Preparation of Carboxymethyl Chitosan Microspheres and Their Application in Hemostasis

2015 ◽  
Vol 11 (6) ◽  
pp. 660-667 ◽  
Author(s):  
Lu Liu ◽  
Qi Lv ◽  
Qingyun Zhang ◽  
Hui Zhu ◽  
Wei Liu ◽  
...  
Keyword(s):  
Hepatic Injury ◽  
Carboxymethyl Chitosan ◽  
Water Soluble ◽  
Cross Linking ◽  
Clotting Time ◽  
Hemostatic Agents ◽  
Hemostatic Effect ◽  
Injury Model ◽  
Chemical Cross Linking

AbstractObjectiveChitosan (CS) is currently used as a hemostatic agent in emergencies and in military settings. However, its application is limited owing to its poor hydrophilia at neutral pH. Carboxymethyl chitosan (CMCS) is an important, water-soluble derivative of CS. In this study, we prepared CS and CMCS microspheres (CSMs and CMCSMs, respectively) and evaluated their hemostatic effect.MethodsTo prepare the microspheres of various sizes, we used the emulsion cross-linking technique. CMCSMs were also loaded with etamsylate (DIC). Clotting time in vitro and in a hepatic injury model was examined to evaluate the hemostatic effect.ResultsCMCSMs swelled more and clotted faster than did CSMs. CMCSMs loaded with DIC had no effect on hemostasis.ConclusionsBoth increasing material hydrophilicity and expanding the contact area promoted clotting, whereas chemical cross-linking hampered it because of decreased swelling. CMCSMs are promising candidates for the production of effective hemostatic agents. (Disaster Med Public Health Preparedness. 2017;11:660–667)

scholarly journals Mapping the Contact Sites of the Escherichia coli Division-Initiating Proteins FtsZ and ZapA by BAMG Cross-Linking and Site-Directed Mutagenesis

2018 ◽  
Vol 19 (10) ◽  
pp. 2928 ◽  
Author(s):  
Winfried Roseboom ◽  
Madhvi Nazir ◽  
Nils Meiresonne ◽  
Tamimount Mohammadi ◽  
Jolanda Verheul ◽  
...  
Keyword(s):  
Site Directed Mutagenesis ◽  
Cross Linking ◽  
Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Binding Interface ◽  
Tetrameric Structure ◽  
Z Ring ◽  
Cross Links ◽  
Chemical Cross Linking

Cell division in bacteria is initiated by the polymerization of FtsZ at midcell in a ring-like structure called the Z-ring. ZapA and other proteins assist Z-ring formation and ZapA binds ZapB, which senses the presence of the nucleoids. The FtsZ–ZapA binding interface was analyzed by chemical cross-linking mass spectrometry (CXMS) under in vitro FtsZ-polymerizing conditions in the presence of GTP. Amino acids residue K42 from ZapA was cross-linked to amino acid residues K51 and K66 from FtsZ, close to the interphase between FtsZ molecules in protofilaments. Five different cross-links confirmed the tetrameric structure of ZapA. A number of FtsZ cross-links suggests that its C-terminal domain of 55 residues, thought to be largely disordered, has a limited freedom to move in space. Site-directed mutagenesis of ZapA reveals an interaction site in the globular head of the protein close to K42. Using the information on the cross-links and the mutants that lost the ability to interact with FtsZ, a model of the FtsZ protofilament–ZapA tetramer complex was obtained by information-driven docking with the HADDOCK2.2 webserver.

Preparation of N, O-Carboxymethyl Chitosan with Different Substitutional Degree and its Application for Hemostasis

2013 ◽  
Vol 798-799 ◽  
pp. 1061-1066 ◽  
Author(s):  
Yan Wei Zhao ◽  
Lu Liu ◽  
Xiang Han ◽  
Jing Guan
Keyword(s):  
Carboxymethyl Chitosan ◽  
X Ray Diffraction ◽  
Clotting Time ◽  
X Ray ◽  
Angle Measurements ◽  
Contact Angle Measurements ◽  
Ft Ir ◽  
Ubbelohde Viscometer

