[Music therapy as an effective method of neurorehabilitation]. / Éffektivnost' muzykoterapii v reabilitatsii bol'nykh s insul'tom.

AIM: To assess the role of music therapy in the recovery of motor, speech and autonomic functions in patients with ischemic stroke (II). MATERIAL AND METHODS: Forty-five patients with II in the middle cerebral artery were examined. The patients were randomized into three groups (main, comparison and control) of 15 individuals each. With patients of the first and the second groups on the 3rd, 5th, 7th and 9th days of the rehabilitation period the special set of exercises with music and without that respectively was fulfilled. The third group received a basic set of physical exercises (a control group).The third group was control. Dynamics of patients' state was estimated by the NIHSS, the Rivermead Mobility Index, the Action Research Arm Test and the modified scale for speech evaluation on the 2nd, 4th, 6th, 8th and 10th day of disease. A study of cardiorespiratory synchronization was conducted since the 6th day of stroke. RESULTS AND CONCLUSION: The statistically significant efficacy of music therapy was shown for all parameters. The authors suggest that neuroplasticity may underlie the mechanisms of the programs used in the study.

[Genetic, phenotypic, and phytochemical polymorphism in Eastern European populations of Mentha arvensis L.]

Variability of M. arvensis from five geographically distanced populations was examined using morphological traits and phytochemical composition of essential oil and with the help of DNA fingerprinting using ISSR markers. The population differentiation based on morphological traits was weak. Analysis of the essential oil composition provided the subdivision of the sample into three groups and, on the basis of the composition of ISSR amplicons, into four groups of specimens. A high degree of genetic polymorphism of M. arvensis and substantial, though incomplete, population differentiation were identified. It was demonstrated that the population of M. arvensis from the Komi Republic was the most genetically isolated, while the populations from Moscow and Penza provinces were weakly differentiated from each other. The population from the Republic of Belarus (near Grodno) was genetically and phytochemically considerably different from the other studied populations, although morphologically indistinguishable from them. We argue that the differentiation was caused not only by the isolation by distance but also owing to the formation of three different ecotypes adapted to different climatic conditions.

A New Bacillus licheniformis Mutant Strain Producing Serine Protease Efficient for Hvdrolvqis of Sov Meal Proteins.

Induced mutagenesis with y-irradiation of the industrial strain Bacillus licheniformis-60 VKM B-2366,D was used to obtain a new highly active producer of an extracellular serine protease, Bacillus licheni- formis7 145. Samples of dry.concentrated preparations of serine protease produced by the original and mutant strains were obtained, and identity of their protein composition was'established. Alkaline serine protease sub- tilisin DY was the main component of the preparations. The biochemical and physicochemical properties of the Protolkheterm-145 enzyme preparation obtained from the mutant strain were studied. It exhibited pro- teolytic activity (1.5 times higher than the preparation from the initial strain) within broad ranges of pH (5- 11) and temperature (30-70'C).-Efficient hydrolysis of extruded soy meal protein at high concentrations (2 to 50%) in-the reaction mixture was.the main advantage of the Protolikheterm 145 preparation. Compared to,. the preparation obtained using the initial strain, the new preparation with increased proteolytic-activity pro- vided for more complete hydrolysis of the main non-nutritious soy,proteins.(glycinin and 0-conglycinin) with the yield of soluble protein increased by 19-28%, which decreased the cost of bioconversion of the protein- aceous material and indicated promise of the new preparation in resource-saving technologies for processing soy meals and cakes.

[Interstitial endocrine apparatus of testes of experimental animals in conditions of chromium-benzene intoxication].

With the use of histological, morphometric and statistical methods there was shown a gonadotropic effect of chromium, benzene and also their mixtures in male mice (CBA x C57Bl6) F1. The established structural changes in the testes of exposed animals showed the suppression of their germinative and endocrine functions. The response of Leydig cells in the chromium group expresses a development of the compensatory process in the relation with the destruction of seminiferous epithelium.

[Phenotypic and phytochemical differences between Mentha arvensis L. and Mentha canadiensis L].

A taxonomic study of anatomical, morphological, and phytochemical characteristics of Mentha arvensis L. and Mentha canadiensis L. using hierarchical cluster analysis has been conducted and the differences between the species studied have been revealed. The ratio between the lengths of the calyx tube and the calyx lobes, the number of secretory glands on the upper and lower surfaces of the leaf, and the composition of the essen- tial oil were shown to be the most appropriate parameters for classification.

