Doehlert Design-desirability Function Multi-criteria Optimal Separation Of 17 Phenolic Compounds From Extra-virgin Olive Oil By Capillary Zone Electrophoresis.

In Brazil, the consumption of extra-virgin olive oil (EVOO) is increasing annually, but there are no experimental studies concerning the phenolic compound contents of commercial EVOO. The aim of this work was to optimise the separation of 17 phenolic compounds already detected in EVOO. A Doehlert matrix experimental design was used, evaluating the effects of pH and electrolyte concentration. Resolution, runtime and migration time relative standard deviation values were evaluated. Derringer's desirability function was used to simultaneously optimise all 37 responses. The 17 peaks were separated in 19min using a fused-silica capillary (50μm internal diameter, 72cm of effective length) with an extended light path and 101.3mmolL(-1) of boric acid electrolyte (pH 9.15, 30kV). The method was validated and applied to 15 EVOO samples found in Brazilian supermarkets.

Quantitative Proteomic Analysis Reveals Metabolic Alterations, Calcium Dysregulation, And Increased Expression Of Extracellular Matrix Proteins In Laminin α2 Chain-deficient Muscle.

Congenital muscular dystrophy with laminin α2 chain deficiency (MDC1A) is one of the most severe forms of muscular disease and is characterized by severe muscle weakness and delayed motor milestones. The genetic basis of MDC1A is well known, yet the secondary mechanisms ultimately leading to muscle degeneration and subsequent connective tissue infiltration are not fully understood. In order to obtain new insights into the molecular mechanisms underlying MDC1A, we performed a comparative proteomic analysis of affected muscles (diaphragm and gastrocnemius) from laminin α2 chain-deficient dy(3K)/dy(3K) mice, using multidimensional protein identification technology combined with tandem mass tags. Out of the approximately 700 identified proteins, 113 and 101 proteins, respectively, were differentially expressed in the diseased gastrocnemius and diaphragm muscles compared with normal muscles. A large portion of these proteins are involved in different metabolic processes, bind calcium, or are expressed in the extracellular matrix. Our findings suggest that metabolic alterations and calcium dysregulation could be novel mechanisms that underlie MDC1A and might be targets that should be explored for therapy. Also, detailed knowledge of the composition of fibrotic tissue, rich in extracellular matrix proteins, in laminin α2 chain-deficient muscle might help in the design of future anti-fibrotic treatments. All MS data have been deposited in the ProteomeXchange with identifier PXD000978 (http://proteomecentral.proteomexchange.org/dataset/PXD000978).

Matrix Metalloproteinases And Left Ventricular Function And Structure In Spinal Cord Injured Subjects.

Subjects with spinal cord injury (SCI) exhibit impaired left ventricular (LV) diastolic function, which has been reported to be attenuated by regular physical activity. This study investigated the relationship between circulating matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) and echocardiographic parameters in SCI subjects and the role of physical activity in this regard. Forty-two men with SCI [19 sedentary (S-SCI) and 23 physically-active (PA-SCI)] were evaluated by clinical, anthropometric, laboratory, and echocardiographic analysis. Plasmatic pro-MMP-2, MMP-2, MMP-8, pro-MMP-9, MMP-9, TIMP-1 and TIMP-2 levels were determined by enzyme-linked immunosorbent assay and zymography. PA-SCI subjects presented lower pro-MMP-2 and pro-MMP-2/TIMP-2 levels and improved markers of LV diastolic function (lower E/Em and higher Em and E/A values) than S-SCI ones. Bivariate analysis showed that pro-MMP-2 correlated inversely with Em and directly with E/Em, while MMP-9 correlated directly with LV mass index and LV end-diastolic diameter in the whole sample. Following multiple regression analysis, pro-MMP-2, but not physical activity, remained associated with Em, while MMP-9 was associated with LV mass index in the whole sample. These findings suggest differing roles for MMPs in LV structure and function regulation and an interaction among pro-MMP-2, diastolic function and physical activity in SCI subjects.

Identification Of Corynebacterium Spp. Isolated From Bovine Intramammary Infections By Matrix-assisted Laser Desorption Ionization-time Of Flight Mass Spectrometry.