We prepared N, O-carboxymethyl chitosans (CMCSs) with different substitutional degrees (SDs) to evaluate their effects of hemostasis, and provided experimental basis on biomedical materials. Chloroethanoic acid was used to synthesize CMCSs. The structure were characterized by Fourier transform infrared (FT-IR) and wide-angle X-ray diffraction (WXRD). Potentiometric titration and Ubbelohde viscometer were adopted to determine the SD and intrinsic viscosity of CMCSs. Contact angle measurements were investigated to determine surface wettability. Method of dynamic clotting time and coagulation test in vivo were used to evaluate their effects of hemostasis. SDs of CMCSs were from 50% to 110%. As the SD increased, molecular weight decreased. CMCS powder with SD 63% possessed excellent hemostasis both in vitro and in vivo. CMCS powder owned hemostatic capability prior to CS. CMCS powder with SD 63% (neither too high, nor too low) possessed excellent hemostasis both in vitro and in vivo.

scholarly journals Influence of Cross-Linker Polarity on Selectivity towards Lysine Side Chains

2020 ◽  
Author(s):  
Jan Fiala ◽  
Zdeněk Kukačka ◽  
Petr Novák
Keyword(s):  
Structural Information ◽  
Protein Complexes ◽  
Water Soluble ◽  
Cross Linking ◽  
Side Chains ◽  
Model Protein ◽  
Cross Linker ◽  
Progressive Technology ◽  
The Impact ◽  
Chemical Cross Linking

The combination of chemical cross-linking and mass spectrometry is currently a progressive technology for deriving structural information of proteins and protein complexes. In addition, chemical cross-linking is a powerful tool for stabilizing macromolecular complexes for single particle cryo-electron microscopy. Broad pallets of cross-linking chemistry, currently available for the majority of cross-linking experiments, still rely on the amine-reactive N-hydroxysuccinimide esters targeting mainly N-termini and lysine side chains. These cross-linkers are divided into two groups: water soluble and water insoluble; and research teams prefer one or another speculating on the benefits of their choice. However, the effect of cross-linker polarity on the outcome of cross-linking reaction has never been studied. Herein, we use both polar (bis(sulfosuccinimidyl) glutarate) and non-polar (disuccinimidyl glutarate) cross-linkers and systematically investigated the impact of cross-linker hydrophobicity on resulting distance constraints, using bovine serum albumin as a model protein.

scholarly journals Localization of Calmodulin and Dynein Light Chain Lc8 in Flagellar Radial Spokes

2001 ◽  
Vol 153 (6) ◽  
pp. 1315-1326 ◽  
Author(s):  
Pinfen Yang ◽  
Dennis R. Diener ◽  
Joel L. Rosenbaum ◽  
Winfield S. Sale
Keyword(s):  
Light Chain ◽  
Crucial Role ◽  
Cross Linking ◽  
Dynein Light Chain ◽  
Flagellar Motility ◽  
Radial Spoke ◽  
Size And Shape ◽  
Radial Spokes ◽  
Chemical Cross Linking

Genetic and in vitro analyses have revealed that radial spokes play a crucial role in regulation of ciliary and flagellar motility, including control of waveform. However, the mechanisms of regulation are not understood. Here, we developed a novel procedure to isolate intact radial spokes as a step toward understanding the mechanism by which these complexes regulate dynein activity. The isolated radial spokes sediment as 20S complexes that are the size and shape of radial spokes. Extracted radial spokes rescue radial spoke structure when reconstituted with isolated axonemes derived from the radial spoke mutant pf14. Isolated radial spokes are composed of the 17 previously defined spoke proteins as well as at least five additional proteins including calmodulin and the ubiquitous dynein light chain LC8. Analyses of flagellar mutants and chemical cross-linking studies demonstrated calmodulin and LC8 form a complex located in the radial spoke stalk. We postulate that calmodulin, located in the radial spoke stalk, plays a role in calcium control of flagellar bending.

scholarly journals Effect of Various Electrolytes on Theophylline Loaded Sodium Alginate Beads Prepared by Ionic Cross Linking Technique

