Characterisation of PDO olive oil Chianti Classico by non-selective (UV-visible, NIR and MIR spectroscopy) and selective (fatty acid composition) analytical techniques.

An authentication study of the Italian PDO (protected designation of origin) extra virgin olive oil Chianti Classico was performed; UV-visible (UV-vis), Near-Infrared (NIR) and Mid-Infrared (MIR) spectroscopies were applied to a set of samples representative of the whole Chianti Classico production area. The non-selective signals (fingerprints) provided by the three spectroscopic techniques were utilised both individually and jointly, after fusion of the respective profile vectors, in order to build a model for the Chianti Classico PDO olive oil. Moreover, these results were compared with those obtained by the gas chromatographic determination of the fatty acids composition. In order to characterise the olive oils produced in the Chianti Classico PDO area, UNEQ (unequal class models) and SIMCA (soft independent modelling of class analogy) were employed both on the MIR, NIR and UV-vis spectra, individually and jointly, and on the fatty acid composition. Finally, PLS (partial least square) regression was applied on the UV-vis, NIR and MIR spectra, in order to predict the content of oleic and linoleic acids in the extra virgin olive oils. UNEQ, SIMCA and PLS were performed after selection of the relevant predictors, in order to increase the efficiency of both classification and regression models. The non-selective information obtained from UV-vis, NIR and MIR spectroscopy allowed to build reliable models for checking the authenticity of the Italian PDO extra virgin olive oil Chianti Classico.

Monitoring of alcoholic fermentation using near infrared and mid infrared spectroscopies combined with electronic nose and electronic tongue.

Effective fermentation monitoring is a growing need due to the rapid pace of change in the wine industry, which calls for fast methods providing real time information in order to assure the quality of the final product. The objective of this work is to investigate the potential of non-destructive techniques associated with chemometric data analysis, to monitor time-related changes that occur during red wine fermentation. Eight micro-fermentation trials conducted in the Valtellina region (Northern Italy) during the 2009 vintage, were monitored by a FT-NIR and a FT-IR spectrometer and by an electronic nose and tongue. The spectroscopic technique was used to investigate molecular changes, while electronic nose and electronic tongue evaluated the evolution of the aroma and taste profile during the must-wine fermentation. Must-wine samples were also analysed by traditional chemical methods in order to determine sugars (glucose and fructose) consumption and alcohol (ethanol and glycerol) production. Principal Component Analysis was applied to spectral, electronic nose and electronic tongue data, as an exploratory tool, to uncover molecular, aroma and taste modifications during the fermentation process. Furthermore, the chemical data and the PC1 scores from spectral, electronic nose and electronic tongue data were modelled as a function of time to identify critical points during fermentation. The results showed that NIR and MIR spectroscopies are useful to investigate molecular changes involved in wine fermentation while electronic nose and electronic tongue can be applied to detect the evolution of taste and aroma profile. Moreover, as demonstrated through the modeling of NIR, MIR, electronic nose and electronic tongue data, these non destructive methods are suitable for the monitoring of must-wine fermentation giving crucial information about the quality of the final product in agreement with chemical parameters. Although in this study the measurements were carried out in off-line mode, in future these non destructive techniques could be valid and simple tools, able to provide in-time information about the fermentation process and to assure the quality of wine.

Minimisation of instrumental noise in the acquisition of FT-NIR spectra of bread wheat using experimental design and signal processing techniques.

Spectral resolution (R) and number of repeated scans (S) have a significant effect on the S/N ratio of Fourier transform-near infrared (FT-NIR) spectra, but the optimal values of these two parameters have to be determined empirically for a specific problem, considering separately both the nature of the analysed matrix and the specific instrumental setup. To achieve this aim, the instrumental noise of replicated FT-NIR spectra of wheat samples was modelled as a function of R and S by means of the Doehlert design. The noise amounts in correspondence to different experimental conditions were estimated by analysing the variance signals derived from replicate measurements with two different signal processing tools, Savitzky-Golay (SG) filtering and fast wavelet transform (FWT), in order to separate the "pure" instrumental noise from other variability sources, which are essentially connected to sample inhomogeneity. Results confirmed that R and S values leading to minimum instrumental noise can vary considerably depending on the type of analysed food matrix and on the different instrumental setups, and helped in the selection of the optimal measuring conditions for the subsequent acquisition of a wide spectral dataset.