Comparative study of biochemical properties of glucoamylases from the filamentous fungi Penicillium and Aspergillus.

Here we report the first isolation to homogeneous forms of two glucoamylases from the fungus Penicillium verruculosum and their study in comparison with known glucoamylases from Aspergillus awamori and Aspergillus niger. Genes that encode glucoamylases from P. verruculosum were cloned and expressed in the fungus Penicillium canescens, and the recombinant glucoamylases were obtained with subsequent study of their molecular weights, isoelectric points, optimal temperature and pH values, and stability. The catalytic activities of the recombinant glucoamylases were determined in relation to soluble potato starch. Changes in molecular mass distribution and content of low molecular weight products during starch hydrolysis by glucoamylases from P. verruculosum, A. awamori, and A. niger were studied. An exo-depolymerization mechanism was established to be the pathway for destruction of starch by the glucoamylases.

Cloning, purification, and characterization of galactomannan-degrading enzymes from Myceliophthora thermophila.

Genes of ß-mannosidase 97 kDa, GH family 2 (bMann9), ß-mannanase 48 kDa, GH family 5 (bMan2), and α-galactosidase 60 kDa, GH family 27 (aGal1) encoding galactomannan-degrading glycoside hydrolases of Myceliophthora thermophila C1 were successfully cloned, and the recombinant enzymes were purified to homogeneity and characterized. bMann9 displays only exo-mannosidase activity, the K(m) and k(cat) values are 0.4 mM and 15 sec(-1) for p-nitrophenyl-ß-D-mannopyranoside, and the optimal pH and temperature are 5.3 and 40°C, respectively. bMann2 is active towards galactomannans (GM) of various structures. The K(m) and k(cat) values are 1.3 mg/ml and 67 sec(-1) for GM carob, and the optimal pH and temperature are 5.2 and 69°C, respectively. aGal1 is active towards p-nitrophenyl-α-D-galactopyranoside (PNPG) as well as GM of various structures. The K(m) and k(cat) values are 0.08 mM and 35 sec(-1) for PNPG, and the optimal pH and temperature are 5.0 and 60°C, respectively.

[Bacteriolytic enzymes of blood plasma from sheep].

In the present work the studies ofbacteriolytic factors from sheep blood plasma have been performed. Three novel enzymes have been identified and characterized. Two of them have a molecular weight 15 +/- 2 kDa and able to lyse the gram-negative Escherichia coli bacteria. The third enzyme has a molecular weight 34 +/- 4 kDa and is able to lyse both gram-negative Escherichia coli and gram-positive Micrococcus luteus bacteria. The bacteriolytic reactions have been studied for all three enzymes; particularly, pH-optima have been identified with respect to the substrate. To identify the enzymes trypsinolysis and consequent MALDI-TOF mass spectrometry studies were performed. The results were compared to data from publicly available databases, such as Swiss-Prot, NCBI, MSDB.

[Creation of a heterologous gene expression system on the basis of Aspergillus awamori recombinant strain].

A heterologous gene expression system was created in a domestic Aspergillus awamori Co-6804 strain, which is a producer of the glucoamylase gene. Vector pGa was prepared using promoter and terminator areas of the glucoamylase gene, and A. niger phytase, Trichoderma reesei endoglucanase, and Penicillium canescens xylanase genes were then cloned into pGa vector. Separation of enzyme samples using FPLC showed the amount of the recombinant proteins to be within the 0.6-14% range of total protein.

[Influence of the cycle number in processing of cellulose from waste paper on its ability to hydrolysis by cellulases].

Hydrolytic ability of laboratory enzyme preparations from fungus of the Penicillium genus was investigated using kraft pulp from nonbleached softwood and bleached hardwood cellulose as substrates. The enzyme preparations were shown to efficiently hydrolyze both softwood and hardwood cellulose. The yields of glucose and reducing sugars were 24-36 g/l and 27-37 g/l from 100 g/l of dry substrate in 48 h, respectively, and depended on the number of substrate grinding cycles.

[Increase in glucoamylase productivity of Aspergillus awamori strain by combination of radiating mutagenesis and plasmid transformation methods].

Increase in the level of amylolytic genes activator protein encoded by amyR gene was shown to result in enhancement of glucoamylase productivity of A. awamori strain by 30%. However, the same effect equal to 30% increase can be achieved by introduction of extra copies of gla gene encoding glucoamylase. These two effects were not additive, which gave the possibility to suggest an additional limitation in the egulation mechanism of glucoamylase gene expression in Aspergillus family strains while introducing an additional copies of amyR and gla genes.

