Clinical and experimental studies of intraoperative autotransfusion using a new filtration device

The Haemocell S-350 device has recently been introduced for intraoperative autotransfusion. The system uses a novel membrane filter to process shed blood. In the first part of this study a 0.2-micron pore size filter was used in a randomized trial comparing the use of autotransfusion (n = 8) with bank blood controls (n = 9) during aortic reconstruction. This part of the trial was abandoned because of unexpected non-surgical bleeding. Bank blood requirements fell from a median of 3.0 (range 0.0-9.0) units to 1.5 (range 0.0-7.0) units when autotransfusion was used, but these patients had a greater perioperative blood loss (1791 (range 932-3104) versus 1140 (range 440-3840) ml). There was evidence of postoperative heparin excess with an activated partial thromboplastin time ratio of 1.3 (range 0.9-3.0) versus 1.0 (range 1.0-1.2) in controls and an activated clotting time of 206 (range 143-280) versus 137 (range 107-142) s. This was confirmed by raised plasma heparin levels and a prolonged thrombin time normalized by protamine. To improve performance a 0.6-micron pore size filter was studied in ten patients. Filtration efficiency doubled from 19 to 38 per cent. Electron micrographs demonstrated better filter clearance, but 44 per cent of the original concentration of heparin remained in the reinfusate. The S-350 device may be an attractive alternative to centrifugation for intraoperative autotransfusion but, until efficiency is improved, it should only be used for cardiovascular surgery when excess heparin can be reversed with protamine.     

Estimation of impurity profiles of drugs and related materials: part 19: theme with variations. Identification of impurities in 3-oxosteroids

Due to the varied reactions leading to the 3-oxo group in steroids and the reactivity of its environment, a large number of impurities related to this group are formed during the reaction steps and the degradation studies. In this paper the experiences from the authors laboratory with the 3-oxo-related impurities in 19-nor-4-ene-3-oxosteroids (norgestrel, norethisterone, nandrolone, its esters and Nestorone) as well as corticosteroids (prednisolone, mazipredone, etc) are presented. The impurities include saturated 3-ones, 1-ene-3-ones, 5(10)-ene-3-ones, 3-deoxo and 3-ethinyl-3,5-diene derivatives, 6-ene, 8(14)-ene, 6,8(14)-diene, 6-hydroxy (alpha and beta), 10beta-hydroxy and 6-one derivatives in 4-ene-3-oxosteroids and 8(9)-ene, 9(11)-ene, 11alpha-hydroxy, 11-oxo and 4-ene-3-one derivatives in 11beta-hydroxy-1,4-diene-3-oxosteroids. The chromatographic, spectroscopic and hyphenated techniques used in this study include TLC, GC, HPLC with diode array UV detector, GC-MS, LC-MS and NMR methods.     

Synthesis,Characterization and Application of Sucrose Carboxylic Acid Polyesters — A Prototype of Non-Caloric or Surface Active Compounds

Sucrose carboxylic acid polyesters are in dependence of the degree of acylation and fatty acid composition more or less undigestible or emulsifying fat substitutes and functional additives for foods. In this connection the relationships between their molecular parameters and functional properties are discussed. New procedures of the solvent-free synthesis are presented. Furthermore, open biochemical and toxicological problems of pinocytosis and accumulation of intact polyesters in organs and tissue being still under investigations are considered.     

Upgrading of gas oils: the HDS kinetics of dibenzothiophene and its derivatives in real gas oil

The hydrodesulphurization (HDS) of dibenzothiophene (DBT), 4-methyl dibenzothiophene (4 M-DBT), 4,6-dimethyl dibenzothiophene (4,6 DM-DBT) and 4,6-diethyl dibenzothiophene (4,6 DE-DBT) as real gas oil components on NiMo/Al₂O₃ catalyst was investigated. On the basis of the first order rate constants of HDS of the individual sulphur compounds reactivities of the investigated compounds decreased in the order DBT ≫ 4 M-DBT > 4,6 DE-DBT ≈ 4,6 DM-DBT. Apparent activation energies of HDS of above sulphur compounds increased from 80.0 to 120.5 kJ/mol.     