Corynebacterium species (spp.) are among the most frequently isolated pathogens associated with subclinical mastitis in dairy cows. However, simple, fast, and reliable methods for the identification of species of the genus Corynebacterium are not currently available. This study aimed to evaluate the usefulness of matrix-assisted laser desorption ionization/mass spectrometry (MALDI-TOF MS) for identifying Corynebacterium spp. isolated from the mammary glands of dairy cows. Corynebacterium spp. were isolated from milk samples via microbiological culture (n=180) and were analyzed by MALDI-TOF MS and 16S rRNA gene sequencing. Using MALDI-TOF MS methodology, 161 Corynebacterium spp. isolates (89.4%) were correctly identified at the species level, whereas 12 isolates (6.7%) were identified at the genus level. Most isolates that were identified at the species level with 16 S rRNA gene sequencing were identified as Corynebacterium bovis (n=156; 86.7%) were also identified as C. bovis with MALDI-TOF MS. Five Corynebacterium spp. isolates (2.8%) were not correctly identified at the species level with MALDI-TOF MS and 2 isolates (1.1%) were considered unidentified because despite having MALDI-TOF MS scores >2, only the genus level was correctly identified. Therefore, MALDI-TOF MS could serve as an alternative method for species-level diagnoses of bovine intramammary infections caused by Corynebacterium spp.

Effect Of Composite Containing An Iodonium Salt On The Bond Strength Of Brackets To Bovine Enamel.

This study investigated the effect of the incorporation of an iodonium salt in experimental composites, on the bond strength of metallic brackets bonded to bovine teeth. Two hundred and seventy bovine teeth were embedded in self-curing acrylic resin and divided into 18 groups (n=15), according to the experimental composite with an iodonium salt at molar concentrations 0 (control), 0.5, or 1%; the light-activation times (8, 20 and 40 s); and the storage times (10 min or 24 h). Metallic brackets were fixed on the tooth surface using experimental composites. Photoactivation was performed with a quartz-tungsten-halogen light-curing unit curing unit for 8, 20 and 40 s. The specimens were stored in distilled water at 37 °C for 10 min or 24 h and submitted to bond strength test at 0.5 mm/min. The data were subjected to three-way ANOVA and Tukey's test (α=0.05). The Adhesive Remnant Index (ARI) was used to classify the failure modes. The shear bond strengths (MPa) at 10 min for light-activation times of 8, 20 and 40 s were: G1 - 4.6, 6.9 and 7.1; G2 - 8.1, 9.2 and 9.9; G3 - 9.1, 10.4 and 10.7; and at 24 h were: G1 - 10.9, 11.1 and 11.7; G2 - 11.8, 12.7 and 14.2; G3 - 12.1, 14.4 and 15.8. There was a predominance of ARI score 3 for groups with 10 min storage time, and ARI score 2 for groups with 24 h storage time. In conclusion, the addition of iodonium salt (C05 and C1) to the experimental composite may increase the bond strength of brackets to bovine enamel using reduced light exposure times.

Inactivation Of Ampk Mediates High Phosphate-induced Extracellular Matrix Accumulation Via Nox4/tgfß-1 Signaling In Human Mesangial Cells.

High phosphate (Pi) levels and extracellular matrix (ECM) accumulation are associated with chronic kidney disease progression. However, how high Pi levels contribute to ECM accumulation in mesangial cells is unknown. The present study investigated the role and mechanism of high Pi levels in ECM accumulation in immortalized human mesangial cells (iHMCs). iHMCs were exposed to normal (0.9 mM) or increasing Pi concentrations (2.5 and 5 mM) with or without diferent blockers or activators. NOX4, phosphorylated AMPK (p-AMPK), phosphorylated SMAD3 (p-SMAD3), fibronectin (F/N), collagen IV (C-IV) and alpha-smooth muscle actin (α-SMA) expression was assessed via western blot and immunofluorescence. Lucigenin-enhanced chemiluminescence, and dihydroethidium (DHE) assessed NADPH oxidase activity and superoxide (SO), respectively. In iHMCs, a Pi transporter blocker (PFA) abrogated high Pi-induced AMPK inactivation, increase in NADPH oxidase-induced reactive oxygen species (ROS) levels, NOX4, p-SMAD3, α-SMA and C-IV expression. AMPK activation by AICAR, NOX4 silencing or NADPH oxidase blocker prevented high Pi-induced DHE levels, p-SMAD3, F/N, C-IV and α-SMA expression. AMPK inactivation with NOX4-induced ROS formation and transforming growth factor ß-1 (TGFß-1) signaling activation mediates high Pi-induced ECM accumulation in iHMCs. Maneuvers increasing AMPK or reducing NOX4 activity may contribute to renal protection under hyperphosphatemia.

Influence Of The Stage Of Ripeness On The Composition Of Iridoids And Phenolic Compounds In Genipap (genipa Americana L.).

