Boronic acid functionalized fibrous cellulose for the selective enrichment of glycopeptides.

Enrichment of glycoproteins has been important because of their dynamicity and role in biological systems. Study of glycoproteins is complex because of the simultaneous glycosylation and deglycosylation inside the body. Often employed affinities for glycopeptides are hydrazide, boronic acid, or physiosorbed lectin on support materials. Cellulose, a natural polysaccharide, has rich surface chemistry, stable structure, low cost and availability in different variants. In present study, fibrous cellulose is oxidized using periodate to modify with boronic acid. Attachment of boronic acid is confirmed by Fourier transform infrared spectroscopy. Particle size and morphology of boronic acid@fibrous cellulose is studied by scanning electron microscopy. The enrichment efficiency is evaluated by using horseradish peroxidase as model protein. Boronic acid@fibrous cellulose is selective up to 1:250 for spiked horseradish peroxidase in bovine serum albumin digest, sensitive down to 0.1 femtomol and recovering 88.15% glycopeptides. Moreover, protein binding capacity is determined as 213 mg/g and 41% sequence coverage of horseradish peroxidase protein with all eight glycosylation sites detected. Total of 18 glycopeptides are enriched from immunoglobulin digest showing ability of boronic acid@fibrous cellulose to enrich glycoproteins from multiglycoforms. Enrichment from human serum recovers 18% extracellular and 72% secreted glycoproteins via bottom-up approach and online tools.

Preparation and properties of cellulose nanocrystals, gelatin, hyaluronic acid composite hydrogel as wound dressing.

Gelatin (GA), hyaluronic acid (HA) and cellulose nanocrystals (CNC) are promising materials for skin wound care. In this study the GA-HA-CNC hydrogels were prepared by cross-linking and freeze-drying. The composition and mechanism of GA-HA-CNC hydrogels were confirmed by FTIR. The morphology and pore size were obtained by SEM. We accessed the physical property from rheological results and swelling ratio. NIH-3T3 cells were inoculated into the hydrogels and cultured for different days, then we analyzed the cytotoxicity of the prepared hydrogels by CCK-8 methods and live/dead pictorial diagram using staining kits. FTIR revealed the combination between GA, HA and CNC was attributed to the amide bond and hydrogen bonding. SEM results showed that the drying GA-HA-CNC hydrogels were spongy, with the pore diameter about 80-120 µm. CNC significantly enhanced the property of the hydrogels and play a vital role according to the rheology and swelling results. The cells culture results showed that NIH-3T3 cells can attached to, grow, and proliferate well on the GA-HA-CNC hydrogels. In conclusion, the natural GA-HA-CNC hydrogel has great potential for the skin wound repair.

Nasal drug absorption from powder formulations: The effect of three types of hydroxypropyl cellulose (HPC).

Despite the numerous advantages of powder formulations, few studies have described their nasal drug absorption. The first aim of this study was to compare the drug absorption from powder formulation with that from a liquid formulation in rats. Since pharmaceutical excipients are usually added to most powder formulations, the second aim of the study was to investigate the effect of hydroxypropyl cellulose (HPC) on nasal drug absorption from the powder. Three types of HPC with different polymerization degrees were used: HPC(SL), HPC(M), and HPC(H). The model drugs were warfarin (BCS Class I), piroxicam (BCS Class II), and sumatriptan (BCS Class III). The absorption of these model drugs in the powder form was higher than that from the solution. All HPCs failed to enhance warfarin absorption, while the piroxicam absorption was enhanced only by HPC(M). Sumatriptan absorption was not enhanced by HPC(SL), but by HPC(M) and HPC(H). The differences in nasal absorption of the three model drugs promoted by HPCs depend on the permeability and solubility of the drug. Moreover, the nasal retention of different formulations was increased by HPCs. Because HPCs showed no toxic effect on the nasal epithelium. These findings indicate that powder formulations supplemented with HPC are a valuable and promising approach to increase the nasal absorption of highly soluble and poorly permeable drugs.

On the non-linear attachment characteristics of blood to bacterial cellulose/kaolin biomaterials.

