Next generation probiotics: an overview of the most promising candidates.

Research in the field of human microbiota and its impact on human health has opened new possibilities for the diagnosis, prevention or treatment of certain pathological conditions. A negative change in the composition of the intestinal microbiota, dysbiosis, is associated with diseases such as inflammatory bowel diseases, obesity, diabetes mellitus, or Clostridium difficile infections. For the use of human microbiota or its biologically active products in clinical practice, it is necessary to thoroughly identify and characterize properties that may be beneficial to human health. The use of the latest technology enables such research to be carried out, and we are already aware of several potential candidates for the so-called probiotics of the next generation. The aim of this article is to summarize available information on the bacteria Akkermansia muciniphila, Bacteroides fragilis, and Faecalibacterium prausnitzii, which are among the most promising and studied candidates.

Influence of lipid imbalance on butyrylcholinesterase activity and biotransformation efficiency.

Butyrylcholinesterase (EC 3.1.1.8, BChE) is highly active in plasma, skin and lung, the tissues that first contact xenobiotics, supporting a role for BChE in detoxication of xenobiotics including medicaments. A possible involvement of BChE in lipid metabolism has been suggested. Elevated BChE activity in obese individuals correlates with some parameters of lipid metabolism including increased levels of triacylglycerols (TAG) and cholesterol. The aim of this study was to estimate the BChE activity in rats on subcellular and inter-organ levels under the conditions of untreated and treated primary hypertriacylglycerolemia with the TAG lowering agent fenofibrate. No changes in BChE activity were observed in obese animals. However fenofibrate administration led to significant increase of BChE activity in all examined tissues (plasma, liver, white adipose tissue). The impact of lipid metabolic imbalance on BChE biotransformation ability was tested by measuring the rate of hydrolysis of 0,1 to 8 mM concentrations of the antimicrobial agent N-(2-benzoyloxyethyl)-ethyldimethylammonium bromide (BCH2). The results revealed a complete shift in the BChE kinetics in all studied models. In animals with hypertriacylglycerolemia the Km value of liver BChE rised 4,6-fold, but the total enzyme efficiency expressed as Vmax/Km dropped 40% comparing to control. In contrast, in animals treated with fenofibrate the BChE efficiency increased in liver 1,6-fold. We conclude here that BChE detoxification capacity is essentially altered under conditions of disturbed lipid metabolism. Clinically, this knowledge could be important in a view of xenobiotic elimination, especially when routinely prescribed medicaments are concerned.

[Characterization of polyphenoloxidase from the latex of greater celandine (Chelidonium majus L.)]. / Charakterizácia polyfenoloxidázy z latexu lastovidníka väcseho (Chelidonium majus L.).

Greater celandine, similarly as other plants of the family Papaveraceae, produces benzylisoquinoline alkaloids, primarily benzophenanthridines. Polyphenoloxidase (PPO) is most probably involved in the formation of dopamine, which is one of the precursors of norcoclaurine, the first intermediate with the benzylisoquinoline structure. This study has revealed that PPO present in the latex of greater celandine is localized in the organelles, which serve to store alkaloids (the so-called 1000 g organelles). The enzyme was purified by means of affinity chromatography into electrophoretic homogeneity. It possesses a relative molecular mass of approximately 65 kDa and exerts two activities, the monophenolase and diphenolase ones. With the use of a polymerase chain reaction, it was possible to amplify a part of the PPO gene from the region of the active site.

[Enzymology of production of benzylisoquinoline alkaloids]. / Enzymológia tvorby benzylizochinolínových alkaloidov.

This review paper summarizes the current knowledge of enzymes participating in the production of benzylisoquinoline alkaloids. This group of alkaloids comprises, e.g., morphine, codeine, thebaine, and sanginarine, which have an irreplaceable position in pharmaceutical practice. For the time being, chemists have not managed to prepare them synthetically with sufficient efficacy, and therefore the study of the enzymology of their formation remains a topical problem. The paper pays particular attention to the knowledge of individual enzymes on the molecular, or gene level. This very knowledge is essential for possible introduction of molecular-genetic approaches to the cultivation of plants producing therapeutically interesting benzylisoquinoline alkaloids.