Genipap fruits, native to the Amazon region, were classified in relation to their stage of ripeness according to firmness and peel color. The influence of the part of the genipap fruit and ripeness stage on the iridoid and phenolic compound profiles was evaluated by HPLC-DAD-MS(n), and a total of 17 compounds were identified. Geniposide was the major compound in both parts of the unripe genipap fruits, representing >70% of the total iridoids, whereas 5-caffeoylquinic acid was the major phenolic compound. In ripe fruits, genipin gentiobioside was the major compound in the endocarp (38%) and no phenolic compounds were detected. During ripening, the total iridoid content decreased by >90%, which could explain the absence of blue pigment formation in the ripe fruits after their injury. This is the first time that the phenolic compound composition and iridoid contents of genipap fruits have been reported in the literature.

Supercritical Carbon Dioxide Extraction Of Capsaicinoids From Malagueta Pepper (capsicum Frutescens L.) Assisted By Ultrasound.

Extracts from malagueta pepper (Capsicum frutescens L.) were obtained using supercritical fluid extraction (SFE) assisted by ultrasound, with carbon dioxide as solvent at 15MPa and 40°C. The SFE global yield increased up to 77% when ultrasound waves were applied, and the best condition of ultrasound-assisted extraction was ultrasound power of 360W applied during 60min. Four capsaicinoids were identified in the extracts and quantified by high performance liquid chromatography. The use of ultrasonic waves did not influence significantly the capsaicinoid profiles and the phenolic content of the extracts. However, ultrasound has enhanced the SFE rate. A model based on the broken and intact cell concept was adequate to represent the extraction kinetics and estimate the mass transfer coefficients, which were increased with ultrasound. Images obtained by field emission scanning electron microscopy showed that the action of ultrasonic waves did not cause cracks on the cell wall surface. On the other hand, ultrasound promoted disturbances in the vegetable matrix, leading to the release of extractable material on the solid surface. The effects of ultrasound were more significant on SFE from larger solid particles.

A Metabolomic Protocol For Plant Systematics By Matrix-assisted Laser-desorption/ionization Time-of Flight Mass Spectrometry.

Matrix-assisted laser desorption/ionization time-of flight mass spectrometry (MALDI-TOF MS) has been widely used for the identification and classification of microorganisms based on their proteomic fingerprints. However, the use of MALDI-TOF MS in plant research has been very limited. In the present study, a first protocol is proposed for metabolic fingerprinting by MALDI-TOF MS using three different MALDI matrices with subsequent multivariate data analysis by in-house algorithms implemented in the R environment for the taxonomic classification of plants from different genera, families and orders. By merging the data acquired with different matrices, different ionization modes and using careful algorithms and parameter selection, we demonstrate that a close taxonomic classification can be achieved based on plant metabolic fingerprints, with 92% similarity to the taxonomic classifications found in literature. The present work therefore highlights the great potential of applying MALDI-TOF MS for the taxonomic classification of plants and, furthermore, provides a preliminary foundation for future research.

Anthocyanin And Other Phenolic Compounds In Ceylon Gooseberry (dovyalis Hebecarpa) Fruits.

Ceylon gooseberry is a deep-purple exotic berry that is being produced in Brazil with great market potential. This work aimed to determine major phenolic compounds in this specie by HPLC-PDA-ESI/MS. Samples were collected in two different seasons. Pulp and skin were analyzed separately. Non-acylated rutinoside derivatives of delphinidin (∼60-63%) and cyanidin (∼17-21%) were major anthocyanins tentatively identified. All anthocyanins had higher concentration in skin than in pulp (64-82 and 646-534mg of cyaniding-3-glucoside equivalents/100g skin and pulp, respectively). Moreover, anthocyanin profile changed between sampling dates (p<0.05). Mainly for delphinidin-3-rutinoside which could be a result of season variation. In this specie, non-anthocyanin polyphenols represent less than 35% of total extracted polyphenols. The tentative identification proposed a flavonol and three ellagitannins as major compounds of the non-anthocyanin phenolics fraction. Finally, anthocyanin is the major phenolic class in this fruit and its composition and content are significantly affected by season.

Matrix Support And Institutional Support: Analyzing Their Construction.

This is an analysis of the theoretical and practical construction of the methodology of Matrix Support by means of studies on Paideia Support (Institutional and Matrix Support), which is an inter-professional work of joint care in recent literature and official documents of the Unified Health System (SUS). An attempt was made to describe methodological concepts and strategies. A comparative analysis of Institutional Support and Matrix Support was also conducted using the epistemological framework of Field and Core Knowledge and Practices.

