The biofilms formed by pathogenic microorganisms seriously threaten human health and significantly enhance drug resistance, which urgently call for developing drugs specifically targeting on biofilms. Chitooligosaccharides extracted from shrimp and crab shells are natural alkaline oligosaccharides with excellent antibacterial effects. Nevertheless, their inhibition efficacy on biofilms still needs to be improved. Spirulina (SP) is a microalga with negatively charged surface, and its spiral structure facilitates colonization in the depth of the biofilm. Therefore, the complex of Spirulina and chitooligosaccharides may play a synergistic role in killing pathogens in the depth of biofilm. This research first screened chitooligosaccharides with significant bactericidal effects. Subsequently, Spirulina@Chitooligosaccharides (SP@COS complex was prepared by combining chitooligosaccharides with Spirulina through electrostatic adsorption. The binding of the complex was characterized by zeta potential, z-average size, and fluorescence labeling. Ultraviolet-visible spectroscopy (UV-Vis) showed the encapsulation efficiency and the drug loading efficiency reached up to 90% and 16%, respectively. The prepared SP@COS2 exhibited a profound synergistic inhibition effect on bacterial and fungal biofilms, which was mainly achieved by destroying the cell structure of the biofilm. These results demonstrate the potential of Spirulina-chitooligosaccharides complex as a biofilm inhibitor and provide a new idea for addressing the harm of pathogenic microorganisms.
Humans ; Spirulina ; Anti-Bacterial Agents/chemistry* ; Chitosan/pharmacology* ; Biofilms ; Chitin/pharmacology*

Chitosan-carboxymethyl-chitosan complex film was prepared by freeze drying. Some tests in vivo and in animal were employed, in order to evaluate it on biology. All results indicated that the film has not only good surface compatibility but also good structural compatibility. It can be more suitable for GTR technology.
Animals ; Biocompatible Materials ; chemical synthesis ; pharmacology ; Chitin ; analogs & derivatives ; Chitosan ; Materials Testing ; Membranes, Artificial ; Rabbits ; Rats ; Skin Irritancy Tests

In this work a system which consists of chitosan (CS) microcores entrapped within enteric polymer is presented. Vitamin D2, used as a model drug, was efficiently entrapped within CS microcores using spray-drying and then microencapsulated into ethylic cellulose(EC). The morphology and release properties of microcapsules were tested. The influential factors of preparation conditions included molecular weight of chitosan, concentration of chitosan solution, concentration of acetic acid, loading of vitamin D2 were discussed. The results of in vitro release studies showed that the microcapsules prepared in this article could realize sustained release in intestine.
Capsules ; Cellulose ; analogs & derivatives ; pharmacology ; Chitin ; analogs & derivatives ; pharmacology ; Chitosan ; Delayed-Action Preparations ; Drug Compounding ; Drug Delivery Systems ; Ergocalciferols ; pharmacology ; In Vitro Techniques

Chitinases were produced by a lot of microorganisms. Chitinase gene expression in most of the chitinase producing bacteria was inducible by chitin. Low levels of chitinase were observed in the presence of glucose. To date, however, the regulation of such chitinase gene in Bacillus thuringiensis had not been well studied. In this paper, all 77 Bacillus thuringiensis strains were grown in the medium with or without chitin. We measured quantitatively the chitinase activity of the cultures. Moreover, we investigated the suppressive effect of glucose on chitinase of 4 strains. Also we studied the relationship between chitin induction and glucose suppression on chitinase. This investigation demonstrated that all tested B. thuringiensis strains could produce chitinase without chitin. After induction, the chitinolytic activity of 31 tested strains had no obvious response to the inducer, whereas 44 stains increased in different degree. Among these strains, most of them did not markedly increase the levels of chitinase, and many stains simultaneously displayed the expression mode of inducible and constitutive. The glucose inhibited the inductive effect of chitin, but it could not inhibit the basal expression of chitinase. Two strains No. 38 and No. 75 belonged to different expression types. But we just found several different bases in the regulatory region of chitinase genes chiA and chiB from them.
Bacillus thuringiensis ; enzymology ; growth & development ; Base Sequence ; Chitin ; pharmacology ; Chitinases ; biosynthesis ; genetics ; Culture Media ; chemistry ; Culture Techniques ; Glucose ; pharmacology ; Molecular Sequence Data ; Polymorphism, Genetic

