Real-time PCR for quantitative detection of chamois (Rupicapra rupicapra) and pyrenean ibex (Capra pyrenaica) in meat mixtures

A real-time quantitative polymerase chain reaction (PCR) technique was developed for the quantification of chamois and pyrenean ibex DNAs in meat mixtures by using a SYBR green detection platform. Two species-specific systems and a eukaryotic endogenous system were combined in the real-time PCR approach to quantify the target species. In the specific systems, a 133 base pair (bp) fragment of the 12S rRNA gene was amplified from chamois DNA, and an 88 bp fragment from the D-loop region was amplified from pyrenean ibex DNA. In the endogenous system, universal primers amplified a 141 bp fragment on the nuclear 18S rRNA gene from eukaryotic DNA. The threshold cycle values obtained with the 18S rRNA primers were used to normalize those obtained from chamois- or pyrenean ibex-specific systems, serving as endogenous control for the total content of PCR-amplifiable DNA in the sample. Analysis of experimental raw and heat-treated binary mixtures of chamois and pyrenean ibex meat in a swine meat matrix demonstrated the suitability of the assay for the detection and quantification of the target DNAs in the range of 0.1-0.8%, depending on the species and treatment of the meat samples.     

Analysis of mitochondrial DNA for authentication of meats from chamois (Rupicapra rupicapra), pyrenean ibex (Capra pyrenaica), and mouflon (Ovis ammon) by polymerase chain reaction-restriction fragment length polymorphism

The prevention of fraudulent labeling of game meat constitutes an important part of food regulatory control and quality assurance systems. A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis based on mitochondrial deoxyribonucleic acid (DNA) was developed for authentication of meats from chamois (Rupicapra rupicapra), pyrenean ibex (Capra pyrenaica), and mouflon (Ovis ammon). Amplification and restriction site analysis of a DNA fragment about 720 base pairs (bp) from the mitochondrial 12S rRNA gene of all analyzed species permitted the selection of Msel and Apol endonucleases for meat speciation. The 12S rRNA restriction profiles obtained allowed the unequivocal identification of chamois, pyrenean ibex, and mouflon/sheep and their differentiation from meats of domestic species such as cattle, goat, and swine. The highly variable mitochondrial D-loop gene was also targeted to attempt discrimination between mouflon and sheep meats. A D-loop region (700-1000 bp) was amplified and sequenced in all game and domestic species analyzed, and a primer set was designed for the selective amplification of a 370 bp DNA fragment from mouflon and sheep. PCR-RFLP analysis with the selected Maell enzyme generated a single electrophoretic profile characteristic for sheep, whereas 3 different fragment patterns were obtained for mouflon meats. Consequently, the PCR-RFLP technique developed can be routinely applied in inspection programs in order to verify the correct labeling of game species.     

Reconstruction of the isotopic history of animal diets by hair segmental analysis

Carbon and nitrogen isotope signatures (delta(13)C and delta(15)N) of animal tissues provide information about the diet and, hence, the environment in which the animals are living. Hair is particularly useful as it provides a stable archive of temporal (e.g. seasonal) fluctuations in diet isotope composition. It can be sampled easily and with minimal disturbance from living subjects. However, derivation of the temporal record along the hair length may be subject to errors and uncertainties. This study investigates (and suggests means to minimize) several sources of error, including (a) incomplete sampling, (b) sampling during the quiescent (telogen) phase, (c) non-representative sub-sampling, (d) ignorance of hair growth rate, i.e. time-position relationship of isotope signatures, and (e) non-optimal compromise between analytical/procedural precision and effort/cost. Cattle tail switch hair was collected from animals of different breed, sex and age. Hair was washed, sectioned, and 5- or 10-mm-long sections were analyzed for C and N isotope composition. Signatures along paired hairs were similar (r(2) approximately 0.8) and distances between isotopic minima and maxima nearly identical, indicating that a single hair constituted a representative sample and (except for telogen hair) hair growth rate was the same for paired hairs. However, cutting hair, instead of plucking, caused a variable loss of recently grown hair and information. Telogen hair was identified and data loss due to cutting error reduced when more than one hair from the same animal and sampling region was compared to spot and delimit common and missing regions. Similarly, comparison of isotopic profiles from hair collected at different times identified the segment produced during the respective interval and allowed calculation of average hair growth rate, which varied between animals (0.69-1.06 mm d(-1)). Analysis of alternate 10-mm-long sections for two hairs per animal provided a good compromise between precision/resolution and effort. The method should be applicable to other mammalian species including man.     

