A fast method using a new hydrophilic–lipophilic balanced sorbent in combination with ultra-high performance liquid chromatography for quantification of significant bioactive metabolites in wines

This manuscript describes the development and validation of an ultra-fast, efficient, and high throughput analytical method based on ultra-high performance liquid chromatography (UHPLC) equipped with a photodiode array (PDA) detection system, for the simultaneous analysis of fifteen bioactive metabolites: gallic acid, protocatechuic acid, (−)-catechin, gentisic acid, (−)-epicatechin, syringic acid, p-coumaric acid, ferulic acid, m-coumaric acid, rutin, trans-resveratrol, myricetin, quercetin, cinnamic acid and kaempferol, in wines. A 50-mm column packed with 1.7-μm particles operating at elevated pressure (UHPLC strategy) was selected to attain ultra-fast analysis and highly efficient separations. In order to reduce the complexity of wine extract and improve the recovery efficiency, a reverse-phase solid-phase extraction (SPE) procedure using as sorbent a new macroporous copolymer made from a balanced ratio of two monomers, the lipophilic divinylbenzene and the hydrophilic N-vinylpyrrolidone (Oasis™ HLB), was performed prior to UHPLC–PDA analysis. The calibration curves of bioactive metabolites showed good linearity within the established range. Limits of detection (LOD) and quantification (LOQ) ranged from 0.006 μg mL−1 to 0.58 μg mL−1, and from 0.019 μg mL−1 to 1.94 μg mL−1, for gallic and gentisic acids, respectively. The average recoveries ± SD for the three levels of concentration tested (n = 9) in red and white wines were, respectively, 89 ± 3% and 90 ± 2%. The repeatability expressed as relative standard deviation (RSD) was below 10% for all the metabolites assayed. The validated method was then applied to red and white wines from different geographical origins (Azores, Canary and Madeira Islands). The most abundant component in the analysed red wines was (−)-epicatechin followed by (−)-catechin and rutin, whereas in white wines syringic and p-coumaric acids were found the major phenolic metabolites. The method was completely validated, providing a sensitive analysis for bioactive phenolic metabolites detection and showing satisfactory data for all the parameters tested. Moreover, was revealed as an ultra-fast approach allowing the separation of the fifteen bioactive metabolites investigated with high resolution power within 5 min.

Socio-economic status, growth, physical activity and fitness: the Madeira growth study

Within a country social conditions change over time and these conditions vary from country to country. The associations between these conditions, somatic growth, physical activity and fitness reflect these changes. Aim: The study documented variation in somatic growth, physical activity and fitness associated with socio-economic status (SES). Subjects and methods: The study involved 507 subjects (256 boys and 251 girls) from the Madeira Growth Study, a mixed longitudinal study of five cohorts (8, 10, 12, 14 and 16 years of age) followed at yearly intervals over 3 years (1996–1998). A total of 1493 observations were made. Anthropometric measurements included lengths, body mass, skeletal breadths, girths and skinfolds. Physical activity and SES were collected via questionnaire and interview. Physical fitness was assessed using the Eurofit test battery. Variation in somatic growth, physical activity and physical fitness by SES (high, average and low) was tested with analysis of variance. Results: Significant differences between SES groups were observed for height, body mass and skinfolds. Boys and girls from high SES groups were taller, heavier and fatter (subscapular and triceps skinfolds) than their peers from average and low SES groups. At some age intervals, the high SES group had larger skeletal breadths (girls) and girths (boys and girls) than low SES. Small SES differences were observed for physical activity (sport and leisure-time indices). SES was significantly associated with physical fitness. At some age levels, boys from the low SES group performed better for muscular and aerobic endurance whereas girls from the high SES group performed better for power. Conclusion: Considerable variation in somatic growth and physical fitness in association with SES has been demonstrated, but little association was found for physical activity.

New method for determination of (E)resveratrol in wine based on microextraction using packed sorbent and ultra-performance liquid chromatography

An ultra-fast and improved analytical methodology based on microextraction by packed sorbent (MEPS) combined with ultra-performance LC (UPLC) was developed and validated for determination of (E)-resveratrol in wines. Important factors affecting the performance of MEPS such as the type of sorbent material (C2, C8, C18, SIL, and M1), number of extraction cycles, and sample volume were studied. The optimal conditions of MEPS extraction were obtained using C8 sorbent and small sample volumes (50–250mL) in one extraction cycle (extract–discard) and in a short time period (about 3 min for the entire sample preparation step). (E)-Resveratrol was eluted by 1 250mL of the mixture containing 95% methanol and 5% water, and the separation was carried out on a highstrength silica HSS T3 analytical column (100 mm 2.1 mm, 1.8mm particle size) using a binary mobile phase composed of aqueous 0.1% formic acid (eluent A) and methanol (eluent B) in the gradient elution mode (10 min of total analysis). The method was fully validated in terms of linearity, detection (LOD) and quantification (LOQ) limits, extraction yield, accuracy, and inter/intra-day precision, using a Madeira wine sample (ET) spiked with (E)-resveratrol at concentration levels ranging from 5 to 60mg/mL. Validation experiments revealed very good recovery rate of 9575.8% RSD, good linearity with r2 values 40.999 within the established concentration range, excellent repeatability (0.52%), and reproducibility (1.67%) values (expressed as RSD), thus demonstrating the robustness and accuracy of the MEPSC8/UPLC-photodiode array (PDA) method. The LOD of the method was 0.21mg/mL, whereas the LOQ was 0.68mg/mL. The validated methodology was applied to 30 commercial wines (24 red wines and six white wines) from different grape varieties, vintages, and regions. On the basis of the analytical validation, the MEPSC8/UPLC-PDA methodology shows to be an improved, sensitive, and ultra-fast approach for determination of (E)-resveratrol in wines with high resolving power within 6 min.

Skeletal maturity and socio-economic status in portuguese children and youths: the Madeira growth study

Skeletal maturity is used to evaluate biological maturity status. Information about the association between socio-economic status (SES) and skeletal maturity is limited in Portugal. Aims: The aim of this study is to document the skeletal maturity of youths in Madeira and to evaluate variation in maturity associated with SES. Subjects and methods: The study involved 507 subjects (256 boys and 251 girls) from the Madeira Growth Study, a mixed-longitudinal study of five cohorts (8, 10, 12, 14 and 16 years of age) followed at yearly intervals over 3 years (1996–1998). A total of 1493 observations were made. Skeletal age was estimated from radiographs of the hand and wrist using the Tanner–Whitehouse 2 method (TW2). Social class rankings were based on Graffar’s (1956) method. Five social rankings were subsequently grouped into three SES categories: high, average and low. Results: Median for the radius, ulna and short finger bones (RUS scores) in the total sample of boys and girls increased curvilinearly across age whereas median for the 7 (without pisiform) carpal bones (Carpal scores) increased almost linearly. The 20-bone maturity scores demonstrated distinctive trends by gender: the medians for boys increased almost linearly while the medians for girls increased curvilinearly. SES differences were minimal. Only among children aged 10–11 years were high SES boys and girls advanced in skeletal maturity. Madeira adolescents were advanced in skeletal maturity compared with Belgian reference values. Conclusion: The data suggests population variation in TW2 estimates of skeletal maturation. Skeletal maturity was not related to SES in youths from Madeira.