The chemical memory of smoking tobacco.

The concentration in urine of N-acetyl-hydroxy-propyl-cisteine (3HPMA), an acrolein metabolite, has been employed as a marker of the risk of illness of smokers and the relative concentration of creatinine has been evaluated to verify the effect of moving from the practice of burning tobacco to nicotine vaping. From the results concerning the urine samples of 38 subjects, collected from 2021 to 2023 and analyzed by LC-MS/MS, corresponding to 5 active smokers, 13 previously heavy smokers who replaced traditional tobacco by vaping, and 20 non-smokers, a dramatic reduction was found in 3HPMA/creatinine in urine. 3HPMA varied from values of 2150-3100 µg gcreatinine-1 to levels of 225-625 µg gcreatinine-1 found for non-smokers, with the time decay described by the equation y = 0.3661x2 - 94.359x + 6246.4 (R2: 0.757), providing a time of approximately 10 years for tobacco memory after the cessation of the consumption of burned tobacco.

Biomarkers of exposure in urine of active smokers, non-smokers, and vapers.

The exposure to smoking related products has been evaluated through urine illness risk marker determination through the analysis of urine samples of smokers and vapers. Biomarkers and their metabolites such as N-acetyl-S-(2-cyanoethyl)-L-cysteine (CEMA), N-acetyl-S-(3,4-dihydroxybutyl)-L-cysteine (DHBMA), N-acetyl-S-[1-(hydroxymethyl)-2-propen-1-yl)-L-cysteine (MHBMA), N-acetyl-S-(3-hydroxypropyl)-L-cysteine (3HPMA), 2R-N-acetyl-S-(4-hydroxybutan-2-yl)-L-cysteine (HMPMA), and N-acetyl-S-(3-carboxy-2-propyl)-L-cysteine (CMEMA) together with nicotine and cotinine were identified and quantified by LC-HRMS and LC-MS/MS, and data found normalized to the creatinine level. One hundred two urine samples were collected from smokers, non-smokers, and vapers, spanning an age range from 16 to 79 years. Results obtained showed that CEMA was only detected in urine samples from smokers and MHBMA was in the same order of magnitude in all the urine samples analyzed. HMPMA was found in the urine of vapers at the same order of concentration as in non-smokers. 3HPMA in vapers was lower than in the urine of smokers, presenting an intermediate situation between smokers and non-smokers. On the other hand, DHBMA in vapers can reach similar values to those found for smokers, while CMEMA shows concentrations in the urine of vapers higher than in the case of non-smokers and traditional smokers, requiring new research to link this metabolite to the use of electronic cigarettes and possible alternative metabolomic routes. In general, this study seems to verify that traditional smoking practice constitutes a major source of carcinogenic chemicals compared with substitutive practices, although those practices are not free of potential harm.

A green methodology for the determination of cocaine in camouflaged samples.

A methodology based on the ultrasound-assisted extraction with ethanol and the dry film attenuated total reflectance infrared spectroscopy (DF-ATR-FTIR) measurement of extracts has been developed for a fast evaluation of non-conventional ("exotic") solid-sized cocaine samples. The method provides quantitative results in less than three minutes with a limit of detection in the solid sample of 1.6 µg g-1 of cocaine with a variation coefficient lower than 7%. Results found for seized samples of different natures were compared with those obtained by a reference gas chromatography method and the greenness of the whole proposed procedure was evaluated and compared using the analytical eco-scale, green analytical procedure index (GAPI), and analytical greenness metric (AGREE). The green evaluation of the proposed methodology provided green scores by considering different evaluation criteria.

Date-rape evidence through fast determination of γ-butyrolactone in adulterated beverages.

An infrared spectroscopy (IR) based methodology has been developed to determine γ-butyrolactone (GBL) in adulterated beverages. The proposed method permits the direct screening of GBL in beverages and involves a minimum sample treatment requiring less than 2 min for quantitative determination of GBL. Sensitivity of IR method was improved by using liquid-liquid extraction (LLE) providing detection limits of 0.023 mg g-1. Accuracy of the proposed methodology was evaluated through the analysis of soft beverages and alcoholic cocktails spiked with GBL at concentration levels ranging from 0.075 to 10 mg g-1 providing recovery values from 91 to 100%. GBL was determined in twelve blind-spiked beverages, including from mineral water to wine and cocktails. Results obtained were statistically comparable to those provided by a liquid chromatography (LC) reference methodology and consistent with the spiked values. Therefore, the use of LLE-FTIR allowed a simple, sensitive and quantitative determination of GBL in soft beverages and alcoholic cocktails, thus evidencing its use for sex submission intention.