2013 ◽  
Vol 11 (2) ◽  
pp. 181-189
Author(s):  
Sheikh Tasnim Jahan ◽  
Sams Mohammad Anowar Sadat ◽  
Muhammad Rashedul Islam ◽  
ATM Zafrul Azam ◽  
Jakir Ahmed Chowdhury
Keyword(s):  
Drug Release ◽  
Sodium Alginate ◽  
Electrolyte Concentration ◽  
Research Work ◽  
Three Dimensional ◽  
Alginate Beads ◽  
Water Soluble ◽  
Cross Linking ◽  
Bonding Structure

The purpose of t h e present research work was to prepare alginate beads containing water soluble drug theophylline using ionic cross linking technique, with electrolyte type and concentration as variables. In this study, the beads were characterized and evaluated in respect of their surface morphology, swelling index and in vitro kinetics. The comparative study among the three polyvalent cationic cross linking agents CaCl2 , BaCl2 and Al2 (SO4)3 were investigated based on their cationic charges. Divalent cation, Ca2+ and Ba2+ containing beads showed simultaneous decrease in drug release with increasing electrolyte amount. In case of Al3+ -alginate beads, the delay in release was due to the ability of Al3+ to form three dimensional bonding structure with the sodium alginate inside the beads. As a result, swelling of beads is delayed leading to slow disintegration. Scanning electron microscope (SEM) photomicrographs revealed that with the increase in the electrolyte concentration the density of the cross link is also increased. When the electrolyte concentration is 5 % then the beads surface is rough and rod shape drug is visible. But when the electrolyte concentration is increased from 10 % to 15 % the surface is comparatively smoother and both the swelling property and in vitro drug release are decreased. Most of the formulations followed Higuchi drug release model. DOI: http://dx.doi.org/10.3329/dujps.v11i2.14578 Dhaka Univ. J. Pharm. Sci. 11(2): 181-189, 2012 (December)

Synthesis of N-Carboxymethyl Chitosan Beads for Controlled Drug Delivery Applications

2006 ◽  
Vol 514-516 ◽  
pp. 1015-1019 ◽  
Author(s):  
Rangasamy Jayakumar ◽  
Rui L. Reis ◽  
João F. Mano
Keyword(s):  
Drug Delivery ◽  
Drug Release ◽  
Phosphate Buffer Solution ◽  
Controlled Drug Delivery ◽  
Gastric Fluid ◽  
Carboxymethyl Chitosan ◽  
Water Soluble ◽  
Buffer Solution ◽  
Chitosan Beads

N-Carboxymethyl chitosan (NCMC) is a water soluble derivative of chitosan. The NCMC beads were prepared by using ionotropic gelation process with the counter polyanion tripolyphoshate at pH 4.0 and characterized by scanning electron microscopy. The swelling behavior of the beads at different time intervals was monitored at different pH conditions. The in vitro drug release behavior in various pH solutions was studied using indomethacin as a model drug with two different concentrations (0.3 and 0.6% w/w). The release percent of indomethacin from NCMC beads was found to increase with increasing of pH in phosphate buffer solution medium due to the ionization of carboxymethyl group and high solubility of indomethacin in alkaline medium. These results indicated that the NCMC beads are useful for controlled drug delivery systems through oral administration by avoiding the drug release in the highly acidic gastric fluid region of the stomach.

scholarly journals P1 ParB Domain Structure Includes Two Independent Multimerization Domains

1999 ◽  
Vol 181 (19) ◽  
pp. 5898-5908 ◽  
Author(s):  
Jennifer A. Surtees ◽  
Barbara E. Funnell
Keyword(s):  
Amino Acids ◽  
Domain Structure ◽  
Cross Linking ◽  
Yeast Two Hybrid ◽  
Yeast Two Hybrid System ◽  
C Terminus ◽  
Self Association ◽  
Two Hybrid ◽  
Chemical Cross Linking