[Effect of a fungal elicitor on levels of sanguinarine and polyphenoloxidase activity in a suspension culture of Papaver somniferum L]. / Zmeny obsahu sanguinarínu a aktivity polyfenoloxidázy vplyvom fungálneho elicitora v suspenzných kultúrach maku siateho Papaver somniferum L.

The opium poppy (Papaver somniferum L.) is still a source for isolation of codeine and morphine. Cell cultures from this plant lose their ability to produce morphinans. Their major alkaloid is sanguinarine. The elicitation of the opium poppy cell cultures by fungal preparation lead to a nine-fold increase in the content of sanguinarine. The specific activity of polyphenoloxidase (PPO) was three-times higher in the elicited compared to the nonelicited cells. Two isoforms of PPO (Mr 63 kDa, 41 kDa) were identified in opium poppy cell cultures by PAGE. The number of PPO isoforms was not affected by elicitation. Phenyl-Sepharose CL-4B was used for affinity purification of PPO. In a single purification step the specific activity of PPO was enriched 14-fold.

[Role of amine oxidase in the biosynthesis of alkaloids]. / Uloha aminooxidázy v biosyntéze alkaloidov.

The review paper deals with the contemporary theoretical knowledge about the role of Cu-aminooxidase in the biosynthesis of alkaloids in plants. In the biosynthesis of tropane and piperidine alkaloids, aminooxidase participates in the conversion of amines into aldehydes which are the first important intermediates in the biosynthesis of these alkaloids. Norkoklaurine, the precursor of benzylisoquinoline alkaloids, is formed by condensation of dopamine and tyral. In the biosynthesis of benzylisoquinoline alkaloids of protoberberine and berberine type, tyral, the aldehyde condensation unit, is produced by the action of aminooxidase. In morphinan alkaloids, the catalytic role of aminooxidase in the formation of tyral have not been demonstrated yet. The paper pays special attention to the mechanism of the aminooxidase-catalyzed reaction, the structure of the active site of the enzyme, and the molecular-biological properties of Cu-aminooxidases.

Biochemical characterization of antigens detected with anti-platelet monoclonal antibodies.

A panel of 18 monoclonal antibodies (mAbs) defined by the third workshop as specific for platelets, clustered in three preliminary groups: PC7, PC13 and PC27. These mAbs were further analysed by immunoprecipitation using extracts of iodinated and biotinylated peripheral blood mononucleated cells (PBMC) and platelets. We could confirm the existence of mAbs with specificities to WC9 (in PC7) and CD41/61 (in PC13). Two mAbs formed a new cluster, WC13, which may be homologous to human CD31 (in PC27). The influence of EDTA and thrombin on the expression of the different antigens on the platelet membrane was assessed by flow cytometry (FCM) analysis, as well as cross-reactivity with platelets from different species.

Immunohistochemical reactivity of anti-platelet monoclonal antibodies.

Ten mAbs of preliminary clusters PC13 and PC27 with specificity for bovine platelets were studied by immunohistochemistry. Cryostat sections of bovine lymph node, spleen, thymus, small intestine, liver, kidney and smears of bone marrow cells were used. Five mAbs (CAPP2, IVA30, IVA125, IL-A164 and IL-A166) assigned to cluster PC13 (CD41/CD61) stained platelets and non-lymphocytic cells of various tissues. Our data confirm the presence of two specificities in PC27: three mAbs (IVA120, IVA197 and IVA198) specific for fibrinogen strongly reacted with the endothelial and reticular tissues whereas the other two mAbs Co-3D1D4 and Buf13 (WC13) were negative.

A polymorphic monoclonal anti-BoLA class I antibody.

Cytotoxic monoclonal antibody IVA 44 was generated after the intraperitoneal immunization with peripheral blood mononuclear (PBM) cells and the boost by the intrasplenic inoculation of skin graft. The detected membrane antigen isolated by immunoprecipitation appears to be composed of two subunits characteristic for the MHC class I molecules. The antibody IVA 44 exhibited a different reactivity: it recognized the BoLA A14 (A8) specificity in animals typed in the Fifth BoLA workshop, while it reacted with all A8 positive animals including subtypes A14 and A15 in Czech and Slovak cattle. It is concluded that mAb IVA 44 might detect the broad subtype of A8 covering A14 and certain A15 split(s). The diverse A15 reactivity of this mAb in the workshop and our population could be explained by the different occurrence of A15 splits in both populations.