Delineating the inherent functional descriptors and biofunctionalities of pectic polysaccharides.

Pectin is a plant-based heteropolysaccharide macromolecule predominantly found in the cell wall of plants. Pectin is commercially extracted from apple pomace, citrus peels and sugar beet pulp and is widely used in the food industry as a stabilizer, emulsifier, encapsulant, and gelling agent. This review highlights various parameters considered important for describing the inherent properties and biofunctionalities of pectins in food systems. These inherent descriptors include monosaccharide composition, galacturonic acid content, degree of esterification, molecular weight, structural morphology, functional group analysis, and functional properties, such as water and oil holding capacity, emulsification, foaming capacity, foam stability, and viscosity. In this study, we also delineate their potential as a nutraceutical, prebiotic, and carrier for bioactive compounds. The biofunctionalities of pectin as an anticancer, antioxidant, lipid-lowering, and antidiabetic agent are also conceptually elaborated in the current review. The multidimensional characteristics of pectin make it a potential candidate for use in food and biomedical science.

Human oocytes: maturation in chemically defined media.

Human oocytes from ovarian follicles resume meiosis in F10, a defined medium, in numbers comparable to that obtained in medium containing serum. Several simple Krebs-Ringer media also support maturation, which suggests a similarity of nutritional requirements between human and mouse oocytes. Fewer oocytes reach metaphase II when the investing follicular cells are removed prior to culture.

Cr(VI) adsorption on the blends of Henna with chitosan microparticles: Experimental and statistical analysis.

In this work, the capability of blends of Henna with chitosan microparticles as novel adsorbent was studied in order to remove Cr(VI) ions. Response surface method (RSM) as a statistical technique was used to minimize number of experiments (26 runs), and optimize the operating conditions. In order to study Cr(VI) adsorption on the adsorbent, the effects of process parameters namely pH (2-9), initial Cr(VI) solution concentration (10-100 mg/L), Henna dose (0.1-1 g/L), chitosan microparticles dose (0.1-1 g/L) and contact time (20-150 min) were carefully considered. The adsorption did not change from pH 2 to 9 while it was reduced from 100% to 98.67% by growing Cr(VI) initial concentration. In addition, the adsorption was increased from 98.04% to 100% by increasing the adsorbent amount. The result of the isotherm study fitted well to Langmuir model (with R2 = 0.9826).

Immobilization of yeast inulinase on chitosan beads for the hydrolysis of inulin in a batch system.

An extracellular inulinase was partially purified by ethanol precipitation and gel exclusion chromatography from a cell free extract of Kluyveromyces marxianus. Partially purified inulinase exhibited 420 IU/mg specific activity and it was immobilized on chitosan beads. Activity yield of immobilized inulinase was optimized with glutaraldehyde concentration (1-5%), glutaraldehyde treatment time (30-240min), enzyme coupling-time (2-16h) and enzyme loading (5-30 IU) as functions. Under the optimized conditions maximum yield 65.5% of immobilized inulinase was obtained. Maximum hydrolysis of inulin 84.5% and 78.2% was observed at 125rpm after 4h by immobilized and free enzyme, respectively. A retention-time of 4h and 5h was found optimal for the hydrolysis of inulin under agitation (125rpm) by free and immobilized enzyme, respectively. The recycling of the developed immobilized biocatalyst was carried out after 5h of inulin hydrolysis in a batch system. The developed immobilized biocatalyst was successfully used for the hydrolysis of inulin for 14 batches. This is the first report on the immobilization of yeast inulinase on chitosan beads for the hydrolysis of inulin in a batch system.

Active water-insoluble derivatives of papain and other enzymes based on preformed diazonium-type supports.