A history of thermochemistry through the tribulations of its devotees

The history and the evolution of thermochemistry are covered in a number of excellent books. In order not to repeat what has already been written, this paper will be concerned with thermochemists as persons through their odd stories and anecdotes, of hot, enduring and even fierce debates, of accidents in the laboratory. This revised version was published online in July 2006 with corrections to the Cover Date.     

Temperature dependencies of high-temperature reduction on conversion products and their isotopic signatures

High-temperature reduction (HTR) is widely used for oxygen and hydrogen isotope determination. Decomposition of cellulose, sucrose and polyethylene foil by HTR is quantitative for temperatures around 1450 degrees C. For lower reaction temperature production of CO(2), water and the deposition of carbon inside the reactor are significant and thus the element of interest for isotopic analysis is split into different pools, leading to isotope fractionation. After reduction of cellulose or sucrose at 1125 degrees C less than 60% of the oxygen is found as CO, which is monitored with the isotope ratio mass spectrometer to determine the delta(18)O value. The remaining oxygen is unevenly distributed between CO(2) and H(2)O, preferentially as CO(2). Raising the reaction temperature to 1425 degrees C yields almost quantitative conversion of oxygen into CO and results in a 3 per thousand more positive delta(18)O value. Similarly, only 40-50% of the carbon of cellulose and sucrose is transformed into CO in the HTR reactor at 1125 degrees C. This is far from the stoichiometric expected value of 83% for quantitative carbon transfer for cellulose and 92% for sucrose. Of the carbon 40-50% is deposited in the reactor and the remainder can be found as CO(2). Based on the comparison of carbon isotope results from HTR and those obtained from combustion, we hypothesize that CO produced during the HTR originates partly from sample carbon and glassy carbon. A combined combustion and HTR carbon isotope determination may provide an insight into the intramolecular carbon distribution of organic substances. These results suggest that HTR should be carried out at temperatures above 1450 degrees C to make sure that fractionations associated with the reduction process are minimal. If this is not possible frequent calibration is required using reference materials of the same structure as the sample.     

Radiobarites from the Cenozoic volcanic region of the Bohemian Massif: radiochemical study,history, and lead isotopic composition

A specific, radium bearing type of barites has been known from the Ohře (Eger) Rift, a region of Cenozoic volcanic activity in northwestern Bohemia, Czech Republic, since 1904. Barites from Karlovy Vary, Teplice and Děčín areas have been studied to elucidate geochemical history of these specific samples. Low background radiometric method using high-resolution gamma-ray spectrometry has shown that radioactivity of these barites (up to 8 Bq g⁻¹) is carried by ²²⁶Ra, which is not in equilibrium with the parent ²³⁸U. In all samples, uranium activity was below 0.005 Bq g⁻¹––the detection limit achieved by instrumental neutron activation analysis. The alpha track method on a crystal plate has shown that ²²⁶Ra is accumulated in surface layers or crystal cracks, therefore, the crystal rims must be relatively young, not older than 10–15 thousand years. In a lead fraction isolated from the barites, concentration and isotopic composition of lead were determined by photon activation analysis and ICP MS, respectively.     

Development of non-destructive isotopic analysis methods using muon beams and their application to the analysis of lead

Journal of Radioanalytical and Nuclear Chemistry - Elemental isotopic ratios are measured in various research fields and provide useful information regarding age, origin, geological and biological...     