In this communication, we report a non-linear variation in the strength of blood attachment to bacterial cellulose/kaolin biomaterials as the fractions of bacterial cellulose to kaolin are increased. The changes observed for attachment strength are elucidated following both experimental and numerical investigations on both the biomaterial and the blood-biomaterial interface. Our research reveals that the non-linear strength of attachment of blood is related to topographical characteristics on the surface of the biomaterial, the maleability of the biomaterial and the intermolecular strength of attraction between clotted blood proteins (fibrinogen) with the cellulose/kaolin components of the biomaterial.

Assessment of the within- and between-lot variability of Whatman™ FTA(®) DMPK and 903(®) DBS papers and their suitability for the quantitative bioanalysis of small molecules.

BACKGROUND: To ensure that PK data generated from DBS samples are of the highest quality, it is important that the paper substrate is uniform and does not unduly contribute to variability. This study investigated any within and between lot variations for four cellulose paper types: Whatman™ FTA(®) DMPK-A, -B and -C, and 903(®) (GE Healthcare, Buckinghamshire, UK). The substrates were tested to demonstrate manufacturing reproducibility (thickness, weight, chemical coating concentration) and its effect on the size of the DBS produced, and the quantitative data derived from the bioanalysis of human DBS samples containing six compounds of varying physicochemical properties. RESULTS & DISCUSSION: Within and between lot variations in paper thickness, mass and chemical coating concentration were within acceptable manufacturing limits. No variation in the spot size or bioanalytical data was observed. CONCLUSION: Bioanalytical results obtained for DBS samples containing a number of analytes spanning a range of chemical space are not affected by the lot used or by the location within a lot.

First-in-human phase 1/2a trial of CRLX101, a cyclodextrin-containing polymer-camptothecin nanopharmaceutical in patients with advanced solid tumor malignancies.

Patients with advanced solid malignancies were enrolled to an open-label, single-arm, dose-escalation study, in which CRLX101 was administered intravenously over 60 min among two dosing schedules, initially weekly at 6, 12, and 18 mg/m(2) and later bi-weekly at 12, 15, and 18 mg/m(2). The maximum tolerated dose (MTD) was determined at 15 mg/m(2) bi-weekly, and an expansion phase 2a study was completed. Patient samples were obtained for pharmacokinetic (PK) and pharmacodynamic (PD) assessments. Response was evaluated per RECIST criteria v1.0 every 8 weeks. Sixty-two patients (31 male; median age 63 years, range 39-79) received treatment. Bi-weekly dosing was generally well tolerated with myelosuppression being the dose-limiting toxicity. Among all phase 1/2a patients receiving the MTD (n = 44), most common grade 3/4 adverse events were neutropenia and fatigue. Evidence of systemic plasma exposure to both the polymer-conjugated and unconjugated CPT was observed in all treated patients. Mean elimination unconjugated CPT Tmax values ranged from 17.7 to 24.5 h, and maximum plasma concentrations and areas under the curve were generally proportional to dose for both polymer-conjugated and unconjugated CPT. Best overall response was stable disease in 28 patients (64 %) treated at the MTD and 16 (73 %) of a subset of NSCLC patients. Median progression-free survival (PFS) for patients treated at the MTD was 3.7 months and for the subset of NSCLC patients was 4.4 months. These combined phase 1/2a data demonstrate encouraging safety, pharmacokinetic, and efficacy results. Multinational phase 2 clinical development of CRLX101 across multiple tumor types is ongoing.

Improved bioavailability of darunavir by use of kappa-carrageenan versus microcrystalline cellulose as pelletisation aid.