Electrodeposition and dissolution of yttrium-hexacyanoferrate layers

The electrodeposition and dissolution of yttrium-hexacyanoferrate [YHCNFe(II)] were investigated by electrochemical quartz crystal microbalance technique (EQCM). The electrodeposition was carried out by potential cycling or stepping from solutions of Y(NO₃)₃ and K₃[Fe^III(CN)₆] of different concentrations. The ratio of the reactants was also varied. No deposition was found in dilute solutions (c < 10⁻³ mol dm⁻³). The increase of concentrations led to an intense deposition of YHCNFe(II) in the course of reduction of [Fe^III(CN)₆]³⁻. At high concentrations of the reactants a coagulation deposition of YHCNFe(III) at open-circuit has also been detected. During the reduction the first phase is the nucleation which requires saturation or oversaturation in respect to the reacting species near the gold surface. The growth phase is much faster than the formation of nuclei, and its rate depends on the concentration and the concentration ratio of the species. The composition of the deposits has been determined by total reflection X-ray fluorescence (TXRF) spectrometry. From the molar ratio of atomic constituents (K, Y and Fe) of the slightly soluble deposit (solubility: 5 × 10⁻⁵ mol dm⁻³) formed after reduction of Fe(III) a formula K_0.46Y_1.18[Fe^II(CN)₆] can be derived. This value is in good accordance with the molar mass calculated from the results of EQCM experiments which also revealed that the deposit contains ca. 2 mol H₂O/mol YHCNFe(II). The solubility of YHCNFe(III) is substantially higher (s = 2 × 10⁻³ mol dm⁻³), and according to the results of TXRF measurements, its composition is Y[Fe^III(CN)₆]. The reoxidation of YHCNFe(II) takes place in two steps. The first one is a partial oxidation which is accompanied by the desorption of K⁺ ions from the layer. During further oxidation a fast dissolution occurs due to the high solubility of YHCNFe(III).     

SDS electrophoresis of legume seed proteins in horizontal ultrathin-layer pore gradient gels

Summary Horizontal SDS electrophoresis of 18 legume seed protein extracts was performed in ultrathin-layer polyacrylamide gels on foil supports. Separation results of the SD S pore gradient electrophoresis (T=4–22.5%) are compared to those of SDS electrophoresis in a constant pore size gel (T=10%). Resolution as well as the sensitivity (0.1 g protein per band) of the ultrathin-layer SDS pore-gradient electrophoresis were extremely high. Because of the very low gel thickness, separation, staining and drying were completed in substantially shorter times than achieved with conventional thick gels. An easy technique for casting ultrathin-layer (360 gm) concave gradient gels for 10 cm separation distance and a width of 25 cm is described. The even distribution of the concave exponential pore-gradient over the whole gel width is demonstrated. Molecular weights of the legume proteins are detected from 5,000 to 110,000 daltons. The protein patterns are genus- and species-specific.

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Horizontale Ultradünnschicht SDS-Gradientengel-Elektrophorese von Leguminosensamenproteinen

Zusammenfassung Die ausgezeichnete Trennschärfe der horizontalen Ultradünnschicht-SDS-Gradienten-gel Elektrophorese wird am Beispiel von Samenproteinen 18 verschiedener Leguminosengattungen, -arten und -sorten gezeigt. Es werden die Trennergebnisse der SDS-Elektrophorese mit Gelgradienten (T=4-22,5%) bzw. mit Gelen konstanter Porengröße (T=10%) verglichen. Das höchste Auslösungsvermögen und die beste Trennschärfe zeigen ultradünne Gradientengele. Die Nachweisempfindlichkeit ist bei allen Ultradünn-schicht-SDS-Elektrophoresen sehr hoch (0,1 g Protein/Bande). Da die auf Folie polymerisierten Gele sehr dünn sind (360 m) kann mit wesentlich verkürzten Trenn-, Färbe-, Entfärbe- und Trocknungszeiten gear-beitet werden. Es wird eine einfache Herstellung ultradünner Polyacrylamidgele mit exponentiellen konkaven Gradienten für die Trenndistanz von 10 cm mit einer Breite von 25 cm beschrieben. Die gerade und gleichmäßige Verteilung des Gradienten über die gesamte Gelbreite wird gezeigt. Mit der beschriebenen Methode werden bei den untersuchten Leguminosen-proteinen Molekulargewichte von 5 000 his 110 000 Dalton gefunden. Die Proteinmuster erweisen sich als gattungs- und artspezifisch.

