Feeding cows with hay, silage, or fresh herbage on pasture or indoors affects sensory properties and chemical composition of milk and cheese.

In European countries, silage-free feeding is an ancient tradition and has a particularly positive reputation among consumers. In the present study, we compared grass-based forages from the same plot conserved as hay or silage or fed fresh either on pasture or indoors, and we evaluated the differences in sensory properties of milk and uncooked pressed cheese. All herbage from the first cut of a grassland dominated by perennial ryegrass was harvested on the same day and preserved either as hay or silage. The first regrowth of the same plot was used for strip grazing or green feeding indoors. Balanced by breed, 24 Montbéliarde and 24 Holstein cows were allocated to the 4 treatments. Apart from the forages, the late-lactation cows received 3 kg/d of dry matter from concentrate. After 2 wk of dietary adaptation, the bulk milk of 3 subgroups, each with 4 cows, was collected. Part of the milk was pasteurized, and part was left raw and partly transformed to small-sized Cantal-type cheese ripened for 9 wk. Milk and cheese underwent descriptive sensory analysis by a trained sensory panel, as well as analyses of physicochemical traits. Volatile organic compounds of the cheeses were also analyzed. Raw and pasteurized milk from hay-fed cows had less intense odors of cooked milk, cream, and barnyard than milk from grazing cows, whereby the effect of pasteurization did not differ between herbage utilization methods. Cheeses obtained from cows fed fresh herbage (grazing and indoors) were clearly yellower than cheeses from silage- and hay-fed cows, which coincided with the color intensity perceived by the panelists. Moreover, cheeses from cows fed fresh herbage had more intense barnyard and dry fruit flavors, were perceived as creamier and having less lactic odor, and exhibited more fat exudation than those from cows fed conserved herbage. Only a few differences were observed in milk and cheeses from hay-fed compared with silage-fed cows, and those differences were far less pronounced than those of milk and cheeses from cows fed fresh herbage. In conclusion, the present study did not substantiate assumptions of clear sensory differences of milk and uncooked pressed cheese from hay-fed compared with silage-fed cows. For the first time, this study reports that the global flavor intensity of cheeses from indoor green-fed cows is similar to that of cheeses derived from cows fed conserved forages, whereas cheeses from grazing cows have the greatest global flavor intensity.

Claimed effects, outcome variables and methods of measurement for health claims proposed under European Community Regulation 1924/2006 in the framework of protection against oxidative damage and cardiovascular health.

BACKGROUND AND AIMS: The high number of negative opinions from the European Food Safety Authority (EFSA) to the requests for authorization of health claims is largely due to the design of human intervention studies, including the inappropriate choice of outcome variables (OVs) and of their methods of measurement (MMs). The present manuscript reports the results of an investigation aimed to collect, collate and critically analyse the information in relation to claimed effects, OVs and MMs, in the context of protection against oxidative damage and cardiovascular health compliant with Regulation 1924/2006. METHODS AND RESULTS: Claimed effects, OVs and the related MMs were collected from EFSA Guidance documents and applications for authorization of health claims under Articles 13.5 and 14. The OVs and their MMs were evaluated only if the claimed effect was sufficiently defined and was considered beneficial by EFSA. The collection, collation and critical analysis of the relevant scientific literature consisted in the definition of the keywords, the PubMed search strategies and the creation of databases of references. The critical analysis of the OVs and their MMs was performed on the basis of the literature review and was aimed at defining the appropriateness of OVs and MMs in the context of the specific claimed effects. CONCLUSIONS: The information provided in this document could serve to EFSA for the development of further guidance on the scientific requirements for health claims, as well as to the stakeholders for the proper design of human intervention studies aimed to substantiate such health claims.

Characterization of the volatile profile of thistle honey using headspace solid-phase microextraction and gas chromatography-mass spectrometry.