The aim of this study was to produce pellet formulations containing a high drug load (80%) of the poorly soluble HIV-protease inhibitor darunavir, using wet extrusion/spheronization with kappa-carrageenan or microcrystalline cellulose (MCC) as pelletization aid. Drug release was assessed in vitro by a standardized paddle-dissolution test and in vivo by a single-dose pharmacokinetic study in dogs. Mean dissolution time (MDT) was 78.2+/-3.5 h from MCC pellets (1301+/-301 microm) and 6.1+/-0.7 min from kappa-carrageenan pellets (966+/-136 microm). In contrast to kappa-carrageenan pellets, MCC pellets did not disintegrate and showed a diffusion-controlled drug release. In line with the in vitro findings, the darunavir peak plasma levels and exposure after the administration of a 300 mg dose were more than 60-fold higher when formulated with kappa-carrageenan pellets when compared with MCC pellets, and 10-fold higher after co-administration with 10mg/kg of ritonavir. The relative bioavailability of darunavir versus the reference tablet (F(rel)) was 155% with kappa-carrageenan pellets and 2% with MCC pellets without ritonavir, while 78% and 9%, respectively, in presence of ritonavir. In conclusion, when compared with MCC pellets, the bioavailability of darunavir was substantially improved in kappa-carrageenan pellets, likely due to their better disintegration behavior.

MAP20, a microtubule-associated protein in the secondary cell walls of hybrid aspen, is a target of the cellulose synthesis inhibitor 2,6-dichlorobenzonitrile.

We have identified a gene, denoted PttMAP20, which is strongly up-regulated during secondary cell wall synthesis and tightly coregulated with the secondary wall-associated CESA genes in hybrid aspen (Populus tremula x tremuloides). Immunolocalization studies with affinity-purified antibodies specific for PttMAP20 revealed that the protein is found in all cell types in developing xylem and that it is most abundant in cells forming secondary cell walls. This PttMAP20 protein sequence contains a highly conserved TPX2 domain first identified in a microtubule-associated protein (MAP) in Xenopus laevis. Overexpression of PttMAP20 in Arabidopsis (Arabidopsis thaliana) leads to helical twisting of epidermal cells, frequently associated with MAPs. In addition, a PttMAP20-yellow fluorescent protein fusion protein expressed in tobacco (Nicotiana tabacum) leaves localizes to microtubules in leaf epidermal pavement cells. Recombinant PttMAP20 expressed in Escherichia coli also binds specifically to in vitro-assembled, taxol-stabilized bovine microtubules. Finally, the herbicide 2,6-dichlorobenzonitrile, which inhibits cellulose synthesis in plants, was found to bind specifically to PttMAP20. Together with the known function of cortical microtubules in orienting cellulose microfibrils, these observations suggest that PttMAP20 has a role in cellulose biosynthesis.

Analysis of the release process of phenylpropanolamine hydrochloride from ethylcellulose matrix granules IV.(1)) Evaluation of the controlled release properties for in vivo and in vitro release systems.

In the pharmaceutical preparation of a controlled release drug, it is very important and necessary to understand the release properties. The dissolution test is a very important and useful method for understanding and predicting drug-release properties. It was readily confirmed in the previous paper that the release process could be assessed quantitatively by a combination of the square-root time law and cube-root law equations for ethylcellulose (EC) matrix granules of phenylpropanolamine hydrochloride (PPA). In this paper EC layered granules were used in addition to EC matrix. The relationship between release property and the concentration of PPA in plasma after administration using beagle dogs were examined. Then it was confirmed that the correlativity for EC layered granules and EC matrix were similar each other. Therefore, it was considered that the dissolution test is useful for prediction of changes in concentration of PPA in the blood with time. And it was suggested that EC layered granules were suitable as a controlled release system as well as EC matrix.

The fermentation of different dietary fibers is associated with fecal clostridia levels in men.

Only a few reports have compared the fermentation of pectin and cellulose using the hydrogen-breath test, and no studies have examined the relation between the hydrogen breathing pattern and colonic microflora. Using breath-hydrogen measurements, we investigated whether different dietary fibers (DFs) were fermented differently and whether there were individual differences after ingestion of the same DF; we also examined the relation between individual fecal microflora and the fermentation of DF. Results of hydrogen tests in 14 men were compared after they had ingested 20 g of pectin, 20 g of cellulose, or 6 g of lactulose (a DF-like substance). We examined the relation between the breath hydrogen results and the subjects' fecal microflora. We defined significant fermentation (i.e., positive cases) as a continuous rise in hydrogen in the expiratory air of >19 ppm. The subjects were divided into 3 groups according to their hydrogen breath test pattern, i.e., positive for lactulose and pectin (Group LP, n = 4); positive for lactulose alone (Group L, n = 7); and negative for pectin, cellulose, and lactulose (Group N, n = 3). Individual differences were noted in subjects from Group LP and Group L. The detection frequency of lecithinase-negative clostridia was higher in Group LP than in the other groups (P < 0.05), and the detection frequency and the number of lecithinase-positive clostridia were higher in Groups LP and L than in Group N (P < 0.05). These findings suggest that the Clostridium species are associated with hydrogen production. The hydrogen breath test results of DFs depend on both the type of DF and the individual colonic microflora. The amount and constitution of colonic microflora might be predicted by the hydrogen-breath test using different DFs.

