Development and validation of a LC-MS/MS method for the simultaneous determination of aflatoxins, dyes and pesticides in spices.

Based on several alerts from European countries over the last years concerning spices, we have been encouraged to establish an accurate method for the determination of dyes, aflatoxins and pesticides in various types of spices using reversed-phase (RP) liquid chromatography-tandem mass spectrometry interfaced with electrospray (LC-ESI-MS/MS). A simple sample treatment procedure entailing the use of an extraction step with acetonitrile without further cleanup has been developed. A C18 column with an aqueous ammonium formate/methanol mixture as the mobile phase was used, and gradient elution was performed. Mass spectral acquisition was done in positive ion mode by applying multiple reaction monitoring of at least two fragmentation transitions per compound to provide a high degree of selectivity. The method was in-house validated in terms of linearity, sensitivity, repeatability, recovery and selectivity on six kinds of spices. Satisfactory results in the majority of the cases were obtained for all analytes and matrices, with practical limits of quantitation acceptable for routine monitoring purposes. Extraction recoveries for most of the compounds ranged from 60% to 140% at spiking levels of 0.05 and 0.5 mg kg(-1). The applicability of the method for the simultaneous determination of dyes, aflatoxins and pesticides in several types of spices was demonstrated, and the method successfully applied to a limited number of products from the local market.

Increased expression of extracellular proteins as a hallmark of human endothelial cell in vitro senescence.

A convenient way to study processes of aging in distinct human tissues consists of a molecular analysis of cells from the tissue in question, that were explanted and grown in vitro until they reach senescence. Using human umbilical vein endothelial cells (HUVEC), we have established an in vitro senescence model for human endothelial cells. A major hallmark of HUVEC in vitro senescence is the increased frequency of apoptotic cell death, which occurs as a determining feature of HUVEC senescence. Senescent endothelial cells are also found in vivo in atherosclerotic lesions, suggesting that the presence of such cells may contribute to the development of vascular pathology. To elucidate mechanisms underlying endothelial cell senescence and age-associated apoptosis, gene expression analyses were carried out. In these experiments, we observed the up-regulation of genes coding for extracellular proteins in senescent HUVEC. In particular, a significant upregulation of interleukin-8, VEGI, and the IGF-binding proteins 3 and 5 was observed. Upregulation of these genes was confirmed by both RT-PCR and Western blot. In the case of interleukin-8, a roughly 50-fold upregulation of the protein was also found in cellular supernatants. The extracellular proteins encoded by these genes are well known for their ability to modulate the apoptotic response of human cells, and in the case of interleukin-8, clear links to the establishment of atherosclerotic lesions have been defined. The results described here support a new model, where changes in the secretome of human endothelial cells contribute to vascular aging and vascular pathology.

High-resolution respirometry--a modern tool in aging research.

Alterations in mitochondrial function are believed to play a major role in aging processes in many species, including fungi and animals, and increased oxidative stress is considered a major consequence of altered mitochondrial function. In support of this theory, a lot of correlative evidence has been collected, suggesting that changes in mitochondrial DNA accumulate with age in certain tissues. Furthermore, genetic experiments from lower eukaryotic model organisms, indicate a strong correlative link between increased resistance to oxidative stress and an extended lifespan; in addition, limited experimental evidence suggests that the inhibition of mitochondrial function by selected pharmacologically active compounds can extend lifespan in certain species. However, changes in mitochondrial function may affect aging in a different way in various tissues, and a clear statement about the role of mitochondrial deterioration during physiological aging is missing for most if not all species. At this point, respirometric analyses of mitochondrial function provide a tool to study age-associated changes in mitochondrial respiratory chain function and mitochondrial ATP production within living cells and isolated mitochondria. In the recent years, new instruments have been developed, which allow for an unprecedented high-resolution respirometry, which enables us to determine many parameters of mitochondrial function in routine assays using small samples of biological material. It is conceivable that this technology will become an important tool for all those, who are interested in experimentally addressing the mitochondrial theory of aging. In this article, we provide a synopsis of traditional respirometry and the advances of modern high-resolution respirometry, and discuss how future applications of this technology to recently established experimental models in aging research may provide exciting new insights into the role of mitochondria in the aging process.

[On the diagnostics and rehabilitation of stenosing processes of cerebral vessels (author's transl)]. / Zur Diagnostik und Rehabilitation stenosierender Hirngefässprozesse.

In rehabilitation, patients suffering from stenosing processes of cerebral vessels represent a particular target group with particularly difficult diagnostic and therapeutic problems. On the basis of own experiences it was possible to test on 25 persons (age: 37 to 83 years) suffering from carotid and vertebral stenosis the capability of the cerebral vessels echography--alone and in combination with the DOPPLER flowmetry--:. This highly valuable cerrebrovascular screening method coupled with clinical findings makes it possible to give a more definite indication for angiography, and represents under certain circumstances (inoperability or undue strain in the presence of a reduced general and vascular status) a safe alternative to invasive methods. The modern programs involving in-patient rehabilitation and, in particular, vasoactive, blood pressure stabilising and cardiac protection accentuating long-term treatment were all successful--notwithstanding the sometimes very severe and prognostically unfavorable neurological involvement, the frequent associated internistic diseases, the risk factors and the intercurrent complications: All rehabilitants (age: 37 to 73 years) were discharged partly or fully mobile (6 in each case) following on average a 55-day stay in this ward. In each case it was possible to achieve a distinct improvement in the cerebral functional disorders, the subjective state and general condition, as well as a high degree of social reintegration.