Evaluation and predictive modeling of shelf life of minced beef stored in high-oxygen modified atmosphere packaging at different temperatures.

The aims were: (1) to follow the freshness decay of minced beef stored in high-oxygen modified atmosphere packaging at different temperatures (4.3, 8.1 and 15.5 degrees C) by applying traditional methods (microbiological counts, color evaluation, thiobarbituric acid assay TBA, headspace gas composition) and e-nose; (2) to model the decay kinetics to obtain information about the maximum shelf life as function of storage conditions. The minced beef, packaged in modified atmosphere was supplied by a manufacturer at the beginning of its commercial life. The study demonstrated the ability of the traditional methods to describe the kinetics of freshness decay. The modeling of the experimental data and the comparison with microbiological or chemical thresholds allowed the setting, for each index, of a stability time above which the meat was no longer acceptable. The quality decay of meat was also evaluated by the headspace fingerprint of the same set of samples by means of a commercial e-nose. A clear discrimination between "fresh" and "old" samples was obtained using PCA and CA, determining at each temperature a specific range of stability time. The mean value of the stability times calculated for each index was 9 days at 4.3 degrees C (recommended storage temperature), 3-4 days at 8.1 degrees C (usual temperature in household refrigerators) and 2 days at 15.5 degrees C (abuse temperature). Resolution of the stability times allowed calculation of mean Q(10) values, i.e. the increase in rate for a 10 degrees C increase in temperature. The results show that the Q(10) values from the traditional methods (3.6-4.0 range) overlapped with those estimated with e-nose and color indexes (3.4 and 3.9, respectively).

Shelf life of Crescenza cheese as measured by electronic nose.

The shelf life of Crescenza, a traditional Italian soft cheese, was measured using classical analysis and a commercial electronic nose. Two lots of samples directly supplied by a manufacturer at the beginning of their commercial life were stored at 2 constant temperatures (8 and 15 degrees C) and analyzed until their respective expiration dates. Among the physicochemical parameters, pH, acidity, hue, and apparent yield rheological index appeared to be the best predictors of the quality decay. Changes in these indices were described with a sigmoidal transition function allowing definition of a loose and a severe shelf-life protocol, based on the trend of first and second time derivatives. A time range of 1 to 3 d at 15 degrees C and 4 to 8 d at 8 degrees C was accordingly assessed to maintain the freshness of Crescenza cheese. The quality decay of cheese aroma was evaluated by inspecting the headspace fingerprint of the same set of samples using the electronic nose. Sample classification through the aroma fingerprint confirmed the predicted shelf-life time ranges. A clear discrimination between "fresh," "aged," and "very aged" samples was obtained using principal components analysis, cluster analysis, and linear discriminant analysis statistical techniques. The predictive ability of the linear discriminant analysis classification model was confirmed by considering a new set of cheese samples purchased at the beginning of their commercial life from a local market and analyzed until their expiration date.

Hepatic fibrogenesis requires sympathetic neurotransmitters.

