Comparative analysis of volatile metabolites, quality and sensory attributes of Actinidia chinensis fruit.

Volatile organic compounds, quality and sensory parameters of four yellow- ('Dorì', 'G3', 'Jintao' and 'Soreli') and two green-fleshed ('Hayward' and 'Summer') kiwifruit cultivars were assessed. Statistical analysis was performed on volatiles, quality and sensory data for the identification of biomarkers of different kiwifruit cultivars. Principal component analysis showed that for all six samples a very good discrimination based on the cultivar was achieved. In particular, 2-pentylfuran can be used to distinguish between the green- and yellow-fleshed kiwifruit cultivars, while seven volatiles, can be identified as biomarkers of 'Dorì'. These findings are in agreement with the sensory analysis, which revealed that 'Dorì', the richest cultivar in esters, showed very high values of both ripe fruit smell and sweet sensory traits. Altogether, these results could offer recommendations for future breeding efforts for the production of kiwifruit cultivars with improved nutritional and aroma quality.

Volatile metabolites, quality and sensory parameters of "Ferrovia" sweet cherry cold stored in air or packed in high CO2 modified atmospheres.

Volatile organic compounds, quality and sensory attributes of sweet cherry cv "Ferrovia", cold packaged in Air or in different modified atmospheres (Low-O2 = 1% O2/0.03% CO2; High-CO2 = 16% O2/20% CO2; Mix = 1% O2/20% CO2), were monitored until 21 days of conservation. Results showed that sweet cherry cv "Ferrovia" is sensitive to CO2 accumulation (over 20%) in low oxygen (about 1%) modified atmosphere, as showed by the increase in respiration rate, biosynthesis of fermentative volatile metabolites, and sensory perception of off-odours. However, High-CO2 treatment seemed to preserve quality and sensory traits, presumably due to the high initial concentration of O2 (16%) that could limit the synthesis of ethyl esters and γ-butyrolactone, keeping the accumulation of off-flavours below their sensory perception threshold. Finally, PLSR analysis allowed to select 1-pentanol as putative marker of sensory alteration and hexanal and 2-hexenal as possible predictors of freshness for "Ferrovia" sweet cherries.

Comparison between open and arthroscopic-assisted foveal triangular fibrocartilage complex repair for post-traumatic distal radio-ulnar joint instability.

The aim of this study was to assess the objective and subjective functional outcomes after foveal reattachment of proximal or complete ulnar-sided triangular fibrocartilage complex lesions by two surgical procedures: an open technique or an arthroscopically assisted repair. The study was done prospectively on 49 wrists affected by post-traumatic distal radio-ulnar joint instability. Twenty-four patients were treated with the open technique (Group 1) and 25 by the arthroscopically assisted technique (Group 2). Magnetic resonance imaging demonstrated a clear foveal detachment of the triangular fibrocartilage complex in 67% of the cases. Arthroscopy showed a positive ulnar-sided detachment of the triangular fibrocartilage complex (positive hook test) in all cases. Distal radio-ulnar joint stability was obtained in all but five patients at a mean follow-up of 6 months. Both groups had improvement of all parameters with significant differences in wrist pain scores, Mayo wrist score, Disability of the Arm, Shoulder and Hand questionnaire and Patient-Rated Wrist/Hand Evaluation questionnaire scores. There were no significant post-operative differences between the two groups in the outcome parameters except for the Disability of the Arm Shoulder and Hand questionnaire score, which was significantly better in Group 2 (p < 0.001).

Cross-cultural adaptation and validation of the Italian version of the patient-rated wrist/hand evaluation questionnaire.

An Italian version of the patient-rated wrist/hand evaluation (PRWHE) questionnaire was obtained through the standardized process of cross-cultural adaptation. The PRWHE-Italian (IT) was tested on 63 patients in order to evaluate comprehension, reliability and validity as correlated to the validated version of the disabilities of the arm shoulder and hand (DASH)-IT and SF-36. No patients had difficulty completing the PRWHE-IT questionnaire. Psychometric testing demonstrated high reliability (Cronbach's alpha coefficient = 0.9607) and internal and external validity (Pearson correlation coefficient r = 0.927 with PRWHE, r < 0.810 with DASH and r < -0.476 with SF-36). The Italian version of the PRWHE has equivalent evaluation capacities to the original English version and is a reliable functional outcome measurement instrument for wrist and hand disorders.

Two novel human anti-ErbB2 immunoagents are active on trastuzumab-resistant tumours.