Papain (EC 3.4.22.2) has been coupled to supports of titanium (IV) oxide and cellulose, which are particulate and pre-coated with diazotised 1,3-diaminobenzene, giving water-insoluble and stable derivatives which possess low proteolytic activity but high esterolytic activity. In addition the reversible binding of zinc (II) at the active site of papain has been exploited to inhibit protectively the enzyme during its linkage to the aforementioned supports, thereby yielding water-insoluble derivatives of papain having superior activity upon reactivation with EDTA. Application of the improved procedure of enzyme coupling to macroporous cellulose particles gave a water-insoluble derivative of papain having further enhanced proteolytic activity. Other properties of the water-insoluble derivatives of papain and of similarly prepared water-insoluble conjugates of urease (EC 3.5.1.5) and cholinesterase (EC 3.1.1.8) with cellulose are also reported.

Immobilization of glucoamylase on gelatin by transition-metal chelation.

The potential applicability of glutaraldehyde-crosslinked-gelatin particles for the immobilization of enzymes by encapsulation has been extended by addition of surface-bound enzyme, leading ultimately to a method for the preparation of dual immobilized enzyme conjugates. Attachment of enzyme to the surface of the capsules was achieved by a transition-metal chelation process in which the incoming enzyme becomes a ligand. Glucoamylase was so immobilized, using titanium-urea, -acrylamide, -citric acid, and -lactose complexes or titanium (IV) chloride as means of introducing the titanium chelating centre. The retentions of enzyme activity for both the surface-bound and pre-encapsulated enzymes were functions of the chelating complex chosen. Differences were observed between the action patterns of the two forms of immobilized enzyme. These action patterns and the production of reversion products are discussed in the light of application of gelatin-immobilized glucoamylase to the production of high-DE glucose syrups.

The use of cellulose carbonate-based immunoadsorbents in the isolation of minor allotypic components of rabbit immunoglobulin populations.

Cellulose trans-2,3-carbonate has been used as a new insoluble matrix for the simple coupling of a1- and b4-positive rabbit immunoglobulin to make immunoadsorbents capable of purifying from serum, with great efficiency, alloantibodies to these allotypic determinants. The antibodies have themselves been conjugated to prepare specific antibody immunoadsorbents of high binding activity for their allotypic target molecules. With these anti-allotypic solid-phase reagents it has been possible to affinity purify a1- and b4-positive immunoglobulin molecules and to deplete serum immunoglobulin of these molecules to leave in the eluates only the allotypically uncontaminated minor immunoglobulin components which are a-negative or b-negative (lambda chain-bearing) molecules. lambda chain molecules were also purified in very small quantities by affinity chromatography on a sheep anti-rabbit lambda chain column. This method of purifying minor populations of rabbit immunoglobulin from normal serum by special immunoadsorbent applications offers new opportunities to study the products of rarely expressed immunoglobulin genes in normal rabbits.

Advances in enzymatic transformation of penicillins to 6-aminopenicillanic acid (6-APA).

This article elaborates on the important recent developments in the enzymatic transformation of penicillins to 6-aminopenicillanic acid (6-APA), which is the basic raw material for the industrial production of semisynthetic penicillins such as amoxycillin and ampicillin. Particular emphasis is placed on the improvements in purification, stability, and immobilization of the enzymes, (i.e. penicillin acylases) used for these transformations.

Ovarian dysgenesis and chromosome abnormalities.

The association of ovarian hypoplasia with trisomy 18 in 6 infants and trisomy 13 in 5 infants and 1 fetus is reported. Three infants also had thymic hypoplasia. The possible etiology of ovarian dysgenesis is discussed in relation to a variety of clinical and experimental associations.

Immobilization of pneumococcal polysaccharide vaccine on silicon oxide wafer for an acoustical biosensor.

One the most important aspects of a biosensor is related to immobilization and maintenance of specific reference compounds on sensing surfaces. A method for the immobilization of polysaccharides to a silicon oxide surface intended for Surface Acoustical Waves (SAW) sensors is described. Silicon oxide is a hydrophobic inorganic support used for the fabrication of many electronic devices. The pneumococcal polysaccharide (PPS) vaccine is immobilized via Protein A after pre-treatment of the surface with hydrochloric acid. The effects of non-specific binding are discussed. The results indicate that the immobilization of PPS via Protein A increases the sensitivity of detecting Streptococcus pneumoniae antibodies in human sera and offers greater reproducibility of response compared with ELISA methods. The principles of this technique are simple and are applicable to the immobilization of many capsular polysaccharides.

