A promising 31P NMR-multivariate analysis approach for the identification of milk phosphorylated metabolites and for rapid authentication of milk samples

A fast and reliable method for the identification of milk from different mammalians was developed by using ³¹P NMR metabolite profile of milk serum coupled to multivariate analysis (PCA and classification models UNEQ, SIMCA and K-NN). Ten milk samples from six different mammalians, relevant to human nutrition (human, cow, donkey, mare, goat, sheep), were analyzed and eight monophosphorylated components were identified and quantified: phosphocreatine (PCr), glycerophosphorylcholine (GPC), glycerophosphorylethanolamine (GPE), N-acetylglucosamine-1-phosphate (NAcGlu-1P), lactose-1-phosphate (Lac-1P), galactose-1-phosphate (Gal-1P), phosphorylcholine (PC), glucose-6-phosphate (Glu-6P). PCA showed interesting clustering based on the animal genus. K-NN can be successfully used to discriminate between donkey and cow samples while UNEQ class-modeling resulted more suitable for compliance verification. Results confirm the natural variability of milk samples among different species. These data highlight the great potentials of NMR/multivariate analysis combined method in the rapid analysis of phosphorylated milk serum metabolites for milk origin assessment and milk adulteration detection.     

Predictive markers of safety and immunogenicity of adjuvanted vaccines

Vaccination represents one of the greatest public health triumphs; in part due to the effect of adjuvants that have been included in vaccine preparations to boost the immune responses through different mechanisms. Although a variety of novel adjuvants have been under development, only a limited number have been approved by regulatory authorities for human vaccines. This report reflects the conclusions of a group of scientists from academia, regulatory agencies and industry who attended a conference on the current state of the art in the adjuvant field. Held at the U.S. Pharmacopeial Convention (USP) in Rockville, Maryland, USA, from 18 to 19 April 2013 and organized by the International Association for Biologicals (IABS), the conference focused particularly on the future development of effective adjuvants and adjuvanted vaccines and on overcoming major hurdles, such as safety and immunogenicity assessment, as well as regulatory scrutiny. More information on the conference output can be found on the IABS website, http://www.iabs.org/.     

Kinematic and electromyographic analysis of the Nordic Hamstring Exercise

The Nordic Hamstring Exercise (NHE) has been introduced as a training tool to improve the efficiency of eccentric hamstring muscle contraction. The aim of this study was to perform a biomechanical analysis of the NHE. Eighteen participants (20.4 ± 1.9 years) performed two sets of five repetitions each of the NHE and maximal eccentric voluntary contraction (MEVC) of the knee flexors on an isokinetic dynamometer whilst knee angular displacement and electrical activity (EMG) of biceps femoris were measured. EMG was on average higher during the NHE (134.3% of the MEVC). During the forward fall of the NHE, the angle at which a sharp increase in downward velocity occurred varied between 47.9 and 80.5 deg, while the peak knee angular velocity (pVelocity) varied between 47.7 and 132.8 deg s^?1. A significant negative correlation was found between pVelocity and peak EMG (r = ?0.62, p < 0.01) and EMG at 45 deg (r = ?0.75, p < 0.01) expressed as a percentage of peak MEVC EMG. Some of the variables analyzed exhibited good to excellent levels of intra- and inter-session reliability. This type of analysis could be used to indirectly monitor the level of eccentric strength of the hamstring muscles while performing the NHE and potentially any training- or injury-related changes.     

A new kumamolisin-like protease from Alicyclobacillus acidocaldarius: an enzyme active under extreme acidic conditions

A new serine-carboxyl proteinase, called kumamolisin-ac, was purified from the thermoacidophilic bacterium Alicyclobacillus acidocaldarius. The enzyme is a monomeric protein of 45?kDa, active over a wide temperature range (5.0–70°C) and extremely acidic pHs (1.0–4.0), showing maximal proteolytic activity at pH?2.0 and 60°C. Interestingly, kumamolisin-ac displayed a significant proteolytic activity even at 5°C, thus suggesting a sort of cold-adaptation for this enzyme. The protease was remarkably stable at high temperatures (t_1/2 at 80°C, 10?h, pH?2.0) and over a broad range of pH (2.0–7.0). Substrate analysis indicated that kumamolisin-ac was active on a variety of macromolecular substrates, such as haemoglobin, hide powder azure, and azocoll. In particular, a high specific activity was detected towards collagen. The corresponding gene was cloned, expressed and the recombinant protease, was found to be homologous to proteases of the ‘S53’ family. From the high identity with kumamolisin and kumamolisin-As, known as collagenolytic proteases, kumamolisin-ac can be considered as the third collagenolytic affiliate within the ‘S53’ family. Cleavage specificity investigation of kumamolisin-ac revealed a unique primary cleavage site in bovine insulin B-chain, whereas a broad specificity was detected using bovine α-globin as substrate. Thus, kumamolisin-ac could represent an attractive candidate for industrial-scale biopeptide production under thermoacidophilic conditions.     