Influence of polysulfone and hemophan hemodialysis membranes on phagocytes.

The aim of the study was to compare the effect of hemophane and polysulfone membranes on the phagocyte-derived production of reactive oxygen species (ROS) as well as on neutrophil CD11b and CD62L expression in patients undergoing regular hemodialysis. The effects of hemodialysis membranes were also studied in in vitro conditions after coincubating them with differentiated HL-60 cells. ROS production was measured using chemiluminometric and flow cytometric methods. Expression of CD11b, CD62L and mitochondrial membrane potential were detected by monoclonal antibodies and by the JC-1 fluorescent probe, respectively. Depressed ROS production was observed in patients already before dialysis. Further decrease in ROS production and an increase in CD11b expression were observed especially in patients after hemophan hemodialysis. Decreased ROS production and increased CD11b expression were observed also after incubation of HL-60 cells with hemophan membranes. Mitochondrial membrane potential dropped only after incubating cells with hemophan membranes proving its more serious adverse effects in comparison with the polysulfone membrane. In conclusion, deleterious effects of hemodialysis on the metabolic activity of phagocytes were proved. Combining chemiluminescent and flow cytometric methods for the detection of ROS production and determining mitochondrial membrane potential can be useful tools for the analysis of material biocompatibility.

Serum amyloid A and P protein levels are lowered by dextran sulfate cellulose low-density lipoprotein apheresis.

The present study describes the short-term effect of dextran sulfate cellulose (DSC) low-density lipoprotein (LDL) apheresis using a plasma separator equipped with a polysulfone (PS) membrane filter (PS/DSC-LDL apheresis) on the serum amyloid A (SAA) and P (SAP) protein levels during treatment in a patient with familial hypercholesterolemia (type IIa, heterozygote). PS/DSC-LDL apheresis markedly lowered both the SAA (reduction percentage, 84.1+/-8.2%) and SAP (91.4+/-5%) levels, which returned to their respective initial levels within 4 days. Experimentally, the levels of both proteins also decreased on passage through the DSC minicolumn without a PS membrane, indicating that the DSC resin had an affinity to both proteins. These results suggest that PS/DSC-LDL apheresis may be advantageous for amyloid protein accumulating disorders, including amyloidosis and atherosclerosis.

Modulation of CD11b/CD18 on monocytes and granulocytes following hemodialysis membrane interaction in vitro.

We studied the generation of CD11b/CD18 mobilizing factors in serum after incubation with dialysis membrane fragments of different chemical composition. We also evaluated the relative importance of the alternative and classical pathways of the complement system in the generation of such factors. Monocytes and granulocytes from healthy blood donors were incubated in normal human serum (NHS) and in NHS that had been preincubated with Cuprophan (CU) membrane (NHS-CU), Hemophan (HE) (NHS-HE) or polysulfone (PS) (NHS-PS). NHS-CU caused the highest up-regulation of the CD11b/CD18 receptor on monocytes and granulocytes. The rank in capacity to mobilize CD11b/CD18 on granulocytes was CU > HE > PS (p < 0.001), CU > HE (p < 0.05) and HE > PS (p < 0.001). The rank in capacity to mobilize CD11b/CD18 on monocytes was CU > HE > PS (p < 0.001), CU > HE (p < 0.05) and HE > PS (p < 0.01). NHS-PS induced a lower up-regulation of CD11b/CD18 compared to NHS which indicates that serum factors with the ability to mobilize the CD11b/CD18 receptor on monocytes and granulocytes are deposited on or absorbed by PS. In order to study the relative contribution of the alternative and classical pathways of the complement system in the generation of CD11b/CD18 mobilizing factors in serum, three different serum preparations (1. both pathways intact. 2. only the alternative intact and 3. only the classical pathway intact) were used. The CU membrane activated the classical pathway to a larger extent than the PS membrane (p < 0.01). When only the alternative pathway was intact no difference in the generation of CD11b/CD18 mobilizing factors between the CU and PS membranes was observed. These studies show that CD11b/CD18 mobilizing serum factors are generated after incubation with CU membranes and that such factors are probably adsorbed by PS. The classical pathway of complement activation seems to contribute to the generation of CD11b/CD18 mobilizing factors in serum.