Sorghum Flour Fractions: Correlations Among Polysaccharides, Phenolic Compounds, Antioxidant Activity And Glycemic Index.

Nutrients composition, phenolic compounds, antioxidant activity and estimated glycemic index (EGI) were evaluated in sorghum bran (SB) and decorticated sorghum flour (DSF), obtained by a rice-polisher, as well as whole sorghum flour (WSF). Correlation between EGI and the studied parameters were determined. SB presented the highest protein, lipid, ash, β-glucan, total and insoluble dietary fiber contents; and the lowest non-resistant and total starch contents. The highest carbohydrate and resistant starch contents were in DSF and WSF, respectively. Phenolic compounds and antioxidant activities were concentrated in SB. The EGI values were: DSF 84.5±0.41; WSF 77.2±0.33; and SB 60.3±0.78. Phenolic compounds, specific flavonoids and antioxidant activities, as well as total, insoluble and soluble dietary fiber and β-glucans of sorghum flour samples were all negatively correlated to EGI. RS content was not correlated to EGI.

Determination Of The Phenolic Composition From Brazilian Tropical Fruits By Uhplc-ms/ms.

Although Brazil is the third largest fruit producer in the world, several specimens consumed are not well studied from the chemical viewpoint, especially for quantitative analysis. For this reason and the crescent employment of mass spectrometry (MS) techniques in food science we selected twenty-two phenolic compounds with important biological activities and developed an ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method using electrospray (ESI) in negative ion mode aiming their quantification in largely consumed Brazilian fruits (açaí-do-Amazonas, acerola, cashew apple, camu-camu, pineapple and taperebá). Multiple reaction monitoring (MRM) was applied and the selection of proper product ions for each transition assured high selectivity. Linearity (0.99580%), precision (CV<20%) and extraction recovery rate (>80%) were satisfactory and showed that the method provides an efficient protocol to analyze phenolic compounds in fruit pulp extracts.

Profile Of Phenolic Compounds Of Brazilian Virgin Olive Oils By Rapid Resolution Liquid Chromatography Coupled To Electrospray Ionisation Time-of-flight Mass Spectrometry (rrlc-esi-tof-ms).

In recent years, agronomical researchers began to cultivate several olive varieties in different regions of Brazil to produce virgin olive oil (VOO). Because there has been no reported data regarding the phenolic profile of the first Brazilian VOO, the aim of this work was to determine phenolic contents of these samples using rapid-resolution liquid chromatography coupled to electrospray ionisation time-of-flight mass spectrometry. 25 VOO samples from Arbequina, Koroneiki, Arbosana, Grappolo, Manzanilla, Coratina, Frantoio and MGS Mariense varieties from three different Brazilian states and two crops were analysed. It was possible to quantify 19 phenolic compounds belonging to different classes. The results indicated that Brazilian VOOs have high total phenolic content because the values were comparable with those from high-quality VOOs produced in other countries. VOOs from Coratina, Arbosana and Grappolo presented the highest total phenolic content. These data will be useful in the development and improvement of Brazilian VOO.

Influence Of Successive Light-activation On Degree Of Conversion And Knoop Hardness Of The First Layered Composite Increment.

To evaluate the influence of light-activation of second, third and fourth increments on degree of conversion (DC) and microhardness (KHN) of the top (T) and bottom (B) surface of the first increment. Forty samples (n = 5) were prepared. In groups 1-4, after each increment light-activation (multiple irradiation), T and B of the first increment were measured in DC and KHN. In groups 5-8, only the first increment was made (single irradiation) and measurements of DC and KHN were taken at 15 min intervals. The light-activation modes were (XL) 500 mW/cm(2) × 38 s (G1/G5); (S) 1000 mW/cm(2) × 19 s (G2/G6), (HP) 1400 mW/cm(2) × 14 s (G3/G7); (PE) 3200 mW/cm(2) × 6 s (G4/G8). Data for DC and KHN were analyzed separately by using PROC MIXED for repeated measures and Tukey-Kramer test (α = 0.05). For KHN, B showed lower values than T. PE resulted in lower values of KHN in B surface. For single and multiple irradiations, T and B of first measurement showed the lowest KHN and the fourth measurement showed the highest, with significant difference between them. For single irradiation, first and second increments presented similar KHN, different from the third and fourth increment, which did not differ between them. For multiple irradiations, the second light-activation resulted in KHN similar to first, third and fourth increments. For DC, except QTH, T presented higher DC than B. The light-activation of successive increments was not able to influence the KHN and DC of the first increment.