Transport and modification of humic substances present in Antarctic snow and ancient ice.

We performed a study of fulvic acids extracted from fresh and aged snow, and from recent and ancient ice in Antarctica. The fresh snow samples were collected in coastal and inland sites to evaluate the influence of the distance from the sea on organic matter transport. Moreover, in a site (Melbourne Mountain) samples were collected at different heights to study the influence of altitude on transport. The obtained results showed that dissolved fulvic acid concentrations are influenced neither by distance nor by height while particulate fulvic acid concentrations are influenced by both parameters. Moreover, the results showed that fulvic acids transported for a long distance can undergo chemical modifications. Chemical modifications are better evidenced by the analysis of samples taken in trenches at different depth, which showed structural changes attributable to the loss of nitrogen-containing compounds and to an increase in aromatic character of the structures due to reduction and/or condensation processes. With ageing, the humification process proceeds with heavy carbon losses as demonstrated by results obtained from fulvic acids isolated from ice aged between twenty-five thousand and seventy thousand years.

Quantitative assay of total dsDNA with PicoGreen reagent and real-time fluorescent detection.

We describe a quantitative assay of dsDNA based on real-time PCR measurement of fluorescence due to the interaction of PicoGreen dye with dsDNA. An aliquot of 1 to 5 ml of the sample is mixed with 45 ml of diluted PicoGreen reagent within an optical PCR tube. This is placed into the real-time apparatus set to read SYBR Green I dye at the end of three cycles of 94 degrees C for 30 s and 65 degrees C for 30 s. The averaged fluorescence value is converted into DNA amount using a calibration curve prepared with lambda-DNA standard. The calibration curve has a dynamic linear range from 0.20 to 50 ng and a standard deviation variability below 5.0%. In conclusion, this method allows reliable determinations on minimal amounts of DNA from biological samples and PCR products in clinical applications of molecular biology.

Evaluation of MISPE for the multi-residue extraction of beta-agonists from calves urine.

Methods based on molecular recognition mechanisms for the clean-up of veterinary drugs and their residues, such as immuno-, receptor- and acceptor-affinity and molecularly imprinted polymers (MIPs), have been described as selective tools to improve the selectivity and the reliability of analytical results. In this work, we tested the extraction recovery performances of a MISPE column, designed for multi-residual clean-up of beta-agonists. For this purpose, 18 different samples of calf urine were spiked at 0.25, 0.50 and 1.00 ppb with pooled standard solutions of clenbuterol (Clen), tulobuterol (Tolu), isoxsuprine (Isox), brombuterol (Brom), mapenterol (Mape) and ractopamine (Racto) and analysed on two independent analytical sessions, on a LC-MS/MS ion trap detector. Averaged recoveries, constant for each molecule considered, were 64.6% for Racto, 63.0% for Salm, 59.9% for Form, 54.7% for Brom, 52.0% for Clen, 41.8% for Mape, 38.6% for Tolu and 34.5% for Isox, respectively. Reproducibility studies gave a CV < 11% at the 0.25 ppb level. The decision limit for the identification of the target drugs ranged from 0.01 ppb for mapenterol to 0.19 ppb for salmeterol, when considering one precursor, and two product ions as identification points. Such findings indicate that the choice of the appropriate molecule as template in the MIP preparation is the critical factor to guarantee a reliable analytical multi-residue approach for beta-agonists, despite the structural differences among molecules exploiting almost the same pharmacological effect.