Accumulating evidences of callose priming by indole- 3- carboxylic acid in response to Plectospharella cucumerina.

Indole-3-carboxylic acid (I3CA) is an indolic compound that induces resistance in Arabidopsis adult plants against the necrotrophic fungus Plectosphaerella cucumerina through primed callose accumulation. In this study, we confirm the relevance of ATL31 and SYP121 genes involved in vesicular trafficking in I3CA priming of defenses and we discard camalexin as a mediator of I3CA-induced resistance (IR) in adult plants. In addition, we observed that an intact I3CA biosynthetic pathway is necessary for I3CA-IR functionality.

Fast authentication of tea tree oil through spectroscopy.

Two new procedures, based on infrared spectroscopy in the near infrared (NIR) and mid infrared (MIR), have been developed for the authentication of tea tree oil (TTO) commercial samples. Infrared measurements were made on untreated samples by transmission NIR and attenuated total reflectance (ATR) followed by partial least square discriminant analysis (PLS-DA). These methods offer a fast and low cost alternative to enantiomeric two-dimensional gas chromatography coupled to mass detection usually employed to discriminate between authentic and non-authentic samples. In these studies, a set of 267 samples, including authentic and non-authentic labelled tea tree oil samples, were used to build the models based on the wavenumber range, data pre-processing and latent variables number selection. Infrared methods can be discriminant for authentic and non-authentic TTO samples with a 98% certainty for both ATR and NIR methodologies, employing 92 and 142 external samples respectively. Developed PLS-DA infrared based methodologies and the reference methodology have been evaluated and compared from a Green Analytical point of view.

Mycorrhizal tomato plants fine tunes the growth-defence balance upon N depleted root environments.

In low nutritive environments, the uptake of N by arbuscular mycorrhizal (AM) fungi may confer competitive advantages for the host. The present study aims to understand how mycorrhizal tomato plants perceive and then prepare for an N depletion in the root environment. Plants colonized by Rhizophagus irregularis displayed improved responses to a lack of N than nonmycorrhizal (NM) plants. These responses were accomplished by a complex metabolic and transcriptional rearrangement that mostly affected the gibberellic acid and jasmonic acid pathways involving DELLA and JAZ1 genes, which were responsive to changes in the C/N imbalance of the plant. N starved mycorrhizal plants showed lower C/N equilibrium in the shoots than starved NM plants and concomitantly a downregulation of the JAZ1 repressor and the increased expression of the DELLA gene, which translated into a more active oxylipin pathway in mycorrhizal plants. In addition, the results support a priorization in AM plants of stress responses over growth. Therefore, these plants were better prepared for an expected stress. Furthermore, most metabolites that were severely reduced in NM plants following the N depletion remained unaltered in starved AM plants compared with those normally fertilized, suggesting that the symbiosis buffered the stress, improving plant development in a stressed environment.

Ion mobility spectrometry as a fast analytical tool in benzalkonium chloride homologs determination.

A novel procedure is proposed for the determination by ion mobility spectrometry (IMS) of C12, C14 and C16 benzalkonium chloride (BAC) homologs. The proposed method requires minimum sample treatment and the measurement was made in less than one minute. A high sensitivity was obtained for BAC determination by IMS with limit of detection values from 37 to 69µgL-1. Accuracy of the proposed methodology was evaluated through the analysis of aqueous and alcoholic samples spiked with BAC at concentration levels from 0.002% to 20% (w/v), providing recovery values from 91% to 104%. BAC was determined in sanitary alcohols, nasal sprays, postharvest products, algaecides, and treated swimming pool water. Results obtained by the proposed IMS methodology were statistically comparable to those provided by a liquid chromatography-ultraviolet (LC-UV) reference methodology. The Green Certificate evaluation of the proposed IMS methodology provided 91 score points in the Eco-Scale as compared with 77 for LC-UV method.

Indoor and outdoor determination of pesticides in air by ion mobility spectrometry.

