Additional Concentration Steps Research Articles

From grape to wine: A thorough compound specific isotopic, enantiomeric and quali-quantitative investigation by means of gas chromatographic analysis

In this study, multiple analytical approaches, including simultaneous enantiomeric and isotopic analysis, were employed to thoroughly investigate the volatile fraction in Moscato giallo grape berries and wines. For the qualitative and quantitative profiling, a fast GC-QqQ/MS approach was successfully utilized. However, prior to isotopic analysis, the extracts underwent an additional concentration step, necessitating an assessment of isotopic fractionation during the concentration process. Once the absence of carbon isotopic fractionation was confirmed, this research aimed to develop a suitable gas chromatographic method for the simultaneous detection of both enantiomeric and isotopic ratios of target monoterpenoids in Moscato giallo samples. To address the limitations associated with a one-dimensional approach, multidimensional gas chromatography was employed to enhance separation before IRMS and qMS detections. Utilizing a Deans switch transfer device, the coupling of an apolar column in the first dimension and a chiral cyclodextrin-based stationary phase in the second dimension proved effective for this purpose. The data obtained from the analysis of Moscato giallo samples allowed for the assessment of natural isotopic and enantiomeric distributions in grapes and wines for the first time in the literature. Significant enantiomeric excesses were observed for the target terpenoids investigated. Regarding isotopic distribution, a consistent trend was observed for all detected target terpenols, including the linalool enantiomers. To date, this study represents the first investigation of simultaneous δ13C and chiral investigation of the main terpenoids in oenological products in the literature.

Valorization and Potential Antimicrobial Use of Olive Mill Wastewater (OMW) from Italian Olive Oil Production.

The production of olive oil generates olive mill wastewater (OMW) which essentially derives from the processing, treatment and pressing of olives in mills. Traditional milling processes require a quantity of water varying between 40 and 120 L per quintal of pressed olives, generating a considerable amount of wastewater. It is thus necessary to reduce process water and enhance its use to implement the concept of a circular economy. To this end, our preliminary work was dedicated to water purification by means of suitable and efficient filtration systems. The microfiltered OMW was firstly concentrated through reverse osmosis. Then, an additional concentration step was carried out via vacuum membrane distillation using hydrophobic hollow fiber membranes. The application of the membrane-based processes allowed the recovery of a purified water and the concentration of valuable polyphenols in a smaller volume. The different fractions obtained from the purification have been tested for the determination of the antioxidant power (DPPH assay) and dosage of polyphenols (Folin–Ciocalteu assay) and were characterized using IR spectroscopy. All samples showed relevant antioxidant activity (percentage range: 10–80%) and total phenolic content in the 1.5–15 g GAE/L range. The obtained fractions were tested for their antimicrobial effect on numerous clinical isolates of Gram-positive and Gram-negative species, resistant and multi-resistant to current antibiotic drugs. OMW samples showed widespread activity against the considered (phyto)pathogens (MIC range 8–16 mg/mL) thus supporting the value of this waste material in the (phyto)pharmaceutical field.

Optimized Protocol for the Isolation of Extracellular Vesicles from the Parasitic Worm Schistosoma mansoni with Improved Purity, Concentration, and Yield

In the past decade, the interest in helminth-derived extracellular vesicles (EVs) increased owing to their role in pathogen-host communication. However, the availability of EVs from these parasitic worms is often limited due to the restricted occurrence and culturing possibilities of these organisms. Schistosoma mansoni is one of several helminths that have been shown to release EVs affecting the immune response of their host. Further investigation of mechanisms underlying these EV-induced effects warrants separation of EVs from other components of the helminth excretory/secretory products. However, isolation of high-purity EVs often come to the expense of reduced EV yield. We therefore aimed to develop an optimized protocol for isolation of EVs from S. mansoni schistosomula and adult worms with respect to purity, concentration, and yield. We tested the use of small (1.7 ml) iodixanol density gradients and demonstrated that this enabled western blot-based analysis of the EV marker protein tetraspanin-2 (TSP-2) in gradient fractions without additional concentration steps. Moreover, the concentration and yield of EVs obtained with small iodixanol gradients were higher compared to medium-sized (4.3 ml) or conventional large-sized (12 ml) gradients. Additionally, we provide evidence that iodixanol is preferred over sucrose as medium for the small density gradients, because EVs in iodixanol gradients reached equilibrium much faster (2 hours) and iodixanol but not sucrose was suitable for purification of schistosomula EVs. Finally, we demonstrate that the small iodixanol gradients were able to separate adult worm EVs from non-EV contaminants such as the blood digestion product hemozoin. Our optimized small iodixanol density gradient allows to simultaneously separate and concentrate EVs while reducing handling time and EV loss and can be applied for EVs from helminths and other limited EV sources.