In this study, a headspace solid-phase microextraction method was developed for the characterization of the volatile fraction of thistle honey and compared with a dynamic headspace extraction method. A DVB/CAR/PDMS fibre was used. The effects of extraction time, equilibration time and salt addition on extraction yield were evaluated. The volatile fraction of seven Italian thistle honey samples was extracted under the optimized conditions and analyzed by gas chromatography-mass spectrometry. Characterization of the volatile profile was performed in terms of nature and relative amount of the extracted compounds. A total of 40 compounds, belonging to different chemical classes, were identified. The relative amounts of 16 compounds found in all the analyzed thistle honeys, i.e. nonanal, furfural, decanal, 3,6-dimethyl-2,3,3a,4,5,7a-hexahydrobenzofuran, benzaldehyde, α-linalool, lilac aldehyde (isomer IV), hotrienol, phenylacetaldehyde, 4-oxoisophorone, benzyl alcohol, 2-phenylethanol, a not identified compound, octanoic acid, nonanoic acid and methyl anthranilate, were calculated and submitted to statistical analysis, in order to define for each compound a typical range. On the basis of the obtained data, a characteristic set of values was defined for thistle honey volatile fingerprint. The developed model proved to be effective in recognizing the botanical origin of thistle honey.

Surface expression of a heterologous phosphatase complements CD45 deficiency in a T cell clone.

Expression of CD45, the major transmembrane protein tyrosine phosphatase expressed on lymphoid cells, is required for optimal T cell receptor (TCR) signal transduction. We and others recently have demonstrated that surface expression of the cytoplasmic domain of CD45 in the absence of its extracellular and transmembrane domains is sufficient to restore TCR-mediated signaling events in CD45-deficient cell lines. Here we demonstrate that a single domain nonreceptor tyrosine phosphatase from yeast expressed as a chimeric protein with the extracellular and transmembrane domains of a major histocompatibility complex class I molecule also is able to restore proximal and distal TCR-mediated signal transduction events in the CD45-deficient T cell line J45.01. Ligation of the TCR on the cell line expressing the yeast phosphatase chimera results in the induction of protein tyrosine kinase activity, soluble inositol phosphate generation, and expression of the CD69 activation antigen. Furthermore, a phosphatase-inactive version of this molecule is unable to restore signal transduction, providing the first formal evidence that plasma membrane associated tyrosine phosphatase activity is required for TCR-mediated signaling.

Rescue of signaling by a chimeric protein containing the cytoplasmic domain of CD45.

Surface expression of the CD45 tyrosine phosphatase is essential for the T cell antigen receptor (TCR) to couple optimally with its second messenger pathways. CD45 may be required to dephosphorylate a TCR-activated protein tyrosine kinase, which then transduces an activation signal from the TCR. A chimeric molecule that contained extracellular and transmembrane sequences from an allele of a major histocompatibility class I molecule and cytoplasmic sequences of CD45 restored TCR signaling in a CD45-deficient mutant T cell line. Thus, expression of the complex extracellular domain of CD45 is not required for the TCR to couple to its signaling machinery.

Experimental design for the optimization of the extraction conditions of polycyclic aromatic hydrocarbons in milk with a novel diethoxydiphenylsilane solid-phase microextraction fiber.

An innovative solid-phase microextraction coating based on the use of diethoxydiphenylsilane synthesized by sol-gel technology was used for the determination of polycyclic aromatic hydrocarbons at trace levels in milk. The effects of time and temperature of extraction and acetone addition were investigated by experimental design. Regression models and desirability functions were applied to find the experimental conditions providing the highest global extraction response. The capabilities of the developed fiber were proved obtaining limit of quantitation values in the low microg/l range, enabling the direct analysis of complex matrices like milk and a complete desorption of high-boiling compounds without carryover effects.

Development and validation of an inductively coupled plasma mass spectrometry method with optimized microwave-assisted sample digestion for the determination of platinum at ultratrace levels in plasma and ultrafiltrate plasma.