     

Recombinant antibody fragments that detect enoyl acyl carrier protein reductase in Brassica napus

Purified Brassica napus enoyl acyl carrier protein reductase (ENR) was used to select specific antibodies from a library of antibody fragments, single-chain F_v (scF_v), displayed on filamentous phage. Analysis of the selected clones by BstNl fingerprinting and nucleotide sequencing showed that the scF_v were derived from three different human V_H germline genes. The binding specificities were confirmed by Western blots and ELISA. The scF_v preparations reacted with B. napus ENR, but not with β-keto reductase, nor enoyl reductase from Escherichia coli. Analysis of fragments generated by CNBr treatment indicates that the scF_v 3.13 recognizes an epitope located within the n-terminal 80 amino acids of the enzyme molecule. The scF_v were used to detect ENR directly in extracts of B. napus seeds.     

Gene Variants Related to Cardiovascular and Pulmonary Diseases May Correlate with Severe Outcome of COVID-19

Background: Severe outcomes of COVID-19 account for up to 15% of all cases. The study aims to check if any gene variants related to cardiovascular (CVD) and pulmonary diseases (PD) are correlated with a severe outcome of COVID-19 in a Polish cohort of COVID-19 patients. Methods: In this study, a subset of 747 samples from unrelated individuals collected across Poland in 2020 and 2021 was used and whole-genome sequencing was performed. Results: The GWAS analysis of SNPs and short indels located in genes related to CVD identified one variant significant in COVID-19 severe outcome in the HADHA gene, while for the PD gene panel, we found two significant variants in the DRC1 gene. In this study, both potentially protective and risk variants were identified, of which variants in the HADHA gene deserve the most attention. Conclusions: This is the first study reporting the association between the HADHA and DRC1 genetic variants and COVID-19 severe outcome based on the cohort WGS analysis. Although all the identified variants are localised in introns, they may be correlated and therefore inherited along with other risk variants, potentially causative to severe outcome of COVID-19 but not discovered yet.     

MIL-100(Fe) Sub-Micrometric Capsules as a Dual Drug Delivery System

Nanoparticles of metal–organic frameworks (MOF NPs) are crystalline hybrid micro- or mesoporous nanomaterials that show great promise in biomedicine due to their significant drug loading ability and controlled release. Herein, we develop porous capsules from aggregate of nanoparticles of the iron carboxylate MIL-100(Fe) through a low-temperature spray-drying route. This enables the concomitant one-pot encapsulation of high loading of an antitumor drug, methotrexate, within the pores of the MOF NPs, and the collagenase enzyme (COL), inside the inter-particular mesoporous cavities, upon the formation of the capsule, enhancing tumor treatment. This association provides better control of the release of the active moieties, MTX and collagenase, in simulated body fluid conditions in comparison with the bare MOF NPs. In addition, the loaded MIL-100 capsules present, against the A-375 cancer cell line, selective toxicity nine times higher than for the normal HaCaT cells, suggesting that MTX@COL@MIL-100 capsules may have potential application in the selective treatment of cancer cells. We highlight that an appropriate level of collagenase activity remained after encapsulation using the spray dryer equipment. Therefore, this work describes a novel application of MOF-based capsules as a dual drug delivery system for cancer treatment.