BACKGROUND AND AIMS: Hepatic stellate cells (HSC) are activated by liver injury to become proliferative fibrogenic myofibroblasts. This process may be regulated by the sympathetic nervous system (SNS) but the mechanisms involved are unclear. METHODS: We studied cultured HSC and intact mice with liver injury to test the hypothesis that HSC respond to and produce SNS neurotransmitters to promote fibrogenesis. RESULTS: HSC expressed adrenoceptors, catecholamine biosynthetic enzymes, released norepinephrine (NE), and were growth inhibited by alpha- and beta-adrenoceptor antagonists. HSC from dopamine beta-hydroxylase deficient (Dbh(-/-)) mice, which cannot make NE, grew poorly in culture and were rescued by NE. Inhibitor studies demonstrated that this effect was mediated via G protein coupled adrenoceptors, mitogen activated kinases, and phosphatidylinositol 3-kinase. Injury related fibrogenic responses were inhibited in Dbh(-/-) mice, as evidenced by reduced hepatic accumulation of alpha-smooth muscle actin(+ve) HSC and decreased induction of transforming growth factor beta1 (TGF-beta1) and collagen. Treatment with isoprenaline rescued HSC activation. HSC were also reduced in leptin deficient ob/ob mice which have reduced NE levels and are resistant to hepatic fibrosis. Treating ob/ob mice with NE induced HSC proliferation, upregulated hepatic TGF-beta1 and collagen, and increased liver fibrosis. CONCLUSIONS: HSC are hepatic neuroglia that produce and respond to SNS neurotransmitters to promote hepatic fibrosis.

Clearance kinetics of hepatitis C virus under different antiviral therapies.

Interferon alpha (IFN) has been the standard treatment for hepatitis C virus (HCV) infection. Using the kinetic curves of viral clearance, this study compared three treatment regimes based on IFN alone or in combination with Amantadine or Ribavirin to determine the mechanisms of action and the most suitable way to use these drugs. The early clearance kinetics of HCV were studied in 22 patients with chronic hepatitis C under different antiviral treatments: IFN 3 MU daily (7 pts); IFN 3 MU daily plus Amantadine 200 mg (7 pts); and IFN 3 MU daily plus Ribavirin 1-1.2 gr (8 pts), for 6 months. HCV-RNA was assessed qualitatively and quantitatively on serial samples. The HCV-RNA decay curves suggested a different behaviour of viral clearance induced by the three treatments. While no significant differences were present in the first 6 hours, between 6 to 12 hours Ribavirin induced a rapid decline in the viral load. Amantadine seemed to accelerate it in the third phase (12 to 30 hours) and to provoke a more pronounced viral decline when compared to IFN alone (P < 0.05) or to IFN plus Ribavirin (P < 0.025) (baseline to 30 hours). Thus, while IFN remains the principal antiviral drug, Amantadine upholds the viral decline. Ribavirin, although synergistic with IFN, does not seem to improve the IFN effect during the earliest phase of treatment but probably supports the effects of IFN later on. A new dynamic approach to HCV treatment can therefore be developed.

Hepatitis G virus infection in intravenous drug users with or without human immunodeficiency virus infection.

BACKGROUND/AIMS: To evaluate the HGV infection prevalence in a group of intravenous drug users with or without human immunodeficiency virus coinfection. METHODOLOGY: We studied 57 patients (48 males and 9 females) who were either previous or still ongoing intravenous drug users. Thirty-seven patients were HIV+ve, 55 patients were anti-HCV+ve and 3 patients were HBsAg chronic carriers. Patient sera were tested for HGV-RNA, anti-E2, qualitative and quantitative HCV-RNA as well as for HCV genotypes. Moreover, the ALT level was checked in the serum sample of each patient. RESULTS: We found a high prevalence (35/57; 61.4%) of HGV infection in our patients. HGV-RNA was detected in 16 out of the 57 intravenous drug users (28%). In particular HGV-RNA was positive in 12 out of the 37 HIV+ve patients (32.4%) and in 4 out of the 20 HIV-ve patients (20%). Anti-E2 were detected in 19 out of the 57 patients (33.3%) with greater prevalence among HIV-ve subjects (12/20; 60%) compared to HIV+ve group (7/37; 18.9%). This resulting difference was statistically significant (P < 0.05). All HGV-RNA+ve/anti-E2+ve patients were anti-HCV/HCV-RNA+ve and none of our patients were anti-E2+ve/HGV-RNA+ve at the same time. Significant differences were not found between HGV-RNA+ve and HGV-RNA-ve patients as far as clinical and virological data are concerned. CONCLUSIONS: The prevalence of HGV infection in intravenous drug users proved to be high especially in the HIV+ve group. Moreover HGV was associated with HCV in all our cases. The actual clinical impact of HGV infection remains unclear since HGV does not seems to influence the biochemical, virological or histological alterations caused by HCV infection.