The use of ion mobility spectrometry (IMS) has been evaluated as analytical methodology to detect and evaluate the occupational exposure to pesticides. The developed IMS methodology was used, in positive and negative modes, to determine the presence of pesticides in air and to evaluate possible inhalation exposures of workers and users based on active sampling on Teflon membranes and direct thermal desorption IMS. The negative IMS mode was used to determine bensulfuron, clorpyrifos, diniconazole, diuron, flutolanil and imidacloprid, while the positive mode was employed to evaluate formetanate, metalaxyl, metamitrone, metribuzin, paclobutrazol and pirimicarb. The IMS measurements provided limits of detection from 8pg to 600pg. Indoor air samples, from phytosanitary plants, and outdoor samples, obtained from pesticide treatments in a local farm, were analysed providing pesticide air concentrations in the range of 0.04 to>0.25mgm-3. Occupational exposure of workers and pesticide users were evaluated and compared with values recommended by the authorities, providing useful information to improve the prevention programs in the phytosanitary field.

Passive exposure to nicotine from e-cigarettes.

A procedure based on the use of ion mobility spectrometry (IMS), after liquid-liquid microextraction (LLME), has been successfully employed for the determination of passive exposure to nicotine from cigarette and e-cigarette smoking. Nicotine has been determined in exhaled breath and oral fluids of both, active and passive smokers. The aforementioned studies, made in closed environments, evidenced that the exhaled breath after conventional blend cigarette smoke provides nicotine levels of the order of 220 ng per puff, in the case of experienced smokers, being exhaled only 32 ng in the case of e-cigarettes. On the other hand, the nicotine amount in oral fluids of passive vapers was between 8 and 14 µg L(-1) lower than the average value of 38±14 µg L(-1) found for passive smokers of rolling tobacco and clearly lower than the 79±36 µg L(-1) obtained from passive smokers of classical yellow blend. This study was also placed in the frame of the verification of the e-cigarettes composition.

Green near-infrared determination of copper and mancozeb in pesticide formulations.

A green analytical procedure has been successfully developed for the simultaneous determination of copper and mancozeb in phytosanitary products. The method is based on different direct measurements of diffuse reflectance near-infrared (DR-NIR) spectra. Accuracy of the method has been evaluated by comparison of the obtained copper and mancozeb concentrations with those provided by reference methodologies based on titrimetric procedures. The average relative prediction error was 0.7 and 1.6 % for copper and mancozeb, respectively. The evaluation of the greenness of the DR-NIR procedure provided 100 points, which is the maximum value in the Green Certificate ranking, because of the absence of consumed reagents and waste generation and energy consumption lower than 0.1 kWh.

Cloning and sequencing of a dextranase-encoding cDNA from Penicillium minioluteum.

A cDNA from Penicillium minioluteum HI-4 encoding a dextranase (1,6-alpha-glucan hydrolase, EC 3.2.1.11) was isolated and characterized. cDNA clones corresponding to genes expressed in dextran-induced cultures were identified by differential hybridization. Southern hybridization and restriction mapping analysis of selected clones revealed four different groups of cDNAs. The dextranase cDNA was identified after expressing a cDNA fragment from each of the isolated groups of cDNA clones in the Escherichia coli T7 system. The expression of a 2 kb cDNA fragment in E. coli led to the production of a 67 kDa protein which was recognized by an anti-dextranase polyclonal antibody. The cDNA contains 2109 bp plus a poly(A) tail, coding for a protein of 608 amino acids, including 20 N-terminal amino acid residues which might correspond to a signal peptide. There was 29% sequence identity between the P. minioluteum dextranase and the dextranase from Arthrobacter sp. CB-8.

Cloning of the Penicillium minioluteum gene encoding dextranase and its expression in Pichia pastoris.

The DEX gene encoding an extracellular dextranase was isolated from the genomic DNA library of Penicillium minioluteum by hybridization using the dextranase cDNA as a probe. Comparison of the gene and cDNA sequences revealed that the DEX gene does not contain introns. Amino acid sequences comparison of P. minioluteum dextranase with other reported dextranases reveals a significant homology (29% identity) with a dextranase from Arthrobacter sp. CB-8. The DEX gene fragment encoding a mature protein of 574 amino acids was expressed in the methylotrophic yeast Pichia pastoris by using the SUC2 gene signal sequence from Saccharomyces cerevisiae under control of the alcohol oxidase-1 (AOX1) promoter. Over 3.2 g/l of enzymatically active dextranase was secreted into the medium after induction by methanol. The yeast product was indistinguishable from the native enzyme in specific activity and the N-terminus of both proteins were identical.