Detection of Viruses from Bioaerosols Using Anion Exchange Resin.

This protocol demonstrates a customized bioaerosol sampling method for viruses. In this system, anion exchange resin is coupled with liquid impingement-based air sampling devices for efficacious concentration of negatively-charged viruses from bioaerosols. Thus, the resin serves as an additional concentration step in the bioaerosol sampling workflow. Nucleic acid extraction of the viral particles is then performed directly from the anion exchange resin, with the resulting sample suitable for molecular analyses. Further, this protocol describes a custom-built bioaerosol chamber capable of generating virus-laden bioaerosols under a variety of environmental conditions and allowing for continuous monitoring of environmental variables such as temperature, humidity, wind speed, and aerosol mass concentration. The main advantage of using this protocol is increased sensitivity of viral detection, as assessed via direct comparison to an unmodified conventional liquid impinger. Other advantages include the potential to concentrate diverse negatively-charged viruses, the low cost of anion exchange resin (~$0.14 per sample), and ease of use. Disadvantages include the inability of this protocol to assess infectivity of resin-adsorbed viral particles, and potentially the need for the optimization of the liquid sampling buffer used within the impinger.

Technical assessment of natural deep eutectic solvent (NADES) mediated biorefinery process: A case study

Comprehensive evaluation of an integrated process for cellulosic ethanol production from lactic acid+choline chloride+water (LA-CC-H2O) mixture based natural deep eutectic solvent (NADES) pretreated rice straw; solvent recovery and its reusability; extraction of value added products viz., lignin and xylan; enzymatic saccharification and fermentation were studied. Four separate processes viz. pretreatment at 5% (w/v) and 10% (w/v) solids loading and enzymatic hydrolysis at 10% (w/v) and 25% (w/v) solids loading were investigated. No significant difference was observed in holocellulose recovery and lignin solubility in all the tested processes. Hydrolysis at 10% (w/v) and 25% (w/v) solids loading produced maximum glucose yields of 38.6±1.2g/L and 90.3±2.3g/L, respectively. Additional concentration step of sugars is needed from 10% (w/v) hydrolysis for direct ethanol fermentation using Clavispora NRRL Y-50464, while this could be voided with 25% (w/v) hydrolysis sugars. Maximum ethanol yield of 36.7g/L was obtained with 9% (w/v) glucose with a theoretical conversion efficiency of 79.9% which estimated to produce ~129L of ethanol per ton of raw biomass. Alongside, NADES pretreatment resulted in solubilization of 72-75% (w/v) of biomass bound lignin from which 86–90% (w/v) of lignin was recovered. Additionally, >98% (v/v) of the treated NADES reagent was recovered and reused in three consecutive biomass pretreatment cycles with no loss in pretreatment efficiency. This study clearly demonstrated that pretreatment at 10% (w/v) solids loading followed by enzymatic hydrolysis at 25% (w/v) solids loading is the most effective integrated biorefinery process using NADES pretreated rice straw.

Multiple effect concentration of ethanol by ohmic-assisted hydrodistillation

Our previous work on ethanol distillation by ohmic-assisted hydrodistillation (OAHD) successfully concentrated ethanol from 10 to 50% (v/v). In this work, two additional concentration steps were conducted using the 50% (v/v) product obtained from the first concentration stage at a constant wattage of 168W using the same OAHD device. The second distillation step resulted in a product with ethanol concentration of 76% (v/v) and the third step yielded ethanol concentration of 84% (v/v). Results were compared to conventional hydrodistillation (HD), and were indicative that OAHD reduces one third of energy cost in ethanol distillation process. Not only was OAHD superior to HD in terms of time and energy consumption, but also resulted in higher ethanol concentrations than HD after the third step of distillation: 84.3±0.3% vs. 82.5±0.5%, respectively. OAHD also showed better process control and was able to stop the distillation process faster than HD. In both second and third distillation steps, the time required to stop the process with OAHD (1min) was shorter than for HD process (10min). The findings of this study further confirm our previous work, and reveal that this emerging distillation technique has the potential of yielding high concentrations of ethanol from dilute alcoholic solutions in a more economical and environmental friendly way.