TT virus infection in haemodialysis patients.

BACKGROUND: The recent discovery of a new parenterally transmitted DNA virus called TT virus (TTV) led us to investigate its prevalence in haemodialysis patients, a high-risk group for blood-borne infection, and to evaluate its role in liver disease. Moreover, we compared the TTV prevalence with the prevalence of other hepatitis virus coinfections. METHODS: Serum samples of 78 patients on maintenance haemodialysis were tested for TTV-DNA, hepatitis G virus (HGV)-RNA, anti-E2, anti-hepatitis C virus (HCV) and HCV-RNA. TTV-DNA was detected by semi-nested PCR using the primers from open reading frame 1 (ORF). HGV-RNA was detected by PCR using specific primers for the NS3 and the 5'-UTR genome regions while anti-E2 were checked by an enzyme immunological test. Anti-HCV was tested by the second generation Chiron RIBA HCV test system. HCV-RNA was evaluated by nested PCR with primers directed to the highly conserved 5' non-coding region of the HCV genome. RESULTS: TTV prevalence in our patients was 19% (15/78) while the prevalence of HCV and HGV infection proved to be 20 and 15.4%, respectively. Among TTV positive patients HGV co-infection was present in five cases (33%), HCV in six cases (39.9%), while HBV co-infection was not present in any of the patients. Only three patients proved positive for all three viruses. ALT levels were normal in most cases (13/15; 86%). In particular, patients with TTV infection alone showed normal ALT levels and HCV coinfection was found in the two patients with moderate ALT increases. CONCLUSIONS: TTV prevalence in haemodialysed patients is significant though the real clinical impact is still unclear. However, we must keep in mind that the epidemiological relevance of TTV infection is probably underestimated due to the impossibility in detecting the corresponding antibody.

Soluble beta2-mu-associated and beta2-mu-free HLA class I heavy chain serum levels in interferon-alpha nonresponder chronic hepatitis C patients. Markers of immune activation, and response to antiviral retreatment.

The serum levels of soluble beta2-mu-associated and beta2-mu-free HLA class I heavy chains were determined in 28 interferon-alpha nonresponder chronic hepatitis C patients retreated with interferon-alpha plus ribavirin and in 70 healthy subjects. The baseline levels of beta2-mu-associated and beta2-mu-free HLA class I heavy chains were significantly higher in patients than in healthy controls (P = 0.001). The levels of beta2-mu-associated HLA class I heavy chains significantly increased in responder patients with respect to nonresponders at the third month of treatment (P = 0.03). At the sixth month of treatment and after 6 months of follow up the levels of beta2-mu-associated HLA class I heavy chains decreased in responder patients and increased in nonresponders. The levels of beta2-mu-free HLA class I heavy chains showed only minor changes during and after treatment. We suggest that the determination of hepatitis C virus RNA levels combined with soluble beta2-mu-associated HLA class I heavy chains, as a marker of immune activation, could identify interferon-alpha non responder chronic hepatitis C patients most likely to respond to a retreatment with interferon-alpha plus ribavirin.

Interferon retreatment in chronic hepatitis C: which patients to choose, and what schedule to use.