A highly sensitive inductively coupled plasma mass spectrometry (ICP-MS) method with microwave-assisted sample digestion for the determination of total platinum in rat whole and ultrafiltrate plasma was developed and validated. A first step of this study concerned the optimization of the mineralization procedure, in order to obtain good extraction recovery (higher than 90%) and repeatability (less than 6%) and the absence of matrix effect. ICP-MS analysis was then performed using the "hot plasma/protective ion extraction" mode, achieving high sensitivity and very high signal/noise ratio. Iridium was added as internal standard. The method was then submitted to validation, performed according to the FDA Bioanalytical Validation Methods guidelines and to the Eurachem guide. Validation was carried out in terms of limit of detection (LOD), limit of quantitation (LOQ), linearity, precision, accuracy and stability. An instrumental LOQ of 1.9ngL(-1), corresponding to a concentration of 955ngL(-1) in matrix under the adopted conditions, was obtained, allowing the quantitative analysis of Pt ultratraces. Instrumental linearity was verified in the range 1.9-14,000ngL(-1), corresponding to a concentration range from 955ngL(-1) to 6825microgL(-1) in matrix. Accuracy was evaluated by analyzing control samples for both matrices at different concentration levels; a good agreement (<15%) was obtained. Sample stability was tested by analyzing control samples maintained for 4h at room temperature or submitted to three freezing-thawing cycles. Finally, the developed method was applied to the analysis of plasma and ultrafiltrate plasma of rats treated with oxaliplatin-base drug, thus demonstrating its reliability in pharmacokinetic studies.

Tyrosine phosphorylation of Grb2-associated proteins correlates with phospholipase C gamma 1 activation in T cells.

Ligation of the T-cell antigen receptor (TCR) results in the rapid activation of several protein tyrosine kinases, with the subsequent phosphorylation of numerous cellular proteins. We investigated the requirement for tyrosine phosphorylation of proteins which bind the Grb2 SH2 domain in TCR-mediated signal transduction by transfecting the Jurkat T-cell line with a cDNA encoding a chimeric protein designed to dephosphorylate these molecules. Stimulation of the TCR on cells expressing this engineered enzyme fails to result in sustained tyrosine phosphorylation of a 36-kDa protein likely to be the recently cloned pp36/Lnk. Interestingly, TCR ligation of the transfected cells also fails to induce soluble inositol phosphate production and intracellular calcium mobilization, although receptor-mediated tyrosine phosphorylation of phospholipase C gamma 1 still occurs. TCR-mediated Ras and mitogen-activated protein kinase activation remain intact in cells expressing the engineered phosphatase. These data demonstrate that tyrosine phosphorylation of a protein(s) which binds the SH2 domain of Grb2 correlates with phospholipase C gamma 1 activation and suggest that such a phosphoprotein(s) plays a critical role in coupling the TCR with the phosphatidylinositol second-messenger pathway.

Porcine follicular fluids: comparison of solid-phase extraction and matrix solid-phase dispersion for the GC-MS determination of hormones during follicular growth.

The capabilities of solid-phase extraction (SPE) and matrix solid-phase dispersion (MSPD) for the determination of the hormones 17beta-estradiol, 2-hydroxyestradiol, 4-hydroxyestradiol and 2-methoxyestradiol by gas chromatography-mass spectrometry (GC-MS) in a very complex matrix like porcine follicular fluids were compared, thus proving the highest effectiveness of the SPE technique. Validation was carried out in terms of limit of quantitation (LOQ), precision, accuracy, recovery and stability. LOQ values in the low microg kg(-1) were achieved, with all the other parameters satisfying the acceptance criteria for the validation of bioanalytical methods. The applicability of the method to the determination of the hormones in porcine follicular fluids was demonstrated, thus allowing to observe an increase of the concentration of the hormones during the follicular growth.

Transient sinus node dysfunction after the Cox-maze III procedure in patients with organic heart disease and chronic fixed atrial fibrillation.

OBJECTIVES: This prospective study examined types, frequency and time dependency of the electrophysiologic manifestation of the sinus node dysfunction after the Cox-maze III procedure--the technique of choice for the management of medically refractory atrial fibrillation-in patients with organic heart disease, chronic fixed atrial fibrillation and no preoperatively overt dysfunction of the sinus node. BACKGROUND: The original maze procedure was modified twice in order to reduce the high incidence of the sinus node inability to generate an appropriate sinus tachycardia in response to maximal exercise, and occasional left atrial dysfunction. Despite these modifications, postoperative disturbance of sinus node function can be frequently observed. METHODS: In 15 adult patients, standard electrocardiogram, 24-h Holter monitoring, power spectral analysis of heart variability, exercise testing, Valsalva maneuver and rapid positional changes were performed 3, 6 and 12 months after the Cox-maze III procedure and mitral valve surgery or closure of atrial septal defect. RESULTS: Electrocardiographic manifestations of sinus node dysfunction were identified in 12 patients at 3 months, in 6 patients at 6 months, and in 0 patients at 12 months after surgery. The heart rate response to exercise during the first 6 months was reduced in the maze group and became fully normal at 12 months. Power spectral analysis of heart rate variability showed very low power values at 1 month with inhibited cardiac autonomic activity and no response on sympathetic stress. A potential of recovery of cardiac autonomic activity was documented 12 months after surgery. CONCLUSIONS: The manifestations of sinus node dysfunction following the Cox-maze III procedure were time dependent and their frequency and intensity progressively decreased and disappeared within 12 months after surgery.