A Modified QuEChERS Sample Preparation Method for Simultaneous Determination of 62 Pesticide Residues in Edible Fungi Using Gas Chromatography–Triple Quadrupole Mass Spectrometry

A rapid analytical method for the simultaneous determination of 62 pesticides in edible fungi was established based on modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) sample preparation method with gas chromatography–triple mass spectrometry (GC–MS/MS). Before extracting the target compounds from edible fungi, the dry samples were soaked in distilled water for 30 min to improve the extraction efficiency. The samples were extracted with acetonitrile–water and acetic acid–sodium acetate buffer solution (pH = 5.5) and salted out by anhydrous MgSO4. Then the extracts were cleaned up with n-propyl ethylene diamine (PSA), octadecylsilane (C18), graphitized carbon black (GCB), and anhydrous MgSO4. Additional concentration steps were performed in this to improve the sensitivity of the method. Finally, the cleaned extracts were detected by GC–MS/MS in multi-reaction monitoring (MRM) mode, and the internal standard curve method was applied to quantify the pesticides. In the linear range of each pesticide, good linear calibrations with coefficients (R 2) with additional concentration steps were performed in this to improve the sensitidry Lentinula edodes. Recoveries at three levels (10, 50, and 100 μg/kg) ranged from 75.4 to 117.1 % for all the compounds, with relative standard deviations (RSD) between 3.2 and 15.2 %. The limits of quantification (LOQs) were 0.625–20 μg/kg. The QuEChERS method was more straightforward, less time and labor intensive, as well as more sensitive, selective, and accurate for screening multiple pesticide residues, and it has been applied to edible fungi samples from different provinces in China.

Immunomagnetic Separation Significantly Improves the Sensitivity of Polymerase Chain Reaction in Detecting Giavdia Duodenalis and Cryptosporidium spp. in Dairy Cattle

The effectiveness of molecular methods for the detection of species of Giardia and Cryptosporidium in fecal samples is often reduced by low or intermittent cyst and oocyst shedding, and/or the presence of polymerase chain reaction (PCR) inhibitors. The present study investigates the use of immunomagnetic separation (IMS) as an additional concentration step before PCR in the detection of these common protozoan parasites in dairy cattle. The IMS-PCR assays were optimized for amplifying fragments of the 16S ribosomal RNA (rRNA), β-giardin, and glutamate dehydrogenase (GDH) genes of Giardia duodenalis, as well as fragments of the 18S rRNA, heat shock protein (HSP)-70, and Cryptosporidium oocyst wall protein (COWP) genes of Cryptosporidium spp. In all cases, IMS-PCR was more sensitive than PCR alone. A significantly greater number of Giardia-positive samples were identified using IMS-PCR of the 16S rRNA gene (P < 0.01) and of the GDH gene (P < 0.01), as compared with PCR without any additional concentration step. In the case of Cryptosporidium, IMS-PCR of the COWP gene (P = 0.02) resulted in a significantly greater number of positives than did PCR without the IMS concentration step. The greatest number of positives, however, was obtained using IMS-PCR to amplify a portion of the 16S rRNA gene of Giardia and a portion of the HSP-70 gene of Cryptosporidium. A further comparison of the optimized IMS-PCR assays to immunofluorescence microscopy suggested that the IMS-PCR assays were considerably more sensitive than microscopy was in the detection of Giardia cysts and Cryptosporidium oocysts in fecal samples.

Potassium tartrate-glycerol as a density gradient substrate for separation of small, round viruses from human feces.

Cesium chloride density gradients are frequently used for virus concentration or purification in the preparation of human feces for examination by electron microscopy, Disruption of some of the fecal viruses occurs if they are pelleted from the density gradient in an additional concentration step. This report highlights an important limitation imposed by the use of cesium chloride as a density gradient substrate in attempting to recover small, round, virus-like particles from feces and suggests an alternative substrate which preserves virus morphology without the use of additional protective agents.