BACKGROUND: Overexpression of ErbB2 receptor in breast cancer is associated with disease progression and poor prognosis. Trastuzumab, the only humanised anti-ErbB2 antibody currently used in breast cancer, has proven to be effective; however, a relevant problem for clinical practice is that a high fraction of breast cancer patients shows primary or acquired resistance to trastuzumab treatment. METHODS: We tested on trastuzumab-resistant cells two novel human anti-tumour immunoconjugates engineered in our laboratory by fusion of a human anti-ErbB2 scFv, termed Erbicin, with either a human RNase or the Fc region of a human IgG1. Both Erbicin-derived immunoagents (EDIAs) are selectively cytotoxic for ErbB2-positive cancer cells in vitro and vivo, target an ErbB2 epitope different from that recognised by trastuzumab and do not show cardiotoxic effects. RESULTS: We report that EDIAs are active also on trastuzumab-resistant tumour cells both in vitro and in vivo, most likely because of the different epitope recognised, as EDIAs, unlike trastuzumab, were found to be able to inhibit the signalling pathway downstream of ErbB2. CONCLUSION: These results suggest that EDIAs are immunoagents that could not only fulfil the therapeutic need of patients ineligible to trastuzumab treatment due to cardiac dysfunction but also prove to be useful for breast cancer patients unresponsive to trastuzumab treatment.

A human, compact, fully functional anti-ErbB2 antibody as a novel antitumour agent.

A new human, compact antibody was engineered by fusion of a human, antitumour ErbB2-directed scFv with a human IgG1 Fc domain. Overexpression of the ErbB2 receptor is related to tumour aggressiveness and poor prognosis. This new immunoagent meets all criteria for a potential anticancer drug: it is human, hence poorly or not immunogenic; it binds selectively and with high affinity to target cells, on which it exerts an effective and selective antiproliferative action, including both antibody-dependent and complement-dependent cytotoxicity; it effectively inhibits tumour growth in vivo. Its compact molecular size should provide for an efficient tissue penetration, yet suitable to a prolonged serum half-life.

Identification of adulteration in water buffalo mozzarella and in ewe cheese by using whey proteins as biomarkers and matrix-assisted laser desorption/ionization mass spectrometry.

A rapid and accurate method to identify bovine and ewe milk adulteration of fresh water buffalo mozzarella cheese by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) is described. The differentiation among mozzarella made from water buffalo milk and from mixtures of less expensive bovine and, more recently, ewe milk with water buffalo milk is achieved using whey proteins, alpha-lactalbumin and beta-lactoglobulins as molecular markers. It is worth noting that the method proposed here is, to our knowledge, the first strategy able to characterize possible fraudulent additions of ewe milk in samples of water buffalo milk devoted to the production of water buffalo mozzarella cheese. In addition, a linear relationship was found between the relative response of the molecular ion and the abundance of the analysed whey proteins. This demonstrates that this approach can be used to determine the amount of bovine or ovine milk added to water buffalo milk employed for mozzarella cheese production. Furthermore, this method also appears suitable for the analysis of ewe cheese. Hence these findings open the way to a new field for mass spectrometry in the evaluation of possible fraudulence in dairy industry production.

Identification of adulteration in milk by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

The development is described of a rapid, simply and accurate analytical method aimed at evaluating both the presence of cow milk in either raw ewe and water buffalo milk samples employed in industrial processes and the addition of powdered milk to samples of fresh raw milk, using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). The presence of adulteration is defined by evaluating the protein patterns coming from the most abundant whey proteins, alpha-lactalbumin and beta-lactoglobulin, used as molecular markers. As no pretreatment of the milk samples is required and owing to the speed and ease of use of MALDI-MS the proposed analytical protocol can be used as a routine strategy for the identification of possible adulteration of the raw fresh milk samples that the dairy industry receives from producers every day.

Proteomics of gluten: mapping of subunit 1 Ax2* in Cheyenne cultivar by matrix-assisted laser desorption/ionization.

This paper reports results on the verification of the 1Ax2* high molecular weight glutenin subunit sequence in Cheyenne cultivar. The gene sequence of the protein is known but recently some text changes have been made, and furthermore until now no characterization of post-translational modifications has been reported. The two published sequences, named I and II, differ in four residues at positions 23, 208, 475, and 611. The first sequence contains 20 Arg and 6 Lys residues, producing 26 tryptic fragments, since the Arg(109)-Pro(110) bond is generally not cleaved by trypsin. The second sequence contains 19 Arg and 6 Lys residues, producing 25 tryptic peptides, again because of the Arg(109)-Pro(110) bond. Both sequences generate two cyanogen bromide fragments. Matrix-assisted laser desorption/ionization analysis of the tryptic digest of the high-MW glutenin subunit 1Ax2* resulted in the identification of 24 out of the 26 expected peptides for sequence I, a sequence coverage of 99.5%. These results were sufficient to rule out sequence II and any protein glycosylation and any other post-translational modifications to within the detection limits of the method. It was found that the choice of matrix considerably influenced the sequence coverage in peptide mapping.