Medium-term results of heart transplantation using donors over 63 years of age.

BACKGROUND: The continual shortage of hearts for transplantation (HTx) led to the expansion of the donor pool by accepting older donors. We compared the medium-term follow-up of patients after HTx with older hearts (over the age of 63 years) with those of patients after HTx with younger hearts. PATIENTS AND METHODS: Since April 1994 we have used hearts for HTx from donors older than the age of 63 years. Until November 1998, 309 HTx and 9 re-HTx were performed in 309 adults with a mean age of 50.7+/-10.9 years (range 17-68 years). There were 252 men and 57 women. The patients were divided into two groups: group I--donor age under 63 years (296 patients, mean age 50.4+/-11 years; mean donor age 38.1+/-13 years; mean follow-up 1.7+/-1.6 years); group II-donor age of more than 63 years (13 patients, mean age 57.4+/-5.6 years; mean donor age 65.1+/-2.1; mean follow-up 2.2+/-1.6 years). There were no differences in the etiology of heart failure, gender, or ischemia time between the groups. The patients in group II were significantly older (P = 0.008). Multiple factors were analyzed in the groups, which included changes in the left/right ventricle ejection fraction, early postoperative mortality (up to 30 days), cumulative survival rates and cardiac-dependent morbidity [myocardial infarction, malignant arrhythmias, coronary stenosis (>50% in one of the main coronary arteries) and transplant vasculopathy]. Additionally, freedom from cytomegalovirus infection (rise of titer or seroconversion) and freedom of acute rejection episodes grade > or =2 (International Society of Heart & Lung Transplantation [ISHLT]) were analyzed. RESULTS: After 1 year mean left and right ventricle ejection fraction were good in both groups and did not significantly change for up to 2 years. No Re-HTx was performed in group II. The early postoperative mortality was similar in both groups (P = 0.8). Also, the cumulative survival rates were similar in both groups (P = 0.87). Long-term cardiac morbidity was lower in group I (P = 0.03). The long-term freedom from cytomegalovirus infection in group I was significantly higher when compared with group II (P = 0.0002). The long-term freedom from severe rejection episodes was similar in both groups (P = 0.3) CONCLUSION: The study found a significant increase in long-term cardiac morbidity due to more focal coronary stenosis in group II, and freedom from cytomegalovirus infection, but did not find significant differences in the long-term survival between patients who received hearts from donors of up to 63 years of age and from those more than 63 years. The acceptance of donors older than 63 years old for HTx does not worsen the outcome of the recipients. The careful selection of older donors, with close monitoring of the coronary situation after HTx and expanded indications for revascularization of older hearts, could make HTx with older hearts, even in older recipients, a safe option.

Effects of subcutaneous recombinant IL-2 on humoral immunity in advanced cancer-patients.

Immunotherapy represents a promising biological approach to cancer treatment. Important haemato-immunological modifications during rIL-2 treatment have been demonstrated in all patients treated, independently of the schedule and mode of rIL-2 administration. Generally, there is a lack of correlation between treatment-related modifications and clinical response. rIL-2 cell-mediated responses have been extensively studied but the role of humoral immunity is still unclear. We evaluated the peripheral blood CD4, CD8, CD19 and CD25 lymphocyte subset modifications and serum levels of immunoglobulin, IL-6 and IL-2 in 19 patients with malignant melanoma and renal cell carcinoma undergoing subcutaneous administration of IL-2. Our results suggest that humoral immunity is only marginally involved; however, the parameter variations observed suggest that further studies are necessary.

