Encapsulation of Clofazimine by Cyclodextran: Preparation, Characterization, and In Vitro Release Properties.

This study aimed to evaluate and compare the efficacy of cyclodextrans (CIs) and cyclodextrins (CDs) in improving the water solubility of a poorly water-soluble drug, clofazimine (CFZ). Among the evaluated CIs and CDs, CI-9 exhibited the highest percentage of drug inclusion and the highest solubility. Additionally, CI-9 showed the highest encapsulation efficiency, with a CFZ:CI-9 molar ratio of 0.2:1. SEM analysis indicated successful formation of inclusion complexes CFZ/CI and CFZ/CD, accounting for the rapid dissolution rate of the inclusion complex. Moreover, CFZ in CFZ/CI-9 demonstrated the highest drug release ratio, reaching up to 97%. CFZ/CI complexes were found to be an effective means of protecting the activity of CFZ against various environmental stresses, particularly UV irradiation, compared to free CFZ and CFZ/CD complexes. Overall, the findings provide valuable insights into designing novel drug delivery systems based on the inclusion complexes of CIs and CDs. However, further studies are needed to investigate the effects of these factors on the release properties and pharmacokinetics of encapsulated drugs in vivo, in order to ensure the safety and efficacy of these inclusion complexes. In conclusion, CI-9 is a promising candidate for drug delivery systems, and CFZ/CI complexes could be a potential formulation strategy for the development of stable and effective drug products.

Insights into the molecular interaction of cyclodextran with a guest molecule: A computational study.

Cyclodextran (CI) is a cyclic oligosaccharide in which d-glucose forms a ring structure by α-1,6 glycoside linkage and forms an inclusion complex with various guest molecules. In this study, we conducted quantum chemical calculations and molecular dynamics (MD) simulations to analyze the molecular interaction mechanism of CI with a guest molecule. Calculations of cyclodextrin (CD), in which d-glucose forms a ring structure by α-1,4 glycoside linkage, were also performed for comparison. Here, coenzyme Q10 (CoQ10), a promising molecule for industrial application with CI, was chosen as a guest molecule. Our MD simulation demonstrated that the molecular fluctuation was similarly large for both CI and CD when CoQ10 was absent. In the case that CoQ10 exists, the CD was found to form a rigid one-by-one inclusion structure, while CI does not. Additional simulations including multiple CI-8s indicated the possibility that an inclusion structure is maintained by the existence of other CI-8s.

Carbohydrate-binding module of cycloisomaltooligosaccharide glucanotransferase from Thermoanaerobacter thermocopriae improves its cyclodextran production.

Thermoanaerobacter thermocopriae-derived thermostable cycloisomaltooligosaccharide (CI)-forming enzymes catalyze the production of CIs from dextran. The primary structure of the enzyme is comprised of CI glucanotransferase (TtCITase) at the N-terminal region and long isomaltooligosaccharide-forming enzyme (TtTGase) at the C-terminal region connected by carbohydrate-binding module family 35 (CBM, TtCBM). Three truncated mutants of CI-forming enzymes were successfully produced in Corynebacterium glutamicum, a food-grade host system, and their biochemical properties were characterized. The enzymes had optimum at pH 6.0 and pH-stability (5.0-12.0). Three enzymes had optimum temperature over 55 °C and they maintained 80% activity at 55 °C for 2 h, 12 h, and 18 h, respectively. Enzymes without CBM showed weaker allosteric behavior than those of other enzymes, which suggests the important role of CBM in allosteric behavior. However, CBM bearing enzymes showed high production of CIs with various degree of polymerization. These enzymes have potential application as the encapsulating material for insoluble pharmaceutical biomaterials.

Carboxy-Terminal Region of a Thermostable CITase from Thermoanaerobacter thermocopriae Has the Ability to Produce Long Isomaltooligosaccharides.

