Anna Maria Tarola

Oxidation of ferulic acid by laccase: identification of the products and inhibitory effects of some dipeptides

The laccase activity, in the presence of ferulic acid as substrate, was tested with high performance liquid chromatography (HPLC). The two products initially formed were identified by HPLC-mass spectrometry and HPLC-diode array as trans-5-[(E)-2-carboxyvinyl]-2-(4-hydroxy-3-methoxy-phenyl)-7-methoxy-2,3-dihydrobenzofuran-3-carboxylic acid and (Z)-beta-{4-[(E)-2-carboxyvinyl]-2-methoxyphenoxy}-4-hydroxy-3-methoxy cinnamic acid. The inhibitory effects of some dipeptides due to the formation of copper complexes have also been reported. The results obtained confirm that the inhibitory effects on laccase activity are strongly related to the structure of the dipeptides and confirm the important role of copper(II) in the reaction mechanism.

Bioactive Compounds and Antioxidant Capacity of Camarosa and Selva Strawberries (Fragaria x ananassa Duch.)

Strawberries represent an important source of bioactive compounds due to their vitamin C and phenolic compound levels, which present high antioxidant effects, beneficial for the maintenance of consumer’s health. Argentina is the second largest strawberry producer in The Common Market of the Southern Cone (MERCOSUR), covering the main export destinations of Argentinian strawberries, i.e., Canada, United States, and European Union. Information about the bioactive compound occurrence and antioxidant capacity of these fruits is scarce or not available. Health related compounds of strawberry cultivars (Camarosa and Selva) from different zones of Argentina were investigated. Vitamin C content was in the same range for both studied cultivars. However, Camarosa strawberries, which are the most cultivated, consumed, and exported berries in Argentina, presented higher total phenolic and anthocyanins content, and consequently better in vitro antioxidant capacity. Moreover, there were differences in the occurrence and concentration in the phenolic compound profiles for both cultivars. Camarosa cultivar presented higher content of anthocyanidins, and Selva showed higher total ellagic acid content. The research shows that Argentina’s strawberries are an interesting source of bioactive compounds comparable to those in other parts of the world.

Simultaneous Determination of Phenolic Compounds in Red Wines by HPLC‐UV

Abstract Ten samples of commercially Italian red wines were analyzed in order to determine the phenolic content. Variations in wine types are largely due to differences in concentration and composition of these compounds. Polyphenolic compounds are a large and complex group of substances which constitute one of the most important quality parameters of wine. These constituents of red wine contribute to organoleptic characteristics and to antioxidant and anti‐inflammatory properties. Moderate wine consumption is associated with several beneficial physiological effects, which include anticancer activities, inhibition of platelet aggregation, and inhibition of LDL oxidation which constitutes the initial stage of the pathogenesis of arteriosclerosis. For the analysis, reversed‐phase high performance liquid chromatography (HPLC) method coupled with UV‐Vis detection was used. The method uses a gradient elution to identify nine biologically active phenolic constituents: catechin; epicatechin; trans‐ and cis‐resveratrol; gallic, chlorogenic and caffeic acid; rutin and quercetin in red wine samples. The samples are injected directly without any pretreatment. The method is simple, fast, not expensive and shows good linearity for all constituents, and the detection limits ranged from 0.3–1.6 µg/ml for trans‐resveratrol and gallic acid, respectively. Moreover, the samples were analyzed in different times for estimation of stability of these compounds.

Determination of polycyclic aromatic hydrocarbons in Italian milk by HPLC with fluorescence detection

The presence of polycyclic aromatic hydrocarbons (PAHs) in Italian commercial milk samples is reported. The study was carried out on lactating (cow and goat) and plant (rice, soya, oat) milk. The quantitative determination involved liquid–liquid extraction of PAHs, a pre-concentration and determination by HPLC using a fluorescence detector. The recovery of analytes was in the range of 70–115%. The precision of the method was found to be between 6% and 24%. The detection limit ranged from 0.66 to 33.3 µg l–1 corresponding to 0.03–1.66 µg kg–1 milk (wet weight), at a signal-to-noise ratio of 3, depending on the compound. By this procedure, the levels of more volatile PAHs (two to three aromatic rings) were confirmed in 34 commercial milk and three plant milk samples, whereas benzo[a]pyrene was found only in five pasteurised milk samples at a mean concentration of 0.17 µg kg–1 milk. These results provide evidence that PAH levels are influenced by heat treatments and skimming processes of milk production. Graphical Abstract

Determination of indican and tryptophan in normal and uraemic patients by high-performance liquid chromatography with a new electrochemical detector

A simple analytical procedure has been developed for the determination of indican and tryptophan in biological fluids by reversed-phase liquid chromatography using a new electrochemical detector consisting of a tubular anode obtained by moulding graphitized carbon black and polyethylene. The hydrodynamic voltammetry of these compounds has been carried out and it has been found that, by operating in isocratic conditions with phosphate buffer (pH 4.0)-methanol (93:7), the reported compounds can be determined directly. The procedure can be applied for the determination of the free compounds on ultrafiltered serum as well as of their total content on serum deproteinized with methanol. Levels of both compounds in normal and uraemic patients have been measured and the relative ratios between free and total content yield a useful marker for patients with renal disease. The limits of quantitation of indican and tryptophan in serum were 5 and 10 ng/ml, respectively. The within-day assay coefficient of variation for total indican and tryptophan ranged from 3.0 to 3.6% and from 3.8 to 4.1%, respectively. The day-to-day assay coefficient of variation for total indican and tryptophan ranged from 3.4 to 3.7% and from 4.6 to 5.0%, respectively.

