyme at dietary levels up to 10,000 ppm in the 90-day subchronic toxicity study in rats did not induce noticeable signs of toxicity. In addition, no adverse effects were observed when broiler chicks were fed dietary concentrations of the β-glucanase enzyme up to eight times the daily recommended dose. it is therefore concluded that this β-glucanase preparation is safe for use in feed of the intended target species. However, some occupational hearth precautions should be taken to avoid skin contact and inhalation, as is the case for almost all enzyme proteins.

Osteoarthritis (OA), one of the most common diseases among the elderly, is characterized by the progressive destruction of joint tissues. Its etiology is largely unclear and no effective disease-modifying treatment is currently available. Metabolic fingerprinting provides a novel tool for the identification of biomarkers. A metabolic fingerprint consists of a typical combination of metabolites in a biological fluid and is identified by a combination of 1H NMR spectroscopy and multivariate data analysis (MVDA). The current feasibility study was aimed at identifying a metabolic fingerprint for OA and applying this in a nutritional intervention study. Urine samples were collected from osteoarthritic male Hartley guinea pigs (n = 44) at 10 and 12 mo of age, treated from 4 mo onward with variable vitamin C doses (2.5-3, 30 and 150 mg/d) and from healthy male Strain 13 guinea pigs (n = 8) at 12 mo of age, treated with 30 mg vitamin C/d. NMR measurements were performed on all urine samples. Subsequently, MVDA was carried out on the data obtained using NMR. An NMR fingerprint was identified that reflected the osteoarthritic changes in guinea pigs. The metabolites that comprised the fingerprint indicate that energy and purine metabolism are of major importance in OA. Metabolic fingerprinting also allowed detection of differences in OA-specific metabolites induced by different dietary vitamin C intakes. This study demonstrates the feasibility of metabolic fingerprinting to identify disease-specific profiles of urinary metabolites. NMR fingerprinting is a promising means of identifying new disease markers and of gaining fresh insights into the pathophysiology of disease.