Development of gel filtration and specific analyses of urinary carbohydrate and protein material.

Hitherto investigations into urinary carbohydrate and proteinaceous macromolecular material has largely been restricted to non-dialysable fractions. We found that this limits the investigation to less than 1% of the total material present and we present a method for direct analysis of urine without prior concentration or extraction. This method extends existing techniques allowing deeper investigation into necessary and important chemical aspects of health and disease. The method is based on gel filtration using Bio-Gel P2. Means included to overcome the observed adsorption by the gel matrix of relevant material depend on use of high ionic concentration buffers for column elution. The problems of bacterial contamination of the gel matrix are discussed and the use of presterilised urine samples is proposed to prevent contamination. The coupling of the gel filtration column to three specific automated analyses allows simultaneous determination of uronic acid, neutral sugar and proteinaceous material.

Isolation of thyroxine-binding globulin (TBG) by immunoadsorption chromatography: some physical and immunochemical characteristics of TBG.

Thyroxine-binding globulin (TBG) was isolated from pooled whole human serum by diethylaminoethyl (DEAE) Sephadex anion exchange chromatography followed by immunoadsorption chromatography on a cyanogen bromide-activated Sepharose 4B-sheep anti-human TBG immunoadsorbent. Sodium dodecyl sulphate (SDS)-poly-acrylamide gel electrophoresis (PAGE) of the purified TBG revealed a major protein band with a molecular mass of 65 000 and a weak band of molecular mass 54 000 in both reducing and non-reducing buffers. Sedimentation velocity analysis revealed an S20,w coefficient of 4.5S and a calculated molecular mass of 60 000. Immunochemical analysis confirmed the purity of the TBG preparation which gave a single precipitin peak on two-dimensional immunoelectrophoresis.

In vitro interaction between psychotropic drugs and alcohol dehydrogenase activity.

A series of CNS-stimulating and -depressant drugs have been studied for their in vitro interaction with horse liver alcohol dehydrogenase (ADH) activity. The depressant drugs studied included barbital, phenobarbital, thiopental, nitrazepam, chlorpromazine, sulpiride, clomethiazole, Li2CO3, diazepam, phenytoin, ethosuximide, morphine, and codeine. The stimulant drugs were theophylline, caffeine, amphetamine, imipramine, chlorimipramine, amitriptyline, and tranylcypromine. The results were as follows. First, ADH activity was inhibited by the action of chlorpromazine, tranylcypromine, imipramine, chlorimipramine, amitriptyline, sulpiride, amphetamine, codeine, ethosuximide, morphine, clomethiazole, nitrazepam, Li2CO3, theophylline, and phenobarbital, in descending order of inhibitory effect. Second, inhibition followed by activation of ADH activity was observed for imipramine and chlorimipramine. Third, activation of ADH activity was observed for phenytoin. Finally, the following drugs were not seen to exert any effect on ADH activity: barbital, thiopental, diazepam, and caffeine.

The fully automatic ion-exchange and gel-permeation chromatography of neutral monosaccharides and oligosaccharides with a Jeolco JLc-6AH analyser.

Experiences with a Jeolco JLC-6AH carbohydrate analyser used to provide a fully automated analysis service for neutral mono- and oligo-saccharides have been summarised. Modifications have been made to the equipment to extend its application, and a method for routine separations of oligosaccharides (d.p. 2--15) has been established. The degree of separation achieved is far superior to that which may be obtained by gel-filtration techniques under gravity- or peristaltic-pump-aided flow. The versatility and adaptability of equipment for use with assays other than the standard orcinol-sulphuric acid reaction shows that it is not too inconvenient to interchange between various assays and separations.

Magnetic, immobilised derivatives of enzymes.