Evaluation of phagocytic-activity in neoplastic patients treated with subcutaneous recombinant IL-2.

Immunostimulating and immunosuppresive events have been noted during rIL-2 immunotherapy and cell-mediated immune deficiencies have been reported in the neutrophil and monocyte functions, determining a high incidence of infections in patients during intravenous rIL-2 treatment. The phagocytic activity of monocytes/macrophages and granulocytes was evaluated in 10 advanced solid tumor patients treated with subcutaneous rIL-2. Several indirect parameters of phagocytosis were also considered, such as Neopterin, Beta2 microglobulin, IL-2 soluble receptor, and the C3 and C4 fractions of the complement system. A decreased phagocytosis was demonstrated after subcutaneous rIL-2 administration together with an increase of indirect parameters of granulocyte/macrophage activity. Therefore, cellular activation does not seem to correspond to actual phagocytosis which is probably due to IL-2-induced secondary cytokines or to rIL-2 inhibitory effects.

Subcutaneous ril-2 in advanced melanoma and kidney carcinoma.

The aim of the present study was to verify the efficacy and tolerability of a treatment protocol using subcutaneous administration of rIL-2. Eighteen patients with metastatic disease were treated: 10 with renal carcinoma and 8 with malignant melanoma. The drug was administered as follows: 9 million UI/mq twice daily for two days followed by 1.8 million UI/mq twice daily for five days/week for six consecutive weeks. Of the 15 patients evaluated for clinical response, all completed treatment without rIL-2 dose modifications. No complete response was obtained. One patient with malignant melanoma and 1 with renal carcinoma (13%) had partial response (soft tissue and lymph nodes respectively) which lasted 3 and 4 months. Six patients with renal carcinoma and 3 melanoma patients had stable disease (60%) which lasted 6 months (median). Toxicity was moderate and spontaneously reversing after stopping treatment. Haemato-immunological modifications and pharmacokinetic study of this modality of rIL-2 treatment were also examined.

The Cox maze iii procedure: parallel normalization of sinus node dysfunction, improvement of atrial function, and recovery of the cardiac autonomic nervous system.

OBJECTIVE: The Cox maze III procedure includes isolation of the pulmonary veins and multiple incisions in both atria in what corresponds to partial autotransplantation and partial denervation of the heart. The aim of this prospective longitudinal study was to identify physiologic effects of reinnervation on changes in heart rate at rest and in response to various stimulations and on atrial function after the Cox maze III procedure. PATIENTS AND METHODS: Power spectral analysis of heart rate variability, exercise testing, 24-hour Holter monitoring, electrocardiography, and transthoracic and transesophageal echocardiography were performed in 30 adult patients after the combined Cox maze III procedure and mitral valve surgery (maze group). They were prospectively followed up at 1, 3, 6, and 12 months after the operation. The results were compared with those of 15 heart transplant recipients (transplant group) and normal probands (healthy adults, n = 12). RESULTS: The physiologic effects of denervation with no differences in cardiac autonomic activity between the groups were seen early after the operation. Later, evidence of autonomic reinnervation was observed only in the maze group but not in the transplant group. Inappropriate heart rate responses during physical exercise were clearly evident in both groups after 1 and 3 months, with progressive improvement seen between 6 and 12 months only in the maze group. Left atrial function after the Cox maze procedure improved parallel to the recovery of sinus node function. CONCLUSION: Progressive improvement of sinus node function and atrial contractions with significant functional normalization 1 year after the Cox maze procedure corresponded to functional reinnervation and recovery of the autonomic nervous system.

Management of dystocia caused by a large sacrococcygeal teratoma.

Sacrococcygeal teratoma is a rare cause of dystocia. Reported management strategies to date have yielded few viable infants. In this case report, the infant was partially delivered when extraction was halted. The infant was intubated and resuscitated while preparations for emergency cesarean section were being made. The infant was subsequently delivered abdominally and three years later is normal. Despite absent thoracic cage movement during resuscitation, adequate oxygenation was possible. Increasing use of prenatal ultrasonography should allow early detection, and thus avoid such unanticipated dystocias.

Surgical treatment of a chest-wall penetrating left ventricular pseudoaneurysm.