UNLABELLED: OBJECTIVE; To evaluate the results of a large cohort of non-responder or relapsing responder patients with chronic hepatitis C retreated with various schedules of interferon (IFN). METHODS: Our study included 276 patients (158 non-responders and 118 relapsing responders) who underwent IFN retreatments. Among the non-responder group, 158 patients underwent further courses of IFN. In particular, 108 patients underwent one course of IFN retreatment, 40 patients underwent two courses, eight patients underwent three courses, and two patients underwent four courses. Regarding the relapsing responder group, the 118 patients were retreated with the same dosage for varying periods. In particular, 50 patients were treated for 6 months, 43 patients for 12 months, and 25 for 24 months. Patients in the subgroups of IFN retreatment were homogeneous as far as age and gender distribution, as well as virological and histological characteristics, are concerned. Qualitative and quantitative HCV-RNA was evaluated at baseline, at the end of treatment and at the last check-up of follow-up. HCV genotype was determined on baseline serum samples. Alanine transaminase (ALT) levels were tested monthly. RESULTS: Long-term biochemical (normal ALT levels) and virological (HCV-RNA negative) response was obtained in 2.6% of non-responder retreated patients, and in 33.9% of relapsing responder retreated patients. Evaluation of response on the basis of the duration of treatment showed that 48%, 19% and 16% of relapsing responder patients retreated for 24, 12 and 6 months, respectively, obtained long-term biochemical and virological response. CONCLUSION: Non-responder patient retreatment is inefficient especially in cirrhotic and/or genotype 1 b patients. IFN retreatment is warranted in relapsing responder patients. In particular, 24-month therapy induces significant long-term biochemical and virological response.

Hepatitis G virus infection in haemodialysis and in peritoneal dialysis patients.

The aim of this study was to detect hepatitis G virus RNA (HGV RNA) and antibodies against the virus envelope protein E2 (anti-E2) in 107 patients either on maintenance haemodialysis (n = 78) or peritoneal dialysis (n = 29) to evaluate the prevalence of HGV infection and to establish its role in liver disease. The total prevalence of HGV infection was of 15.4% among haemodialysis patients, whereas it was 10.3% among peritoneal dialysis patients. HGV RNA was detected in 2 haemodialysis patients (2.6%) and in 3 peritoneal dialysis patients (10.3%). Anti-E2 was found in 10 haemodialysis patients (7.8%), whilst all peritoneal dialysis patients resulted negative. In only 1 patient the alanine aminotransferase level was elevated. This patient underwent liver biopsy that did not reveal evidence of chronic hepatitis. The lower HGV prevalence in haemodialysis patients, when compared with data reported by other European authors, should be related to the lower rate of polytransfused patients in our series (29.5%). Multiple blood transfusions should be considered as the main factor to explain the different prevalence of HGV infection among various European dialysis centres. Detection of both antibody and viraemia is important to establish the real rate of the infection.

Chronic hepatitis C. New therapeutic strategies.

BACKGROUND: Aim of this work was to evaluate the early viral decay induced by a daily therapy with alfa-interferon (IFN) and the presence of any synergistic effects of amantadine and ribavirin. METHODS: Twenty patients with a diagnosis of chronic hepatitis C were randomly assigned to receive a course of treatment with: IFN 3MU daily (6 pts); or IFN 3MU daily plus amantadine 200 mg (7 pts): or IFN 3MU daily plus ribavirin 1-1.2 g (7 pts) for 6 months. Blood samples were drown at baseline, at 6, 12, 24, 30 and 48 hrs after the first dose of IFN; at 3, 7, 15 days and at every month. Serum was separated within two hours from the collection and stored at -80 degrees C until use. Viraemia was evaluated qualitatively by the Cobas Amplicor (cut-off 1.00E+02 copies/ml) (Roche Diagnostics, Monza, Milan, Italy) and quantitatively by the Cobas Amplicor Monitor (cut-off 1.00E+03 copies/ml). The HCV genotype was determined for each patient by Inno-LiPA HCV II (Innogenetics, Ghent, Belgium). Liver function tests were evaluated at baseline, at 7 and 15 days and at every month. RESULTS: The analysis of the decay curves showed the presence of a three phase decline in the viraemia. At the end of therapy 7 out of the 20 patients (35%) had normal ALT and undetectable HCV-RNA (2 out of 6 in the IFN group: 33.3%, 3 out of 7: 42.8%; 2 out of 7: 28.6%, in the IFN plus amantadine and IFN plus ribavirin groups respectively). CONCLUSIONS: IFN is the major antiviral effector in the early stage of therapy. The observation of the kinetic curves shows a tendency for the ribavirin to induce a slightly steeper slope of decay in the first 48 hrs, while amantadine seems to induce a slightly deeper abatement of circulating viraemia after 48 hrs.