Improvement of blood compatibility on cellulose dialysis membrane. III. Synthesis and performance of water-soluble cellulose grafted with phospholipid polymer as coating material on cellulose dialysis membrane.

To improve the surface blood compatibility on a cellulose hemodialysis membrane, a blood compatible polymer with a phospholipid polar group, poly[2-methacryloyloxyethyl phosphorylcholine(MPC)], was immobilized on the surface through the coating of a water-soluble cellulose grafted with poly(MPC) (MPC-grafted cellulose, MGC). The MGC was synthesized by graft copolymerization of MPC on a water-soluble cellulose using cerium ion as an initiator. The coating process on the cellulose membrane with an aqueous solution of the MGC was convenient, and the MGC on the surface was not significantly detached even after immersion in water. The permeability and mechanical strength of the membrane coated with the MGC did not decrease compared with the original membranes. The MGC-coated cellulose membrane was blood compatible, as determined by the prevention of platelet adhesion and aggregation after contact with platelet-rich plasma. From these results, it is concluded that the MGC may be a useful material for improving the blood compatibility of the cellulose hemodialysis membrane.

[Changes in leukocyte counts using a cellulose acetate membrane (FB-T) during hemodialysis. A comparison with a cuprophane membrane].

The effect of cellulose acetate as the membrane of dialyzer on changes in white blood cell (WBC), platelet, CH50, C3 and C4 was evaluated in 5 patients with chronic renal failure during hemodialysis. Similarly the effect of cuprophan on these values was also evaluated. Significant changes in WBC (neutrophil and lymphocyte) were observed in all patients with cuprophan, but changes in WBC in those with cellulose acetate were not significant. There were no differences in the platelet count, CH50, C3 and C4 values between the cupro phan and cellulose acetate membrane.

Effect of cellulose phosphate and dietary calcium restriction in primary hyperparathyroidism.

1. The bivalent cation-binding agent, cellulose phosphate, together with a low calcium diet was given for 6 days to nine patients with primary hyperparathyroidism subsequently verified at surgery. 2. Urinary calcium fell promptly by 8-4 mmol/24 h, and by 70% and reached amounts below 4-0 mmol/24 h in five of the nine patients. The magnitude of fall may have been related to increased synthesis of vitamin D by the skin in a sub-tropical environment. Plasma magnesium fell steadily and urinary magnesium fell by 80%. 3. The plasma calcium showed two types of response. In five patients there was no significant change because a reduction in calcium load was offset by a further increase in the already high tubular reabsorption of calcium. In the remaining four patients, the tubular reabsorption of calcium was at a higher level initially and failed to increase further on the experimental regime, with a corresponding fall in plasma calcium. 4. The hypercalcaemia of primary hyperparathyroidism can be explained by increased renal tubular reabsorption of calcium; net bone resorption makes only a small contribution but an additional factor dependent on the blood-bone equilibrium is not ruled out. 5. Comparison with other published data suggests that the fall in urinary calcium in response to a calcium-depleting regimen is prevented by concurrent depletion of inorganic phosphate and may be enhanced by concurrent depletion of magnesium. 6. Persistence of hypercalcaemia combined with an increase in tubular reabsorption of calcium in response to cellulose phosphate may be of diagnostic value in suspected primary hyperparathyroidism. 7. Cellulose phosphate may be of value in stone prevention in patients with primary hyperparathyroidism who are unsuitable for surgical treatment.