Extraction of lithium from GSJ rock reference samples and determination of their lithium isotopic compositions

Li was extracted completely from the Geological Survey of Japan (GSJ) rock reference samples by HF-HC1 decomposition and separated from other alkali metal ions by cation exchange chromatography. The degree of Li isotope fractionation during the preparations of samples for mass spectrometry for Li isotopic ratio measurements was, therefore, negligible. The isotopic compositions of Li extracted from the GSJ samples were measured by double-filament surface ionization mass spectrometry. The permil (%.) deviation, δ⁷Li, of the ⁷Li-to-⁶Li ratio of one of the GSJ samples (JR-1) from that of the Li isotope standard, IRM-016, was +3.9%., with a 95% confidence limit of 0.7%. The δ⁷Li values of the 16 GSJ samples varied in the range −2.5 to +6.4%.. These data are probably the first such data obtained for the GSJ rock samples. No clear dependence of the δ⁷Li value on the kind of rock was observed.     

Extraction of boron from GSJ rock reference samples and determination of their boron isotopic ratios

The extraction of boron from rocks by methyl borate distillation after alkali fushion was studied. Reproducible extraction was obtained when the ratio of the amounts of flux and rock was ?5:1. Isotope dilution experiments showed there was no appreciable boron isotope fractionation during the extraction under these conditions. ¹¹B/¹⁰B ratios for three standard rocks were 4.044–4.047.     

A role of the 9-aminoacridines and their conjugates in a life science

The 9-aminoacridines play an important role in medicine. They were applied first in a treatment of protozoal infections in the beginning of the last century. Recently, it has been shown that the 9-aminoacridines are successful candidates for treatment of cancer, viral and prion diseases. Their conjugation with biomolecules such as peptides and proteins may modulate their activity, bioavailability and applicability. This review deals with the synthesis of 9-aminoacridine, its conjugation with variety of molecules and utilization of such conjugates in several fields of science.     

Thionin-sensitized intrazeolite photooxygenation of trisubstituted alkenes: substituent effects on the regioselectivity as probed through isotopic labeling

The regioselectivity for the intrazeolite photooxygenation of several trisubstituted alkenes with geminal dimethyl groups was examined. The length of the alkyl chain at the lone position was varied, and as end groups, the phenyl or the cyclohexyl functionalities were chosen. The general trend for all alkenes is a significant increase of the reactivity at the twin position compared to the photooxygenation in solution. For the cyclohexyl-substituted alkenes, it was found that the regioselectivity is nearly independent of the alkyl chain length. However, for the phenyl-substituted alkenes, the ene reactivity of the allylic methylene hydrogen atoms at the lone position and the twix/twin regioselectivity depend significantly on the distance of the phenyl group from the double bond. These trends are discussed in terms of cation-pi interactions and conformational effects. Intramolecular and intermolecular isotope effects in the intrazeolite photooxygenation of deuterium-labeled alkenes suggest that a perepoxide-type intermediate is formed in the rate-determining step. Type I photooxygenation that involves reaction of the radical cations of the alkenes with superoxide ion are unlikely.     

Assessment of hydrophobic interactions and their contributions through the analysis of the methane dimer

Hydrophobic Interactions (HIs) are important in many phenomena of molecular recognition in chemistry and biology. Still, the relevance of HIs is sometimes difficult to evaluate particularly in large systems and intramolecular interactions. We put forward a method to estimate the magnitude and the different contributions of a given HI of the C···C, H? C···H, and H···H type through (i) the analysis of the electron density in the intermolecular region for eleven relative orientations of the methane dimer and (ii) the subsequent decomposition of the corresponding interaction energy in physically significant contributions using Symmetry Adapted Perturbation Theory (SAPT). Strong correlations were found between the topological properties of calculated at intermolecular bond critical points and plus its different contributions with the C···C distance of the considered orientations of (CH₄)₂. These correlations were used to construct Mollier‐like diagrams of and its components as a function of the separation between two carbons and the orientation of the groups bonded to these atoms. The ethane dimer and tert‐butylcyclohexane are used as representative examples of this new approach. Overall, we anticipate that this new method might prove useful in the study of both intramolecular and intermolecular HIs particularly of those within large systems wherein SAPT or electronic structure calculations are computationally expensive or even prohibitive. © 2014 Wiley Periodicals, Inc.     