Matrix-assisted laser desorption/ionization mass spectrometric peptide mapping of high molecular weight glutenin subunits 1Bx7 and 1Dy10 in Cheyenne cultivar.

This study describes the verification of the cDNA-deduced amino acid sequences of high molecular weight glutenin subunits 1Dy10 and 1Bx7 in Cheyenne cultivar by direct matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analysis of their tryptic fragments omitting chromatographic pre-separation. These polypeptides have a conserved structure consisting of a long central repetitive domain that prevents the application of conventional sequencing procedures such as Edman degradation. The published sequence of subunit 1Dy10 contains 7 Lys and 13 Arg residues; thus the production of 21 tryptic peptides is expected. The cDNA-deduced sequence for 1Bx7 subunit includes 5 Lys and 15 Arg residues, but the presence of three Arg-Pro bonds, which are normally not cleaved by trypsin, predicts only 19 tryptic peptides. Three different matrices (DHB, SA and HCCA) in combination with the most compatible sample preparation procedures were used in order to obtain the maximum 1Dy10 and 1Bx7 sequence coverage. MALDI analysis of the 1Dy10 tryptic digest resulted in the identification of all 21 expected peptides. In the case of 1Bx7 MALDI analysis resulted in the identification of 17 of the 19 expected peptides, giving a sequence coverage of 99.3%. These results were sufficient to rule out glycosylation of the 1Dy10 and 1Bx7 proteins and to assess the absence of any other post-translational modification, to within the detection limits of the method.

Sequencing of oligosaccharides by collision-induced dissociation matrix-assisted laser desorption/ionization mass spectrometry.

A study of the collision-induced dissociation post-source decay (PSD) spectra of free oligosaccharides is presented. These spectra, when obtained with helium as collision gas, show (1,5)X fragments containing the reducing end sugar. The presence of these fragments permits Y ions and, consequently, B and C peaks to be identified. This is a common behaviour from which it has been possible to delineate a general method for the easy assignment of the peaks in PSD spectra of underivatized neutral sugars, allowing the sequence of a real unknown to be obtained.

Studies on the primary structure of short polysaccharides using SEC MALDI mass spectroscopy.

The introduction of size-exclusion chromatography (SEC) analysis of polysaccharides prior to MALDI mass spectroscopy accounts for the determination of the molecular mass of the repeating unit when neutral homopolymers are investigated. In the case of natural polysaccharides characterised by more complicated structural features (presence of non-carbohydrate substituents, charged groups, etc.), this mass value usually is in agreement with more than one sugar composition. Therefore, it is not sufficient to give the correct monosaccharidic composition of the polysaccharide investigated. To solve this problem, MALDI spectra were recorded on the permethylated sample and post-source decay experiments were performed on precursor ions. In this way, the composition (in terms of Hex, HexNAc, etc.), size and sequence of the repeating unit were determined.

Social rank and sex-biased maternal investment in captive japanese macaques: behavioural and reproductive data.

This study investigated the relationship between social rank and sex-biased maternal investment in captive Japanese macaques (Macaca fuscata) using reproductive and behavioural data. High-ranking mothers showed a significantly male-biased secondary sex ratio, spent more time in contact with and carried male infants for longer than female infants. Low-ranking mothers showed no bias in secondary sex ratio nor in the time spent in contact with male and female infants, but carried female infants for longer. No differences were observed in the interbirth intervals following male and female infants nor in the frequency and intensity of aggression received by mothers with male and female infants, either in high-ranking or in low-ranking mothers. These results show that Japanese macaque mothers can adopt flexible and rank-dependent rearing strategies.

Use of hydroxyacetophenones as matrices for the analysis of high molecular weight glutenin mixtures by matrix-assisted laser desorption/ionization mass spectrometry

A selection of hydroxyacetophenones has been investigated as matrices for the analysis, by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), of high molecular weight (HMW) glutenin mixtures from three wheat varieties. Mass spectra were obtained directly from the HMW glutenin extracts without any preliminary purification and separation steps. According to the quality of the mass spectra, obtained using different hydroxyacetophenones, it has been possible to classify the matrices on the basis of their suitability for the analyte properties. Only two of the matrices considered showed to be compatible with the HMW glutenin mixtures analysis, although a large amount of other highly complex protein mixtures are present. This study indicates that MALDI-MS, as a stand-alone technique, is particularly useful for the direct determination of the complete HMW subunits profile and their molecular weights. Copyright 1999 John Wiley & Sons, Ltd.