Serum intercellular adhesion molecule-1 and long-term response to IFN-alpha2b therapy in chronic hepatitis C.

We have attempted to correlate the outcome of interferon (IFN) therapy with circulating soluble intercellular adhesion molecule-1 (sICAM-1) and the level of viremia in a sample of patients with chronic hepatitis C virus (HCV) infection. Forty-two patients were studied. Eighteen patients were maintained in long-term remission following IFN therapy, whereas 24 did not respond or relapsed. Serum concentrations of sICAM-1 were measured by enzyme-linked immunoassay. Viremia was measured by branched DNA signal amplification assay. Basal sICAM-1 was significantly higher in long-term responders than in nonresponder/relapsing patients. It was found that very high levels (>1000 ng/ml) were closely associated with long-term clinical response. A quantitative evaluation of viremia in basal conditions, which was significantly lower in long-term responders, gave completely opposite results. During treatment, sICAM-1 concentrations significantly decreased in the group of long-term responders but not in the nonresponders. sICAM-1 reduction was apparent as early as 1 month after treatment started. Serum sICAM-1 may be a useful parameter in evaluating the outcome of patients with chronic hepatitis C infection treated with IFN.

Chronic hepatitis induced by Jin Bu Huan.

BACKGROUND/AIMS: Jin Bu Huan and other Chinese herbal products are widely taken remedies. They have been developed as a natural alternative to traditional drugs in the treatment of various ailments. Their ability to induce several side effects such as acute hepatitis has already been described. We report a case of chronic hepatic damage following administration of Jin Bu Huan Anodyne tablets. METHODS: The patient, a 49-year-old man, developed biochemical signs of liver damage 2 months after beginning Jin Bu Huan intake (3 tablets/daily) including biopsy-proven chronic hepatitis with moderate fibrosis. Virological, autoimmune, metabolic or other hepatotoxic causes were excluded. Liver function impairment was resolved by discontinuing Jin Bu Huan intake. CONCLUSIONS: This case reinforces the already known hepatotoxicity of this product and should make us think more about the uncontrolled use of alternative products.

Vertical transmission of hepatitis C virus infection: usefulness of viremia detection in HIV-seronegative hepatitis C virus-seropositive mothers.

Seventy mother-newborn pairs were studied for hepatitis C viremia to evaluate the risk of vertical transmission of hepatitis C virus from human immunodeficiency virus-negative mothers. Forty-five mothers were hepatitis C virus-RNA positive: 4 to 45 children were positive at birth and during follow-up. The level of viremia plays an important role in vertical transmission.

Is hepatitis G virus a real risk for haemodialysis patients?

Haemodialysis patients are at high risk of developing liver disease due to blood-borne viral agents. At present hepatitis C virus (HCV) is the most common cause of infection in these patients. A new RNA virus of the Flaviviridae family, hepatitis G virus (HGV) has recently been cloned. HGV prevalence in haemodialysis patients ranges from 3.1% to 57.5%. The aim of this study has been to detect HGV-RNA in our haemodialysis patients in order to evaluate the prevalence of HGV and to correlate the viral presence to liver disease. A total of 79 patients, on haemodialysis for a mean of 52 months, were tested. 3 patients (3.8%) were HBsAG positive and 19 patients (24%) were HCV positive. 24 of the 79 (30%) patients had been transfused. Only 2 of the 79 patients (2.5%) were HGV positive. These patients were HBsAG and anti HCV negative, both had been previously transfused and showed no signs of liver disease.