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The national antimicrobial resistance monitoring determines the current quantitative resistance level of life-stock pathogens, in order to permit the evaluation and surveillance of the distribution of resistances on a valid basis. During the examination period from June 2002 to July 2003, a total of 1849 pathogens was collected, following a representative German-wide pattern in collaboration with 29 laboratories. The selection of examined bacterial strains included different specimen causing respiratory diseases in fattening pigs (Pasteurella multocida, Bordetella bronchiseptica) and cattle (Pasteurella multocida, Mannheimia haemolytica), respectively, as well as strains causing mastitis in dairy cows (Staphylococcus spp., Streptococcus spp., E. coli). Determination of the in-vitro susceptibility (minimal inhibitory concentration) to at least 17 antimicrobial agents was performed centrally by the BVL using the microdilution broth method. The findings approximately match results of an analogous study conducted in 2001, and correspondingly revealed significantly lower resistance-values in comparison to data published for Germany so far. No correlation could be established between the incidence of resistance and differing stock densities. By means of the resistance monitoring data gathered by the BVL, the risk potential of antimicrobials applied in Germany can be reliably defined. This valuable information about the epidemiological situation of resistance in Germany can be helpful to veterinarians as a decision guidance when choosing appropriate means of therapy. Accordingly, the results stated by the BVL are apt to provide an important contribution to improving the safety of food-animal products. The experiences obtained from the monitoring also show that valid data about the antimicrobial susceptibility can only be raised on an interdisciplinary approach (federal agencies, county diagnostic laboratories, universities, industry). Currently, the BVL put into practice a study with an extended spectrum of bacterial species and indications.

In 2001, the first Germany comprehensive cross-sectional study into the sensitivity to antimicrobial substances of selected pathogenic bacteria from food-producing animal species (dairy cows, fattening pigs) was conducted by the Federal Office of Consumer Protection and Food Safety, BVL (formerly, Federal Institute for Health Protection of Consumers and Veterinary Medicine, BgVV). Initial experience from national resistance monitoring revealed that the necessary organisational structures may be suitably established in a federal system. The quantitative sensitivity results (minimum inhibitory concentration, MIC) for the bacterial species examined showed lower resistance values in contrast to German data published previously and also in comparison with results from other European countries. Based on the experience from this pilot study, an urgent need has been identified to continue this interdisciplinary approach to tackle the resistance problem together with human medicine. After analysing the data from the pilot monitoring study, in 2002 the BVL started a year's study with an extended selection of bacterial species and indications. In the future, bacterial samples from private diagnostic laboratories and universities will also be included in the resistance monitoring system.

Determination of chloramphenicol (CAP) residues in egg by gas chromatography/high-resolution mass spectrometry (GC/HRMS) with negative chemical ionization and gas chromatography with electron capture detection (GC-ECD) is described. A cleanup based on acetonitrile extraction followed by solid-phase extraction with silica gel and gel filtration columns was developed for extraction of CAP residues from whole egg. For quantitation, the internal standards used were the meta isomer of CAP (m-CAP) for GC-ECD and both m-CAP and deuterium-labeled CAP (D5-CAP) for GC/HRMS. For GC/HRMS, evaluation was performed by selecting characteristic ions at m/z 466 for CAP and m-CAP and at m/z 471 for D5-CAP. Both methods were validated with egg samples fortified at 0.4-2.0 micrograms/kg and lyophilized egg samples from animals treated at the 1 microgram/kg concentration level. The coefficient of variation was below 10%. Limits of detection and quantitation of both methods were about 0.3 and 0.5 microgram/kg, respectively. For confirmation of CAP residues, the relative ion abundance (m/z 466 and 468) was calculated. Further confidence was obtained by comparison of accurate monoisotopic masses from a spectrum library.

The creation of the single market and the intended free trade of goods, individuals, capital investments and services lead to the question if the quality standards of veterinary drugs are going to be changed in view of an improved animal health and consumer protection. However, the comparison between registration and surveillance criteria that result from the German Law for Medicinal Products and the criteria valid in the European Union as a consequence of Regulations and Directives shows that no changes leading to a reduced quality are to be expected. On the contrary, the application of EU Directives to the respective national law results in a number of improvements, like for example the environmental assessment of veterinary drugs.

Pharmacokinetic studies on Albendazole after peroral administration demonstrate a rapid and complete biotransformation. The major metabolites Albendazole sulphoxide and -sulphone were detected in the plasma; the parent compound was only found sporadically at very low levels. These results indicate removal by the liver and/or the gut at first pass. After evidence is found that the liver has the capacity to sulphoxidize and to sulphonize Albendazole, biotransformation of the gut was examined using an isolated perfused rat gut model. A high-performance liquid chromatography method was used to simultaneously determinate Albendazole, -sulphoxide and -sulphone. Albendazole was biotransformed partly to Albendazole sulphoxide by the gut, whereby the metabolite but not the parent compound was absorbed. It is concluded, that the gut has the capacity to biotransform Albendazole. However, biotransformation is limited to the first step, the sulphoxidation.

