Assessment of enzymatic methods in the δ18O value determination of the L-tyrosine p-hydroxy group for proof of illegal meat and bone meal feeding to cattle.

The δ(18)O value of the p-hydroxy group of L-tyrosine depends on the biosynthesis by plants or animals, respectively. In animal proteins it reflects the diet and is therefore an absolute indicator for illegal feeding with meat and bone meal. The aim of this investigation was to perform the positional (18)O determination on L-tyrosine via a one-step enzymatic degradation. Proteins from plants, herbivores, omnivores, and carnivores were characterized by their δ(13)C, δ(15)N, and δ(18)O values, the latter for normalizing the positional δ(18)O values. Their L-tyrosine was degraded by tyrosine phenol lyase to phenol, analyzed as (2,4,6)-tribromophenol. Degradation by tyrosine decarboxylase yielded tyramine. The δ(18)O values of both analytes corresponded to the trophic levels of their sources but were not identical, probably due to an isotope effect on the tyrosine phenol lyase reaction. Availability of the enzyme, easy control of the reaction, and isolation of the analyte are in favor of tyrosine decarboxylase degradation as a routine method.

Essential methodological improvements in the oxygen isotope ratio analysis of N-containing organic compounds.

The quantitative conversion of organically bound oxygen into CO, a prerequisite for the (18)O/(16)O analysis of organic compounds, is generally performed by high-temperature conversion in the presence of carbon at ∼1450°C. Since this high-temperature procedure demands complicated and expensive equipment, a lower temperature method that could be utilized on standard elemental analyzers was evaluated. By substituting glassy carbon with carbon black, the conversion temperature could be reduced to 1170°C. However, regardless of the temperature, N-containing compounds yielded incorrect results, despite quantitative conversion of the bound oxygen into CO. We believe that the problems were partially caused by interfering gases produced by a secondary decomposition of N- and C-containing polymers formed during the decomposition of the analyte. In order to overcome the interference, we replaced the gas chromatographic (GC) separation of CO and N(2) by reversible CO adsorption, yielding the possibility of collecting and purifying the CO more efficiently. After CO collection, the interfering gases were vented by means of a specific stream diverter, thus preventing them from entering the trap and the mass spectrometer. Simultaneously, a make-up He flow was used to purge the gas-specific trap before the desorption of the CO and its subsequent mass spectrometric analysis. Furthermore, the formation of interfering gases was reduced by the use of polyethylene as an additive for analytes with a N:O ratio greater than 1. These methodological modifications to the thermal conversion of N-containing analytes, depending on their structure or O:N ratio, led to satisfactory results and showed that it was possible to optimize the conditions for their individual oxygen isotope ratio analysis, even at 1170°C. With these methodological modifications, correct and precise δ(18)O results were obtained on N-containing analytes even at 1170°C. Differences from the expected standard values were below ±1‰ with standard deviations of the analysis <0.2‰.

delta(34)S-value measurements in food origin assignments and sulfur isotope fractionations in plants and animals.

The delta(34)S values of biological material, especially food commodities, serve as indicators for origin assignments. However, in the metabolism of higher plants sulfur isotope fractionations must be expected. As a matter of fact, the delta(34)S values of the sulfate- and organic-S, respectively, of Brassicaceae and Allium species vegetables showed differences between 3 and 6 per thousand, and differences in glucosinolates were between 0 and 14 per thousand. delta(34)S-value differences of total-S between individual tissues of the same plant were approximately 3 per thousand. It is believed that these relatively small and variable fractionations are due to the partition of individual S-metabolism steps to different plant compartments, where they may occur independently and quantitatively. The delta(34)S values of herbivore muscle meat and milk relative to the diet and between an animal and its child had trophic shifts of approximately 1.5 per thousand. (34)S enrichments of up to 4 per thousand were observed for hair, hooves, and horn, an isotope fractionation of -5 per thousand between the diet sulfate and cartilage. Therefore, the reported agreements between delta(34)S value of biomass and primary S sources are true for only bulk material and not for individual compounds or tissues.

A measuring system for the fast simultaneous isotope ratio and elemental analysis of carbon, hydrogen, nitrogen and sulfur in food commodities and other biological material.

The isotope ratio of each of the light elements preserves individual information on the origin and history of organic natural compounds. Therefore, a multi-element isotope ratio analysis is the most efficient means for the origin and authenticity assignment of food, and also for the solution of various problems in ecology, archaeology and criminology. Due to the extraordinary relative abundances of the elements hydrogen, carbon, nitrogen and sulfur in some biological material and to the need for individual sample preparations for H and S, their isotope ratio determination currently requires at least three independent procedures and approximately 1 h of work. We present here a system for the integrated elemental and isotope ratio analysis of all four elements in one sample within 20 min. The system consists of an elemental analyser coupled to an isotope ratio mass spectrometer with an inlet system for four reference gases (N(2), CO(2), H(2) and SO(2)). The combustion gases are separated by reversible adsorption and determined by a thermoconductivity detector; H(2)O is reduced to H(2). The analyser is able to combust samples with up to 100 mg of organic material, sufficient to analyse samples with even unusual elemental ratios, in one run. A comparison of the isotope ratios of samples of water, fruit juices, cheese and ethanol from wine, analysed by the four-element analyser and by classical methods and systems, respectively, yielded excellent agreements. The sensitivity of the device for the isotope ratio measurement of C and N corresponds to that of other systems. It is less by a factor of four for H and by a factor of two for S, and the error ranges are identical to those of other systems.