muramidase and galactomannan

In the present study, lysozyme-galactomannan conjugate (LGC) was fractionated by ion-exchange chromatography, the immune activity of the fractions was confirmed, and a structural analysis of the glycoprotein was performed. A high-molecular-weight fraction of LGC (H-LGC), was characterized by using a method using matrix-assisted laser desorption/ionization time of flight mass spectrometry. The glycated site of H-LGC was determined to be the lysine (Lys)115 residue. In addition, about 1mol of galactomannan (G) was linked to 1mol of lysozyme (L) in LGC based on the binding weight ratio. Conjugation of L and G reduced the aggregation of particles, resulting in a monodispersion based on measurement of dynamic light scattering. LGC in solution showed heterogeneous shapes with a mean size of 337nm. Therefore, we suggest that LGC improves the immune-enhancing activity as G conjugates the site of Lys115 on L, and provides higher solubility with reduced aggregation for the industrial use of LGC as a food constituent.

Topics: Animals; Chromatography, Ion Exchange; Galactose; Glycoproteins; Immunologic Factors; Macrophages; Maillard Reaction; Mannans; Mass Spectrometry; Mice; Muramidase; RAW 264.7 Cells; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

We describe a new aerogel obtained from laccase-oxidized galactomannans of the leguminous plant fenugreek (Trigonella foenum-graecum) and suggest its potential practical use. Laccase/TEMPO oxidation of fenugreek in aqueous solution caused a viscosity increase of over 15-fold. A structured, elastic, stable hydrogel was generated, due to formation of carbonyl groups from primary OH of galactose side units and subsequent establishment of hemiacetalic bonds with available free hydroxyl groups. Upon lyophilization of this hydrogel, a water-insoluble aerogel was obtained (EOLFG), capable of uptaking aqueous or organic solvents over 20 times its own weight. The material was characterized by scanning electron microscopy, FT-IR, elemental analysis and (13)C CP-MAS NMR spectroscopy and its mechanical properties were investigated. To test the EOLFG as a delivery system, the anti-microbial enzyme lysozyme was used as model active principle. Lysozyme was added before or after formation of the aerogel, entrapped or absorbed in the gel, retained and released in active form, as proven by its hydrolytic glycosidase activity on lyophilized Micrococcus lysodeikticus cells wall peptidoglycans. This new biomaterial, composed of a chemo-enzymatically modified plant polysaccharide, might represent a versatile, biocompatible "delivery system" of active principles in food and non-food products.

Topics: Cyclic N-Oxides; Diffusion; Drug Carriers; Freeze Drying; Galactose; Gels; Hydrolysis; Laccase; Mannans; Mechanical Phenomena; Muramidase; Oxidation-Reduction; Solubility; Trigonella; Water

We studied the immune-modulating effect of Maillard-type lysozyme-galactomannan conjugate (LGC). LGC significantly induced nitric oxide, and expressions of tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-8 on the murine macrophage Raw 264.7 cell line. In the mechanism of LGC, while extracellular signal-regulated kinase (ERK) was important for the induction of TNF-α, IL-1β and IL-8, the phosphorylation of C-Jun NH2-termianl kinase (JNK) contributed to the induction of TNF-α and IL-1β to a greater degree. These cytokines were less sensitive to the inhibition of p38. Nuclear factor (NF)-κB was involved in the induction of TNF-α and IL-1β. These data indicate that LGC has immune-modulating effects via JNK, ERK and NF-κB pathways, and that LGC may contribute to host immune defense.

Topics: Animals; Cell Survival; Cytokines; Extracellular Signal-Regulated MAP Kinases; Galactose; Immunologic Factors; JNK Mitogen-Activated Protein Kinases; Mannans; Mice; Muramidase; NF-kappa B; Nitric Oxide; Signal Transduction

Protein-polysaccharide conjugate was prepared as a functional biopolymer using protein and polysaccharide via a Maillard-type reaction. Ovalbumin and lysozyme were conjugated with galactomannan under controlled heating and humidity conditions. The antioxidant effect of ovalbumin and the antimicrobial activity of lysozyme were enhanced by the glycosylation. The emulsifying properties of the egg protein were also significantly improved by the modification. The increase in lipid affinity due to the conjugation resulted in the enhancement of the radical scavenging ability of ovalbumin. The effectiveness of lysozyme and its glycosylated derivative in restricting the activity of a Gram-negative pathogen, Edwardsiella tarda in fish was also investigated.

Topics: Animals; Anti-Bacterial Agents; Antioxidants; Biopolymers; Carps; Crystallization; Edwardsiella; Electrophoresis, Polyacrylamide Gel; Emulsions; Fish Diseases; Galactose; In Vitro Techniques; Lethal Dose 50; Maillard Reaction; Mannans; Muramidase; Mutagens; Ovalbumin; Rats

Hen egg white lysozyme was modified using the Maillard-type glycosylation method prior to the lipophilization with palmitic acid. The yield of lipophilized lysozyme significantly increased by the pre-glycosylation of the protein. The lipophilized lysozyme derivative was separated into two main fractions with different level of glycosylation. All fractions showed a strong antimicrobial activity against Gram-negative bacteria, Escherichia coli. The lipophilization of the lysozyme combined with glycosylation is a promising method for potential industrial applications of the lysozyme due to the enhanced antimicrobial activity and the improved yield.

Topics: Anti-Bacterial Agents; Carbohydrates; Egg White; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Freeze Drying; Galactose; Glycosylation; Maillard Reaction; Mannans; Microbial Sensitivity Tests; Micrococcus; Muramidase; Palmitic Acid

The protective effect of lysozyme-galactomannan or lysozyme-palmitic acid conjugates orally administered to carp, Cyprinus carpio L. was investigated using a virulent strain of Gram-negative Edwardsiella tarda isolated from an infected fish. Lysozyme-galactomannan conjugate was prepared through controlled Maillard reaction. Lysozyme-palmitic acid conjugate was prepared through base-catalyzed ester exchange using N-hydroxysuccinimide ester of palmitic acid. The conjugates provided substantial protection to carp infected with a Gram-negative bacteria fish pathogen E. tarda NG 8104. Lytic activities of lysozyme conjugates with galactomannan and palmitic acid were about 80 and 71% of native lysozyme using Micrococcus lysodeikticus as a substrate. Feeding with lysozyme conjugates, for 8 days, significantly enhanced fish protection against E. tarda infection. The survival rate was 30% for lysozyme-galactomannan conjugate treated fish and 20% for lysozyme-palmitic acid conjugate treated fish after 6 days cultivation while all control fish died within 3 days. On the other hand, a recovery rate of 40% after 6 days was observed in the fish group that were fed lysozyme-palmitic acid conjugate 3 and 2 h before and after E. tarda challenge, respectively, and for 6 consecutive days. The results of this work show the possibility of utilizing lysozyme conjugates with galactomannan or palmitic acid as a therapeutic for infection in fish.

Topics: Animals; Anti-Bacterial Agents; Fish Diseases; Galactose; Gram-Negative Bacteria; Gram-Negative Bacterial Infections; Maillard Reaction; Mannans; Microbial Sensitivity Tests; Muramidase; Palmitic Acid; Time Factors; Virulence