Enzymatic hydrolysis of pectic substances by gel-permeation chromatography

SummaryA rapid liquid chromatographic (HPLC) method for the determination of pectinolytic activity of enzymes produced by Aspergillus Niger and Rhizopus species is reported.Compared with more conventional methods, HPLC is more reliable and has a much improved maximum sensitivity. The limit of quantitation of galacturonic acid is 0.1μg.

Reversed-phase capillary electrochromatography of aloins and related constituents of aloe

SumamryThe separation of some anthrone-C-glucosyl constituents of aloe was carried out by isocratic capillary electrochromatography using a 50 μm l.D. column packed with 3 μm ODS silica gel. In comparison to HPLC mode, CEC resolution of aloins was obtained in a very short time with a high selectivity factor. The influence of mobile phase parameters (pH and ionic strength of the buffer, acetonitrile amount in the eluent etc.) on the retention of aloins was examined in order to improve the separation conditions and to study the retention mechanism. Efficiency was found to increase with decreasing the EOF up to a reduced plate height of about 2 at a reduced linear velocity of 0.3 mm s−1. This effect was ascribed to the fact that, under the buffer conditions used (pH 8), aloins are partially anionically charged, with a consequent low mass transfer between the mobile and stationary phases. However, resolution remains almost the same in a wide pH range permitting baseline separations of aloins in short analysis times. Since aloins are known to undergo fast degradation reaction dependent on pH, a kinetic study was also carried out. The method was extended to the analysis of aloins in food and cosmetic products.

Polyphenol Content and Antioxidant Activity of Merlot and Shiraz Wine

The total polyphenol content, antocyanins, proantoacyanidins, and some phenols were determined in fifteen red wine samples of the same vintage from single-blended Merlot or Shiraz cultivars. The principal goal of this study was to use these compounds for wine identification and to evaluate the correlation with antioxidant activity. High-performance liquid chromatography (HPLC) was used to determine catechin, epicatechin, rutin, quercetin, malvidin, peonidin, petunidin, cyanidin, delphinidin, pelargonidin, trans-resveratrol, gallic acid, chlorogenic acid, and p-coumaric acid. Spectrophotometric methods were employed to determine chromatic profile, total polyphenol content, antocyanins, proanthocyanidins, and antioxidant activity. There were some differences in the concentrations of phenolics for Shiraz and Merlot wines of 2009 vintage, including total polyphenols, antocyanidins, and proanthocyanidins. The analysis revealed a significantly higher antioxidant activity of Merlot than Shiraz and a strong correlation between antioxidant capacity and total polyphenol concentration.

Determination of Polycyclic Aromatic Hydrocarbons in Tea Infusions Samples by High Performance Liquid Chromatography with Fluorimetric Detection

This study focuses on the contamination of 15 polycyclic aromatic hydrocarbons (PAHs), recommended by the US Environmental Protection Agency, in 10 tea brands distributed in Italy. Analyses were carried out with a procedure based on saponification, liquid-liquid extraction, and PAHs determination by high performance liquid chromatography with fluorescence detector. A comparison with ultrasonic extraction in bath water was also reported. Contamination is expressed as the sum of analyzed PAHs and ranged between 347 and 4120 ng/L with a mean value of 1675 ng/L. PAHs with 3-4 rings were dominant with a contribution of 92%, while 7% and 1% were found for PAHs with 5 and 6 rings, respectively. Moreover, data revealed that three samples exceeded the EU 2008 criteria established for drinking water in which the sum of benzo[k]fluoranthene, benzo[b]fluoranthene, benzo[g,h,i]perylene, and indeno[1,2,3-cd]pyrene is considered (<100 ng/L) and two samples exceeded the 10 ng/L level allowed for benzo[a]pyrene.

Polycyclic aromatic hydrocarbons (PAHs) in yogurt samples

The concentrations and distributions of major polycyclic aromatic hydrocarbons (PAHs) were determined in 20 kinds of yogurt specimens collected from Italian supermarkets using reversed phase high-performance liquid chromatography equipped with fluorescence detection. The method was validated by determination of recovery percentages, precision (repeatability) and sensitivity (limits of detection) with yogurt samples fortified at 0.25, 0.5 and 1 µg/kg concentration levels. The recovery of 13 PAHs, with the exception of naphthalene and acenaphthene, ranged from 61% to 130% and from 60% to 97% at all the levels for yogurts with low (0.1%) and high (3.9%) fat content, respectively. The method is repeatable with relative standard deviation values <20% for all analytes. The results obtained demonstrate that acenaphthene, fluorantene, phenanthrene, anthracene, fluoranthene and pyrene were found in all samples with a similar distribution, but different content when yogurts with low and high fats were compared.