Explorations of conical intersections and their ramifications for chemistry through the Jahn-Teller effect

Much recent progress has been made theoretically and computationally towards understanding the importance of conical intersections for chemical reactions. Nonetheless, experimental characterization of conical intersections has proven extremely difficult with one striking exception: the Jahn-Teller conical intersection. This article overviews the fundamental similarity of a variety of conical intersections and demonstrates how the spectroscopy of Jahn-Teller active molecules can be used to characterize them. Specific results are reviewed for four representative Jahn-Teller active molecules, C5H5, C6H6+, Ag3 and CH3O.     

Simple long-range model and scaling relations for the binding of isotopic hydrogen atoms to isotopic helium surfaces

A simple long-range model for the very weak binding of isotopic hydrogen atoms on isotopic helium surfaces is proposed. The one parameter is fixed by the empirical binding energy of ³He on a ⁴He surface. This simple model predicts binding energies in good agreement with recent experimental and theoretical results.     

Analysis of α-keratins in the horns of rhinoceros and buffalo by non-native capillary isoelectric focusing

Summary A capillary isoelectric focusing (CIEF) method under denaturing condition has been developed for separation of α-keratins, highly cross-linked biomacromolecules, in horns of rhinoceros and buffalo. The α-keratins were denatured in 8 M urea and separated in the presence of ampholytes with applicable pH ranges of 3–7 or 3–10. In the preliminary results, it was demonstrated that the α-keratins in the horns of buffalo or rhinoceros might have their own unique isoelectric focusing profiles. The analyses of more samples from different rhinoceros and buffalo are presently underway, with the final goal to establish a fast and convenient method for identification of the source (buffalo or rhinoceros) of an unknown powdered sample.     

The effect of acidification on the determination of organic carbon, total nitrogen and their stable isotopic composition in algae and marine sediment

We investigated the effects of sample acidification on the stable carbon and nitrogen isotopic composition (delta13C and delta15N), as well as the organic carbon (OC) and total nitrogen (TN) composition, of an algal culture and a marine sediment. Replicate measurements of untreated and acid-treated samples were made using 1 M, 2 M and 6 M HCl, 6% H2SO3 and 1 M H3PO4. For all treatments the precision of the analysis for the acid-treated sample was equal to or less than that in the non-acidified sample. For the algae, analysis of variance (ANOVA) indicated no significant differences in the mean OC and TN concentration, or delta13C and delta15N composition, between any acid treatment and non-acidified samples. For the sediment sample a comparison could only be made between the different acid treatments because the untreated contained significant amounts ( approximately 30%) of carbonate carbon. ANOVA indicated that the mean OC determined in sediment samples after the 1 M HCl treatment and the mean delta13C values after the 6% H2SO3 and 1 M H3PO4 treatments were significantly different (p < 0.013 and < .05, respectively) from all other treatments. Mass balance calculations indicate that in some instances delta13C values were biased due to a contribution from unreacted carbonate carbon. There were no significant differences in the mean TN between any acid-treated and non-acidified samples. The mean delta15N values after 6 M HCl, 6% H2SO3 and 1 M H3PO4 treatments were significantly different from the untreated sediment sample (p < 0.044). Based on the significant bias observed for the delta15N and delta13C values, a weak (1-2 M) HCl solution is confirmed as the most appropriate acid for the removal of inorganic carbon from natural materials requiring elemental and isotopic analysis.     

Isolation of peganine and deoxypeganine from the total alkaloids ofPeganum harmala through their complex salts

Abtsract The possibility has been shown of combining the processes of the reduction of deoxyvasicinone and vasicinone to deoxypeganine and peganine and the isolation of the latter in the form of complex salts from the natural total mixture of alkaloids.Institute of the Chemistry of Plant Substances, Academy of Sciences of the Republic of Uzbekistan, Tashkent, fax (3712) 89 14 75. Translated from Khimiya Prirodnykh Soedinenii, No. 1, pp. 70–73, January–February, 1996. Original article submitted April 5, 1994.