The protective effect of two silybin derivatives against phalloidin was tested in isolated rat livers, perfused with an erythrocyte free medium. Both liver swelling and potassium loss were examined for the evaluation of antagonism. Liver swelling and potassium loss were markedly decreased when silybin hemisuccinate was added to the perfusion medium before the application of phalloidin. In contrast, the protective effect of this compound was minimal after the mushroom toxin was partially absorbed by the liver. The protective efficiency of a dimer derivative (disilybin) was much greater than that of the monomer product. The binding of both derivatives in the liver cannot be very strong because the protective effect mainly could be washed out by a single change of the perfusion medium. Spectroscopical studies on isolated hepatocytes indicate that silybin inhibits the interaction of phalloidin with the cell surface.

Cholic acid binding to isolated rat liver plasma membranes was studied using a centrifugal filtration technique which allowed independent determination of free and membrane-bound cholic acid. Binding of cholic acid was very rapid and reversible. Scatchard analysis revealed at least three binding sites with high, medium and low affinity. The high affinity binding a) displayed saturability and isotope replacement, b) was not present in rat liver mitochondria and red blood cell ghosts and c) was temperature dependent. This binding has a very low capacity with a dissociation constant in the physiological range of plasma cholic acid concentration and has an affinity for other common bile acids. Cholic acid binding to the high affinity binding site was not inhibited by estrone, beta-estradiol or cholesterol. These results would suggest that the high affinity binding site represents a specific binding site for cholic acid and may also be specific for other common bile acids. This binding was not dependent on Na and was inhibited by bromosulfophthalein. Cholic acid binding to the high affinity site has some features in common with cholic acid uptake by isolated rat hepatocytes, and this would suggest that the high affinity binding site could be the postulated carrier for hepatic uptake of cholic acid.

Isolated perfused rat liver was used to characterize the choleretic action of 4-[4-chloro-N-(4-methoxyphenyl)-benzamido]-butyric acid (clanobutin). The dose-dependent increase of the bile flow was associated with an increase of the Na+ secretion. In in vitro studies the drug inhibited the Na+-K+-ATPase of the sinusoidal membrane more than that of the canalicular membrane in a dose-dependent manner. The following hypothesis was proposed to explain the choleretic action of clanobutin. The intracellular Na+ concentration increased due to the inhibition of the sinusoidal Na+-K+-ATPase. This is then compensated by transporting the extra Na+ into the bile. However, it cannot be ruled out that Na+ is transported as counterion during hepatobiliary transport of clanobutin or else its metabolites. The effect of the drug seems to be non-specific since other enzymes of the plasma membrane were also inhibited.

The age dependence of bile acid metabolism in rats was investigated and the following results were obtained. 1) Synthesis rate, pool size and total secretion decreased in old rats (24 month) significantly compared to young rats (6 weeks). 2) The quotient of taurine to glycine conjugates is is significant lower in old rats compared to young rats. 3) The lithogenic index remains unchanged in all age groups. 4) Old rats show a significantly higher Km for cholic acid uptake, whereas the diffusion coefficient is decreased. The possible influence of humoral factors on age dependent regulations of bile metabolism is discussed.

Cholic acid uptake was studied in isolated rat hepatocytes using a centrifugal filtration technique to allow rapid sampling. Hepatocytes were found to adsorb as well as to transport cholic acid. The adsorption was characterized by a capacity of 24 nmol X mg cell protein-1 and an association constant of 0.59 X 103 M-1. Cholic acid uptake was linear with respect to concentration at or below 10 degree C, suggesting a unsaturable uptake process which was considered to represent simple diffusion and is quantitated by a diffusion coefficient of 1.76 pmol cholic acid X min-1 X mg protein-1 X muM-1. Above 10 degrees C the uptake curve was biphasic. After subtracting the unsaturable component from uptake rates at higher temperatures, a curve showing saturable kinetics resulted. The apparent Km and V values at 37 degrees C were calculated to be 31muM and 0.8 nmol X min-1 X mg protein-1 respectively. This saturable uptake process was temperature-dependent with an activation energy of 13 kcal X mol-1 (5.44 X 104 J X mol-1) and was inhibited by oligomycin and KCN. Countertransport was demonstrated with cholic, taurocholic and chenodeoxycholic acids. The results suggest that cholic acid is transported by an energy-dependent carrier-mediated process in addition to simple diffusion by hepatocytes, and that the postulated carrier has affinity for other bile acids.

The influence of the choleretic drug methylumbilliferone on bile formation in the isolated perfused rat liver is characterized. The compound induces rapidly an elevation of bile flow, bile acid secretion and soium excretion. The increased production of bile is of canalicular origin. The choleretic effect was defined as "bile acid like" choleresis due to excretion of the drug into the bile. It is discussed that the excretion of methylumbilliferone can influence the transport of bile in form of a positive cooperation on transport mechanism.