This report describes the treatment of a patient who developed a chest-wall penetrating pseudoaneurysm 3 years after coronary bypass grafting and after the resection of a lateral wall left ventricular aneurysm twice. The patient presented with a pulsatile tumor in the left submammilar region. Surgery was done in deep hypothermia, with femoro-femoral cannulation and via a left anterolateral thoracotomy. The perioperative course was uneventful and the patient is still well 5 years after surgery.

Identification of P waves after the Cox-maze procedure: significance of right precordial leads V3R through V6R.

BACKGROUND: The maze circuit lengthens the period of atrial depolarization and may lead to small or absent P waves, which may not be readily apparent in a standard 12-lead electrocardiogram. In this prospective study, we investigate if the right precordial leads V3R through V6R can improve detection of P waves and identification of atrial electrical activity. METHODS: Standard 12-lead electrocardiograms (with leads I through III, aVR, aVL, aVF, and V1 through V6) and right precordial electrocardiograms (V3R through V6R) were recorded in 30 consecutive patients during the first 5 postoperative days and 1, 3, 6, 12, and 24 months after combined Cox-maze III procedure and mitral valve surgery. The P wave identification as an indication of atrial electrical activity and atrial contraction was proved by transesophageal echocardiographic identification of atrial contractions. RESULTS: Despite echocardiographically identifiable atrial contractions, the P waves were not visible on standard 12-lead electrocardiograms in 7 (23%) patients after 1 and 3 months, and in 4 patients after 6 months. In contrast, right precordial electrocardiograms showed P waves in all of these patients that were best seen in lead V4R. Twelve and 24 months postoperatively, only 2 patients had no visible P waves in the standard leads that were still present in the right precordial leads. However, if P waves were easily identifiable in standard 12-lead electrocardiograms (23 patients at 1 and 3 months after surgery), the right precordial leads showed P waves only in 15 patients. CONCLUSIONS: Right precordial electrocardiogram with leads V3R through V6R is a helpful tool for visualizing P waves after the Cox-maze procedure. Standard 12-lead electrocardiogram should be combined with right precordial electrocardiogram in all patients after the Cox-maze procedure.

The biocompound method in coronary artery bypass operations: surgical technique and 3-year patency.

BACKGROUND: Complete arterial revascularization may be unsafe in patients with a high operative risk. In patients with varicose ectatic veins, the biocompound technique, which uses unsuitable autologous veins, enables the surgeon to influence the bypass graft wall stress levels and diameter. This report summarizes the 3-year patency of 53 patients, the survival rate of 200 patients, and operative technical considerations. METHODS: Biocompound grafts were used for aortocoronary bypass in 200 patients who were considered inappropriate subjects for complete arterial revascularization and who had unsuitable saphenous veins. RESULTS: The mortality rate (30 days) of 200 patients was 3.5%. The 3-year survival rate was 88.5%. The patency rate of the left internal thoracic artery (LITA) after 3 years was 97.3%, of the native vein was 68.7%, and of the biocompound graft was 68.3%. The LITA showed a superior patency rate (p = < 0.05). CONCLUSIONS: The LITA is the first choice in coronary bypass operation. The biocompound technique is a reliable method to achieve complete revascularization in patients with a lack of suitable saphenous veins.

Applications of liquid chromatography-mass spectrometry interfacing systems in food analysis: pesticide, drug and toxic substance residues.

This paper reviews applications of different LC-MS techniques for the determination of xenobiotic substances in foods. Specific examples of contaminants discussed are pesticides, herbicides, insecticides and drugs; concerning toxic substances, mycotoxins, phycotoxins, cyanobacterial toxins, mutagenic and heterocyclic amines and beta-carbolines, arsenic, tin and inorganic halogen compounds, packaging materials and various epoxy resins are considered. Advantages and limitations are outlined for the different LC-MS interfacing systems (particle beam, thermospray, atmospheric pressure ionization with electrospray, ionspray and heated pneumatic nebulizer). The impact of developments in instrumental analysis on methodology and the limitations of the various LC-MS methods are discussed. Further, the coupling of LC with element-selective detection systems such as inductively coupled plasma mass spectrometry is discussed, with emphasis on speciation of trace toxic elements in foods.