A novel, water, insoluble, stable coating has been attached to various metals, which is capable of covalent attachment to enzymes. The coating (aminobenzoic acid-formaldehyde resin) can be attached to nickel, cobalt, tin, iron, and aluminium. beta-D-Glucosidase has been attached to the coated metals by diazotisation of the coating. The stability and usefulness of the resulting, metallic enzyme-preparations is discussed with special emphasis on the value of ferromagnetic supports for enzymes.

Effects of titanium compounds on a D-glucose-D-glucose oxidase assay system.

Titanium compounds affect the measurement of D-glucose oxidase (and therefore D-glucose) by the D-glucose-D-glucose oxidase-peroxidase-2,2'-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) assay system. The validity of measurement of the activity of D-glucose oxidase immobilised on supports based on titanium oxides is affected by complexation of the intermediate hydrogen peroxide with the support, and such supports may prove to be unsuitable for the immobilisation of D-glucose oxidase. The formation of titanic peroxides is among the reasons discussed for the various interactions encountered. The use of the assay system for the determination of D-glucose oxidase contaminated with catalase and for the determination of hydrogen peroxide is also described.

The use of cellulose xanthate for the immobilisation of biological molecules.

The mercapto groups of cellulose xanthate can reversibly form disulphide bridges with L-cysteine. This property has been utilised for the immobilisation of a protein and an enzyme. These macromolecules, as polythiol derivatives, formed disulphide linkages with the matrix without serious disturbance of their active sites, became firmly bound to the xanthate, and were not eluted by normal washing conditions. Cellulose xanthate is a cheap, easily prepared matrix which permits a simple coupling reaction. The immobilisation process is selectively reversible.

The use of titanium (IV) oxide for the immobilisation of carbohydrate-directed enzymes.

The use of particles of porous titanium (IV) oxide as a suitable matrix for enzyme immobilisation has been investigated with dextranase. Treatment of the particles with enzyme in the presence and absence of ammonium ions showed that the presence of ammonia induced a greater coupling of protein, whereas a greater retention of enzyme specific activity was achieved in the absence of ammonia. Properties of the immobilised enzyme include a pH-dependence and reversibility of the coupling between enzyme and matrix. The immobilised dextranase was most stable at pH 5.0. Automated analytical techniques for measuring the activity of dextranase and other polysaccharidases in soluble and insoluble forms are also reported.

G.L.C. of the O-trimethylsilyl derivatives of hexuronic acids.

The free acids, sodium salts, and lactones of several hexuronic acids have been studied as their O-trimethylsilyl derivatives by gas-liquid chromatography using SE-30 and XE-60 liquid phases. Silylation was best performed in methyl sulphoxide. The equilibrium between the various forms of a hexuronic acid in methyl sulphoxide was also studied by g.l.c. following silylation. The hexamethyldisilazane used in the silylation disturbed the equilibrium attained in the solvent, but this was overcome by premixing the hexamethyldisilazane with chlorotrimethylsilane. Methyl sulphoxide and the silylating reagents gave a two-phase system in which the derivative was favourably partitioned into the upper layer. Partition coefficients and stabilities of the derivatives were measured, and a g.l.c. method for the analysis of the hexuronic acids was thereby developed. The oximes of the hexuronic acids were studied as alternative derivatives for g.l.c., and their equilibrium compositions and g.l.c. retention times are recorded.

Structural identification of isomeric O-trimethylsilyl derivatives of some hexuronic acids.

The structures of the isomeric products obtained on trimethylsilylation of naturally occurring hexuronic acids and their sodium salts and lactones have been established by the application of n.m.r. (for anomeric configuration) and mass spectrometry (for ring size). The equilibria of some of the hexuronic acids in methyl sulphoxide involved a larger proportion of furanoid forms than those in water or pyridine. The proportion of furanoses was also increased by the addition of hexamethyldisilazane. Kinetic evidence indicated that two molecules of each hexuronic acid interacted autocatalytically during mutarotation.