Isolation and properties of extracellular beta-xylosidases from fungi Aspergillus japonicus and Trichoderma reesei.

Homogeneous beta-xylosidases with molecular mass values 120 and 80 kDa (as shown by SDS-PAGE), belonging to the third family of glycosyl hydrolases, were isolated by anion-exchange, hydrophobic, and gel-penetrating chromatography from enzyme preparations based on the fungi Aspergillus japonicus and Trichoderma reesei, respectively. The enzymes exhibit maximal activity in acidic media (pH 3.5-4.0), and temperature activity optimum was 70 degrees C for the beta-xylosidase of A. japonicus and 60 degrees C for the beta-xylosidase of T. reesei. Kinetic parameters of p-nitrophenyl beta-xylopyranoside and xylooligosaccharide hydrolysis by the purified enzymes were determined, which showed that beta-xylosidase of A. japonicus was more specific towards low molecular weight substrates, while beta-xylosidase of T. reesei preferred high molecular weight substrates. The competitive type of inhibition by reaction product (xylose) was found for both enzymes. The interaction of the enzymes of different specificity upon hydrolysis of glucurono- and arabinoxylans was found. The beta-xylosidases exhibit synergism with endoxylanase upon hydrolysis of glucuronoxylan as well as with alpha-L-arabinofuranosidase and endoxylanase upon hydrolysis of arabinoxylan. Addition of beta-xylosidases increased efficiency of hydrolysis of plant raw materials with high hemicellulose content (maize cobs) by the enzymic preparation Celloviridine G20x depleted of its own beta-xylosidase.

Isolation and properties of fungal beta-glucosidases.

Using chromatography on different matrixes, three beta-glucosidases (120, 116, and 70 kDa) were isolated from enzymatic complexes of the mycelial fungi Aspergillus japonicus, Penicillium verruculosum, and Trichoderma reesei, respectively. The enzymes were identified by MALDI-TOF mass-spectrometry. Substrate specificity, kinetic parameters for hydrolysis of specific substrates, ability to catalyze the transglucosidation reaction, dependence of the enzymatic activity on pH and temperature, stability of the enzymes at different temperatures, adsorption ability on insoluble cellulose, and the influence of glucose on catalytic properties of the enzymes were investigated. According to the substrate specificity, the enzymes were shown to belong to two groups: i) beta-glucosidase of A. japonicus exhibiting high specific activity to the low molecular weight substrates cellobiose and pNPG (the specific activity towards cellobiose was higher than towards pNPG) and low activity towards polysaccharide substrates (beta-glucan from barley and laminarin); ii) beta-glucosidases from P. verruculosum and T. reesei exhibiting relatively high activity to polysaccharide substrates and lower activity to low molecular weight substrates (activity to cellobiose was lower than to pNPG).

Isolation and characterization of extracellular pectin lyase from Penicillium canescens.

Pectin lyase A (molecular weight 38 kD by SDS-PAGE, pI 6.7) was purified to homogeneity from culture broth of the mycelial fungus Penicillium canescens using chromatographic techniques. During genomic library screening, the gene encoding pectin lyase A from P. canescens (pelA) was isolated and sequenced, and the amino acid sequence was generated by applying the multiple alignment procedure (360 residues). A theoretical model for the three dimensional structure of the protein molecule was also proposed. Different properties of pectin lyase A were investigated: substrate specificity, pH- and temperature optimum of activity, stability under different pH and temperature conditions, and the effect of Ca2+ on enzyme activity. In the course of the laboratory trials, it was demonstrated that pectin lyase A from P. canescens could be successfully applied to production and clarification of juice.

[Use of a preparation from fungal pectin lyase in the food industry].

A new enzyme preparation of fungal pectin lyase (EC 4.2.2.10) was shown to be useful for the production of cranberry juice and clarification of apple juice in the food industry. A comparative study showed that the preparation of pectin lyase is competitive with commercial pectinase products. The molecular weight of homogeneous pectin lyase was 38 kDa. Properties of the homogeneous enzyme were studied. This enzyme was most efficient in removing highly esterified pectin.

[Hydrolysis of wheat flour with amylase preparations and individual enzymes].