Cyanidin-3- O -β-glucopyranoside Protects Myocardium and Erythrocytes from Oxygen Radical-mediated Damages

The cyanidin-3- O - &#103 -glucopyranoside (C-3-G) antioxidant capacity towards reactive oxygen species (ROS)-mediated damages was assessed in tissue and cells submitted to increased oxidative stress. In the isolated ischemic and reperfused rat heart, 10 or 30 &#119 M C-3-G protected from both lipid peroxidation (66.7 and 94% inhibition of malondialdehyde (MDA) generation in 10 and 30 &#119 M C-3-G-reperfused hearts, respectively, in comparison with control reperfused hearts) and energy metabolism impairment (higher ATP concentration in 10 and 30 &#119 M C-3-G-reperfused hearts than in control reperfused hearts). These effects were associated to C-3-G permeation within myocardial cells, as indicated by results obtained in the isolated rat heart perfused for 30 min in the recirculating Langendorff mode under normoxia with 10 and 30 &#119 M C-3-G. Protective effects were exerted, in a dose-dependent manner, by C-3-G also in 2 mM hydrogen peroxide-treated human erythrocytes. With respect to MDA formation, an apparent IC 50 of 5.12 &#119 M was calculated for C-3-G (the polyphenol resveratrol used for comparison showed an apparent IC 50 of 38.43 &#119 M). The general indications are that C-3-G (largely diffused in dietary plants and fruits, such as pigmented oranges very common in the Mediterranean diet) represents a powerful natural antioxidant with beneficial effects in case of increased oxidative stress, and at pharmacological concentrations it is able to decrease tissue damages occurring in myocardial ischemia and reperfusion.     

Molecular evidence for hybrid origin of Quercus crenata Lam. (Fagaceae) from Q. cerris L. and Q. suber L.

Abstract

Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeats (ISSR) markers were employed to examine samples from Quercus cerris, Q. suber and Q. crenata in order to test the hypothesis of the hybrid origin of Q. crenata from Q. cerris and Q. suber in a part of its distribution area where the two putative parents do not overlap at present. Leaves from 21 Q. crenata and 37 Q. cerris individual trees were collected at natural sites in northern Italy, where Q. suber is currently lacking; 21 Q. suber and six Q. crenata plants from central Italy were also analysed. Results from Unweighted Pair Group Method with Arithmetic mean (UPGMA) analysis and principal component analysis (PCA) implied that exchange of neutral markers has been considerable between the three species, while differences in morphological characters have remained comparatively stable. The Mantel test indicated low correlation between RAPD- and ISSR-based similarity matrices, showing that the two screening techniques reveal unrelated estimates of genetic relationships. Hybrid indices computed for both markers displayed an intermediate position of Q. crenata individuals between the two putative parents shifted toward Q. cerris. Results from the present study corroborate the hypothesis of a hybrid origin for Q. crenata occurring in northern Italy, and suggest asymmetrical backcrossing with Q. cerris acting as the recurrent parent.     

Irpexil (Methyl-acethydroxamoilacetate)-induced changes in some membrane functions and transmembrane potential in wheat seedling root segments

Abstract

A strong K⁺-dependent, fusicoccin-sensitive increase in acidic extrusion paralleled by a corresponding increase in net K⁺ uptake and hyperpolarization of transmembrane electrical potential was observed in the root segments of wheat seedlings cultured in diluite CaSO₄+0.27 mM Methyl-acethydroxamoilacetate. Moreover, K⁺-induced depolarization was also increased up to three times.     

Modification of chromatin organisation at low water potential in cultured cells of Solanum tuberosum: Possible involvement of dehydrins

ABSTRACT

To understand the mechanisms which enable the nucleus to function under low water potential, the morphology and biochemistry of potato cell nuclei were studied. Conformational modifications were observed in the chromatin of nuclei of cells growing under low water potential. These modifications include a higher number of heterochromatic centres, enlargement of the nuclear diameter, and a different accessibility of DNA to the action of restriction enzymes. Biochemical analyses showed that these chromatin modifications may coincide with quantitative and qualitative variations of several nuclear proteins, some of which may belong to the dehydrin family. We especially focussed our attention on a 45-kDa protein that is heat-stable and is recognised by an antibody raised against the conserved domain of dehydrins. The survival of potato cells in an environment where water availability is low may depend on several simultaneous events regarding the nucleus. The accumulation in the nucleus of specific proteins such as dehydrins could be required to stabilise the chromatin by means of their molecule-salvation action. Further studies are in progress to check whether or not variations in chromatin organisation may be one of the numerous traits that a cell must acquire to become water-stress resistant.