Isomaltooligosaccharides (IMOs) have good prebiotic effects, and long IMOs (LIMOs) with a degree of polymerization (DP) of 7 or above show improved effects. However, they are not yet commercially available, and require costly enzymes and processes for production. The Nterminal region of the thermostable Thermoanaerobacter thermocopriae cycloisomaltooligosaccharide glucanotransferase (TtCITase) shows cyclic isomaltooligosaccharide (CI)-producing activity owing to a catalytic domain of glycoside hydrolase (GH) family 66 and carbohydrate-binding module (CBM) 35. In the present study, we elucidated the activity of the C-terminal region of TtCITase (TtCITase-C; Met740-Phe1,559), including a CBM35-like region and the GH family 15 domain. The domain was successfully cloned, expressed, and purified as a single protein with a molecular mass of 115 kDa. TtCITase-C exhibited optimal activity at 40°C and pH 5.5, and retained 100% activity at pH 5.5 after 18-h incubation. TtCITase-C synthesized α-1,6 glucosyl products with over seven degrees of polymerization (DP) by an α-1,6 glucosyl transfer reaction from maltopentaose, isomaltopentaose, or commercialized maltodextrins as substrates. These results indicate that TtCITase-C could be used for the production of α-1,6 glucosyl oligosaccharides with over DP7 (LIMOs) in a more cost-effective manner, without requiring cyclodextran.

Thermostable CITase from Thermoanaerobacter thermocopriae shows negative cooperativity.

OBJECTIVE: The biochemical properties of a putative thermostable cycloisomaltooligosaccharide (CI) glucanotransferase gene from Thermoanaerobacter thermocopriae were determined using a recombinant protein (TtCITase) expressed in Escherichia coli and purified to a single protein. RESULTS: The 171-kDa protein displayed maximum activity at pH 6.0, and enzyme activity was stable at pH 5.0-11.0. The optimal temperature was 60 °C in 1 h incubation, and thermal stability of the protein was 63% at 60 °C for 24 h. TtCITase produced CI-7 to CI-17, as well as CI-18, CI-19, and CI-20, which are relatively large CIs. Additionally, an unusual kinetic feature of TtCITase was its negative cooperative behavior in the dextran T2000 cleavage reaction. CONCLUSIONS: Based on our results, TtCITase can be applied to produce relatively large CIs at high temperature.

Biochemical Characterization of Alkaliphilic Cyclodextran Glucanotransferase from an Alkaliphilic Bacterium, Paenibacillus daejeonensis.

Cycloisomaltooligosaccharide glucanotransferase (CITase) was isolated from alkaliphilic Paenibacillus daejeonensis via an amino acid homology search for the reported CITase. The recombinant alkaliphilic CITase (PDCITase) from P. daejeonensis was expressed in an Escherichia coli expression system and purified as a single protein band of 111 kDa. PDCITase showed optimum activity at pH 8.0 and retained 100% of activity within a broad pH range (7.0-11.5) after 18 h, indicating alkaliphilic or alkalistable CITase properties. In addition, PDCITase produced CI-7 to CI-17, CI-18, and CI-19, which are relatively large cycloisomaltooligosaccharides yet to be reported. Therefore, these large cycloisomaltooligosaccharides can be applied to the improvement of water solubility of pharmaceutical biomaterials.

PEGylation of cationic, shell-crosslinked-knedel-like nanoparticles modulates inflammation and enhances cellular uptake in the lung.

The airway provides a direct route for administration of nanoparticles bearing therapeutic or diagnostic payloads to the lung, however optimization of nanoplatforms for intracellular delivery remains challenging. Poly(ethylene glycol) (PEG) surface modification improves systemic performance but less is known about PEGylated nanoparticles administered to the airway. To test this, we generated a library of cationic, shell crosslinked knedel-like nanoparticles (cSCKs), including PEG (1.5 kDa PEG; 2, 5, 10 molecules/polymer arm) on the outer shell. Delivery of PEGylated cSCK to the mouse airway showed significantly less inflammation in a PEG dose-dependent manner. PEGylation also enhanced the entry of cSCKs in lung alveolar epithelial cells and improved surfactant penetration. The PEGylation effect could be explained by the altered mechanism of endocytosis. While non-PEGylated cSCKs used the clathrin-dependent route for endocytosis, entry of PEGylated cSCK was clathrin-independent. Thus, nanoparticle surface modification with PEG represents an advantageous design for lung delivery. FROM THE CLINICAL EDITOR: In this study, the effects of PEGylation were studied on cross linked knedel-like nanoparticles in drug delivery through the lungs, demonstrating less airway inflammation in the studied model than with non-PEGylated nanoparticles, which suggests an overall favorable profile of PEGylated nanoparticles for alveolar delivery.