Rheological properties of wheat flour were studied in the course of its processing (cooking and saccharification). The effects of commercial alpha-amylase preparations were compared during flour preparation. Test preparations were equally potent in decreasing the viscosity of an all-grain batch. Homogenous glucoamylases isolated from Aspergillus differed in the presence or absence of the starch-binding domain. The starch-binding domain provided for the high activity of glucoamylase on insoluble starch, but gave no advantages in saccharification of pretreated wheat flour.

[Application of neutral-alkaline pectate lyases to cotton fabric boil off].

Commercial and pilot pectate lyase preparations (EC 4.2.2.2) have been compared. They differ in their effect on pectins with different esterification degrees (ED). The activity of the pilot preparation with respect to a substrate with ED = 70% is tenfold lower than with respect to unesterified polygalacturonic acid. For commercial preparations, this activity ratio ranged within 1.5-2. At equal pectate lyase activities, the commercial preparations better remove pectin from crude cotton fabric during its boil off. The laboratory preparation is more efficient for improving the capillarity (wettability) of the fabric owing to the cooperative effect of the pectate lyase, cellulase, and hemicellulase present in the preparation.

Relationship between changes in the photoperiod and proliferation of Ehrlich ascitic tumor cells.

The effect of changed photoperiod (constant darkness) on the proliferation of Ehrlich ascitic tumor cells was demonstrated. Some mechanisms maintaining the proliferative homeostasis in Ehrlich ascitic tumor under conditions of constant darkness were studied, explaining more leveled pattern of circadian rhythms of cell proliferation in animals kept under conditions of constant darkness and indicating the existence of circadian rhythm in sensitivity of Ehrlich ascitic tumor cell to permanent darkness.

Isolation and properties of pectinases from the fungus Aspergillus japonicus.

Using anion-exchange chromatography on different carriers and phenyl-Sepharose hydrophobic chromatography, five pectolytic enzymes were isolated from the culture liquid of a mutant strain of Aspergillus japonicus: two endo-polygalacturonases (I and II, 38 and 65 kD, pI 5.6 and 3.3), pectin lyase (50 kD, pI 3.8), and two pectinesterases (I and II) with similar molecular weights (46 and 47 kD) and the same pI (3.8). The pectinesterases apparently represent two isoforms of the same enzyme. All purified enzymes were homogenous according to SDS-PAGE and polyacrylamide gel-IEF, except for endo-polygalacturonase II that gave two bands on isoelectric focusing, but one band on electrophoresis. All enzymes had maximal activity in an acid medium (at pH 4.0-5.5). The pectin lyase and pectinesterase were stable at 40-50 degrees C. The thermal stability of both endo-polygalacturonases was much lower (after 3 h of incubation at 30 degrees C, endo-polygalacturonases I and II lost 40 and 10% of the activity, respectively). The activity of endo-polygalacturonases I and II towards polygalacturonic acid strongly depended on NaCl concentration (optimal concentration of the salt was 0.1-0.2 M); the enzymes were also capable of reducing the viscosity of pectin solution, but rather slowly. The pectin lyase had no activity towards polygalacturonic acid. The activity of the pectin lyase increased with increasing degree of methylation of pectins. Both endo-polygalacturonases demonstrated synergism with the pectinesterase during the hydrolysis of highly methylated pectin. On the contrary, in the mixture of pectin lyase and pectinesterase an antagonism between the two enzymes was observed.

Viscometric method for assaying of total endodepolymerase activity of pectinases.

An improved method for assaying of the total endodepolymerase activity of pectinases has been developed. The method is based on the determination of the viscosity of a citrus pectin solution in the presence of the enzyme using an Ostwald viscometer. The depolymerizing activity of different pectinases can be detected including polygalacturonase, polymethylgalacturonase, pectin lyase, and pectate lyase. One unit of the endodepolymerase activity corresponds to the activity resulting in 50% decrease in the relative viscosity of 0.5% citrus pectin solution for 5 min at 40 degrees C and the appropriate pH. Depending on the pH-optima of the enzymes, two modifications of the method are described: 1) for acid pectinases at pH 5.0, and 2) for neutral (mildly alkaline) pectinases at pH 8.0. The modifications differed in the control and in the calculation of the activity. Six enzyme preparations were used to demonstrate the applicability of the method. The parameter used for the calculation of the enzymatic activity was directly proportional to the enzyme concentration (the dependence was linear in the range of at least 10-fold change in the enzyme concentration). The relative error of the method did not exceed 10%.