ABSTRACT ParB is one of two P1-encoded proteins that are required for active partition of the P1 prophage in Escherichia coli. To probe the native domain structure of ParB, we performed limited proteolytic digestions of full-length ParB, as well as of several N-terminal and C-terminal deletion fragments of ParB. The C-terminal 140 amino acids of ParB form a very trypsin-resistant domain. In contrast, the N terminus is more susceptible to proteolysis, suggesting that it forms a less stably folded domain or domains. Because native ParB is a dimer in solution, we analyzed the ability of ParB fragments to dimerize, using both the yeast two-hybrid system and in vitro chemical cross-linking of purified proteins. These studies revealed that the C-terminal 59 amino acids of ParB, a region within the protease-resistant domain, are sufficient for dimerization. Cross-linking and yeast two-hybrid experiments also revealed the presence of a second self-association domain within the N-terminal half of ParB. The cross-linking data also suggest that the C terminus is inhibitory to multimerization through the N-terminal domain in vitro. We propose that the two multimerization domains play distinct roles in partition complex formation.

Procoagulant Assessment of Topical Battlefield Hemostats Using a Novel Semi-Automated Whole Blood Clotting Assay System.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3161-3161 ◽  
Author(s):  
Keith A. Moskowitz
Keyword(s):  
High Pressure ◽  
Whole Blood ◽  
Blood Clotting ◽  
Complete Blood Count ◽  
Clotting Time ◽  
Contact Phase ◽  
Hemostatic Agents ◽  
Platelet Counts ◽  
Arterial Bleeding

A new granular smectite hemostatic agent (WoundStat™ [WS]) is capable of producing hemostasis during high pressure arterial bleeding (Ward et al., J Trauma, 63:276–284, 2007). The mechanism by which WS and other topical hemostatic agents help achieve clotting has not been investigated. In this study, the procoagulant and hemo-compatibility functions of WS were assessed using a novel whole blood clotting time assay, and its performance compared to current hemostatic agents QuikClot® zeolite granules (QC); CELOX™ marine biopolymer hemostatic granules (CX); sterile gauze bandage (SG); and to glass beads (GB). A modified whole blood recalcified clotting time assay was developed using the Actalyke® Mini II activated clotting time (ACT) analyzer and empty G-ACT tubes devoid of contact phase activating reagents. Whole citrated blood recalcified to 10 mM with CaCl2 yielded clot times (CT) of 217+/−6 sec, 212+/−11 sec, and 236+/−5 sec, respectively, for three normal donors. Addition of WS resulted in a time and dose-dependent biphasic decrease in CT. At 0.5 mg/ml, WS lowered CT by over 40% to 124+/−7 sec, 133+/− 9 sec, and 126+/−5 sec, p <0.001. Equal concentrations of QC also diminished CT ∼ 30% to 146+/−15 sec, 139+/−18 sec, and 184+/−22 sec, p< 0.02. Differences in CT between WS and QC were not significant. Surprisingly, 0.5 mg/ml CX did not exhibit procoagulant activity, as CT were on average 8% higher than controls (234+/−4 sec, 227+/−8 sec and 256+/−5 sec). SG and GB gave results within 5% of untreated samples. WS at ≥1.5 mg/ml clotted all samples in <90 sec, (the ACT instrument detection limit) and also significantly diminished CT of heparinized blood from 790 to 135 sec. All agents were hemocompatible as judged by the lack of change in hematocrit and hemoglobin complete blood count values after 45 min incubation. However, citrated blood incubated for <45 min with QC consistently demonstrated a drop in baseline platelet counts by >50,000 /μl while platelet counts in WS-treated specimens were similar to untreated controls (214,000/μl versus 219,000/μl) and clumped granules, presumably platelet-mediated, were visually apparent in the QC-treated bloods. Blood incubated with CX for up to 45 min also exhibited a drop in platelets from 204,000/μl to 150,000/μl. When QC, CX and WS were incubated in blood for 3 hours, corresponding to the maximal times recommended during field application prior to reapplication or surgical debridement, all hemostats resulted in an ∼ 50% reduction in platelet counts from 258,000/μl to 132,000/μl; 158,000/μl and 155,000 /μl, respectively. In summary, the data suggest that WS is hemocompatible, is moderately capable of inducing in vitro platelet aggregates, and may assist in the arrest of high-pressure arterial bleeding, at least in part by promoting the intrinsic contact phase of blood coagulation. Additional studies are required to further elucidate the potential platelet- and plasma-mediated mechanisms by which WS may help achieve hemostasis during severe arterial traumatic injury.

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