P-glycoprotein (P-gp) located in the apicalmembranes of intestinal absorptive cells is an energy-dependent efflux pump which can reduce the bioavailability of a wide range of substrate drugs. There is increasingly interest in enhancing the bioavailability of these molecules by inhibiting intestinal P-gp. A classification of excipient inhibitors of intestinal P-gp nonionic surfactants, poly (ethylene glycol), derivates of beta-cyclodextrin and thiolated chitosan will be presented and then the inhibition mechanism will be discussed. Compared with traditional P-gp inhibitor, excipient inhibitors appear to have minimal nonspecific pharmacological activity, thus potential side effects can be mostly avoided. These excipient inhibitors, which hold the promise of replacing the traditional ones, will be extensively employed to significantly improve the intestinal absorption of poorly soluble and absorbed drugs as a result of P-gp inhibition, and thus to enhance the bioavailability of these drugs. However, the further studies of both the mechanism and clinical application are urgently needed.
ATP-Binding Cassette, Sub-Family B, Member 1 ; antagonists & inhibitors ; pharmacokinetics ; Animals ; Biological Availability ; Chitin ; analogs & derivatives ; pharmacology ; Excipients ; pharmacology ; Glycerol ; analogs & derivatives ; pharmacology ; Humans ; Intestinal Absorption ; drug effects ; Polyethylene Glycols ; pharmacology ; Surface-Active Agents ; pharmacology ; beta-Cyclodextrins ; pharmacology

OBJECTIVETo evaluate the cytotoxicity of polylactic acid-chitin plates in vitro.

METHODSThe cytotoxicity of polylactic acid-chitin plates was evaluated with MTT assay and direct contact assay using the cell line L929 in vitro. L929 cells were cultured with different concentrations of the extracts of polylactic acid-chitin plates (0, 50% and 100%, V/V) in vitro, and MTT assay was performed to evaluate the cell proliferation at 2, 4 and 7 days. The morphologic changes of the cells were observed by inverted microscope and scanning electron microscope.

RESULTSMTT assay revealed that the extracts of polylactic acid-chitin plates enhanced the cell proliferation (P<0.01), but not in a dose-dependent manner (P>0.05). In the direct contact assay, no side effects were found in regard to the cell growth, proliferation and attachment to the polylactic acid-chitin plates as compared with the negative control. The cytotoxicity grade of polylactic acid-chitin plates was 0 or 1, suggesting that the material was almost free of cytotoxicity.

CONCLUSIONThe polylactic acid-chitin plate possesses ideal cellular compatibility, and therefore has great potential for clinical use as an internal bone fixation material.

Animals ; Biocompatible Materials ; pharmacology ; Cell Line ; Cell Proliferation ; drug effects ; Cell Survival ; drug effects ; Chitin ; pharmacology ; Fibroblasts ; cytology ; ultrastructure ; Fracture Fixation, Internal ; instrumentation ; Lactic Acid ; pharmacology ; Materials Testing ; Microscopy, Electron, Scanning ; Polyesters ; Polymers ; pharmacology ; Time Factors

This study examined the effectiveness of Holmium-166 (Ho-166) chitosan complex therapy for a malignant glioma. Cultured C6 glioma cells (100, 000 in 5microliter) were injected into the caudate/putamen of 200 - 250 gram Wistar rats. Five days later, a Ho-166 chitosan complex was injected into the same site of the glioma injection. Four injection doses were administered: the control group received PBS 10microliter, group 1 received an injection of 100micro Ci (10microliter), group 2 received an injection of 50microCi (5microliter), and group 3 received an injection of 10micro Ci (1microliter). The average tumor volume for each group was 1.385 mm3 for the control group, 0.036 mm3 for group 1, 0.104 mm3 for group 2, and 0.111 mm3 for group 3. Compared with the control group, the size of the tumors in groups 1, 2 and 3 was reduced by an average of 97.4%, 92.5% and 91.9%, respectively. The Kaplan-Meier survival curve of group 2 was the longest, followed by groups 3, group 1 and the control. The mean survival was 22.8, 59, 60, and 44.6 days for the control group and groups 3, 2 and 1, respectively. H-E staining revealed that group 2 yielded the best results in the destruction of the malignant glioma. TUNEL staining and immunohistochemical studies indicated apoptotic features. The Ho-166 chitosan complex proved to be effective in destroying the malignant glioma.
Animals ; Brachytherapy ; Brain Neoplasms/mortality/pathology/*radionuclide imaging ; Cell Line, Tumor ; Chitin/analogs & derivatives/*pharmacology ; Disease Models, Animal ; Glioma/mortality/pathology/*radionuclide imaging ; Holmium/*pharmacology ; Radioisotopes/*pharmacology ; Rats ; Rats, Wistar ; Research Support, Non-U.S. Gov't

OBJECTIVETo discuss the electrophysiologic effects of regenerative nerve fibres affected by control releasing of FK506.

METHODSFrom Mar. to Sep. in 2008, the body weigh of 32 Sprague-Dawley rats which was 200 to 250 g,anesthesia was performed with an intraperitoneal injection of 30 mg/kg 1% continal. The sciatic nerve was transected in each rat by the excision of a 10 mm gap just proximal to the trifurcation of the nerve. The 10 mm gap of sciatic nerve had been bridged with the new double channel nerve conduit of fusiform shape, which were randomly divided into two groups basing on the different drug in the channel, each group contained 16 animals. In group A,100 microl of chitin for medical use was injected into the conduit,in group B the two branches of the conduit respectively contained 100 microl of the chitin and 10 microl FK506 (group B2) or physiologic saline (group B1). At 8 and 12 week after operation, the morphology in regenerative nerve and electrophysiologic effects by detect compound muscle active potential (CMAP) and cortical somatosensory evoked potential (CSEP) were evaluated.