A comparative study of two isoforms of laccase secreted by the "white-rot" fungus Rigidoporus lignosus, exhibiting significant structural and functional differences.

Two isoforms of laccase were obtained as the predominant phenol-oxidases in defined medium liquid cultures of the "white-rot" fungus Rigidoporus lignosus (R. lignosus). A characterization of the two laccases was made in terms of molecular mass, isoelectric point, metal content and N-terminal sequence. Furthermore, in order to gain information on the structural features related to the metal centers, a study of their geometric arrangement and their redox ability was made. It turned out that the two isoenzymes greatly differed with regard to pH stability, catalytic and copper centers features. It is proposed that all such differences are dependent on the amino acid sequences, which cause a distortion of the copper sites, thus accounting for the redox potential values and kinetic properties.

Analysis of human serum albumin variants by mass spectrometric procedures.

A new strategy for the structural characterisation of human albumin variants has been developed which makes extensive use of mass spectrometric methodologies. The rationale behind the method is to provide a rapid and effective screening of the entire albumin structure. The first step in this strategy consists in the attempt to determine the accurate molecular mass of the intact variant by electrospray mass spectrometry often providing a first indication on the presence of the variant. An HPLC procedure has been developed io isolate all the seven fragments generated by CNBr hydrolysis of HSA in a single chromatographic step. A rapid screening of the entire albumin structure is achieved by the ESMS analysis of the peptide fragments and the protein region(s) carrying the structural abnormality is identified by its anomalous mass value(s). Mass mapping of the corresponding CNBr peptide, either by Fast Atom Bombardment Mass Spectrometry (FABMS) or by Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDIMS), leads to the definition of the site and the nature of the variation. This combined strategy was applied to the structural characterisation of three HSA genetic variants and provided to be an effective procedure for the rapid assessment of their structural modifications showing considerable advantages over the classical approach.

Within- and between-strain variability in longevity of inbred and outbred rats under the same environmental conditions.

The analysis of 26 longevity curves of different populations of inbred (Fischer 344) and outbred (Sprague-Dawley) rats highlighted a remarkable between-populations variability in survival parameters. This variability is independent of the breeding characteristics of the strain. The two strains differed in the slope of the survival curves, with Fischer 344 rats showing a higher survival over the second year of life as well as a lower interindividual variability. A model-free approach based on principal component analysis allowed us to quantify these differences and to high-light some limitations of the classical Gompertzian approach.

Nonparametric scaling: a descriptive index.

A descriptive index (CI) based on the U statistic is presented. The CI permits to quantify the differences between experimental samples taking into account both the location and variability features of the data without any distributional assumption. This index can be useful to build activity scales for series of compounds.

Effects of concomitant nicotinic and muscarinic blockade on spatial memory disturbance in rats are purely additive: evidence from the Morris water task.

This study reexamined the role played by a concurrent manipulation of nicotinic and muscarinic acetylcholine (ACh) receptors on performance of rats in the Morris water maze. A series of experiments was performed to test decreasing doses of scopolamine, a muscarinic ACh blocker, given concurrently with a fixed dose level of mecamylamine, a nicotinic ACh blocker, down to a subthreshold combination. Both substances were also tested separately. Data were analyzed to distinguish between a summative and a greater than additive (synergistic) effect of the two blocking agents. Our results fully support the important role played by ACh systems on cognitive functions and also show the substantial functional independence of the two ACh receptors in regulating spatial learning processes. In fact, data analysis did not reveal any significant interaction between the two ACh receptor blockers other than their additive effect: the hypothesis of a reciprocal modulation between the two ACh receptors, raised by some authors, cannot be supported for spatial learning mechanisms, at least with regard to the Morris water maze paradigm.

Synthesis and amnesia-reversal activity of a series of 7- and 5-membered 3-acylamino lactams.

A series of 3-(acylamino)-epsilon-caprolactams and 3-(acylamino)-2-pyrrolidinones was synthesized. Some of these compounds reversed at different degrees electroconvulsive shock- and Scopolamine-induced amnesia, using a step-through passive avoidance in mice. Classical nootropic drugs, i.e., Aniracetam, Oxiracetam, and Piracetam, were used as reference compounds. Within the analyses of data performed, we introduced a new parameter, the confrontation index (CI), which is a function of Mann-Whitney's U statistic. The CI permits a common scale of activity of substances to be generated, independently of probabilistic hypotheses, with higher scores representing higher activities. The most active compounds were characterized by the formylamino and [3-(trifluoromethyl)benzoyl]amino groups in the 3-position of the ring. None of the substances assayed showed any effect on spontaneous behavior and neurovegetative system.