RESULTSThere were not significant differences of the regenerative nerve fibres between two channels in group A, but in group B2, the number of the regenerative fibres was much more than that in group B1. The latency of CMAP and CSEP in group B2 was shorter than that in group B1. But its amplitude was higher. There were highly significant difference between the groups (P < 0.01).

CONCLUSIONThe electrophysiologic effects of regenerative nerve fibres can be significantly promoted by FK506, which provide theory base for immunosuppressive treatment of peripheral nerve.

Animals ; Chitin ; administration & dosage ; Delayed-Action Preparations ; Female ; Immunosuppressive Agents ; administration & dosage ; pharmacology ; Male ; Nerve Fibers ; drug effects ; physiology ; Nerve Regeneration ; Rats ; Rats, Sprague-Dawley ; Tacrolimus ; administration & dosage ; pharmacology

OBJECTIVETo study the mechanism of oligochitosan-induced macrophage activation.

METHODSOligochitosan was chemically modified with fluorophore 2-aminoacridone (2-AMAC). The cellular events of 2-AMAC-oligochitosan-macrophage interaction were analyzed with confocal laser microscopy and the fluorescence intensity of cells was analyzed by BD LSR flow cytometer. The mechanism of oligochitosan uptake by macrophages was studied by competitive inhibition test and the effect of calcium, trypsin and colchicine on oligochitosan recognition and internalization were also determined. RT-PCR was performed to investigate the level of TNF-alpha secretion.

RESULTMacrophage could bind and uptake oligochitosan, which was dependent on the temperature: the uptake proceeded rapidly at 37 degrees C and at 4 degrees C macrophage could only bind oligochitosan. EDTA decreased oligochitosan uptake. Trypsin treatment significantly reduced the internalization, and uptake was recovered by trypsin termination. Colchicine significantly inhibited the internalization process and was dose dependent. 0.1 mol/L mannose inhibited TNF-alpha expression induced by oligochitosan.

CONCLUSIONMacrophage could uptake oligochitosan via mannose receptor mediated pinocytosis. Mannose receptor is crucial for the oligochitosan-induced macrophages activation.

Cells, Cultured ; Chitin ; analogs & derivatives ; pharmacology ; Humans ; Lectins, C-Type ; metabolism ; Macrophage Activation ; drug effects ; Macrophages ; cytology ; Mannose-Binding Lectins ; metabolism ; Pinocytosis ; drug effects ; Receptors, Cell Surface ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

AIMTo study the relationship between cellular membrane fluidity and relative bioavailability (Fr) of protein and peptide drugs combined with absorption enhancers after pulmonary administration in rats.

METHODSA series of model drug salmon calcitonin (sCT) solutions with 6 absorption enhancers (Brij78, sodium cholate, sodium caprylate, 2-hydroxypropyl-beta-cyclodextrin, lecithin and chitosan) were prepared and then delivered to rats by pulmonary route. Serum drug concentration was determined by radioimmunoassay method. Using the techniques of electron spin resonance and fluorescence polarography, the effects of enhancers on pulmonary cellular membrane fluidity were investigated.

RESULTSFr values of sCT solution with some absorption enhancers (Brij78, sodium cholate, sodium caprylate, lecithin and chitosan) were significantly higher than those without enhancers. Brij78, lecithin and sodium caprylate, not only increased membrane lipid fluidity but also loosed the constitution of membrane protein. The effect of sodium cholate on membrane protein was low. Lipid fluidity was reduced and protein constitution was changed markedly, after pulmonary cellular membrane was treated by 0.5% chitosan solution. This result showed that the absorption enhancing of chitosan mainly came from its effects on membrane protein. Corresponded with lower Fr after pulmonary administration, 2-hydroxypropyl-beta-cyclodextrin (0.5% and 3%) had not significant effects on both lipid fluidity and protein constitution.

CONCLUSIONThe effects of enhancers on pulmonary absorption of peptide drugs in vivo might be investigated on the grounds of determination of cellular membrane fluidity in vitro.

Absorption ; Animals ; Biological Availability ; Calcitonin ; administration & dosage ; pharmacokinetics ; Caprylates ; pharmacology ; Cell Membrane ; metabolism ; Chitin ; analogs & derivatives ; pharmacology ; Chitosan ; Drug Synergism ; Lung ; metabolism ; Male ; Membrane Fluidity ; drug effects ; Membrane Proteins ; drug effects ; Molecular Conformation ; Phosphatidylcholines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sodium Cholate ; pharmacology