Protective effect of Carbopol on enzymatic degradation of a peptide-like substrate. I: Effect of various concentrations and grades of Carbopol and other reaction variables on trypsin activity.

The purpose of this study was to determine and compare the effect of various concentrations and grades of Carbopol on trypsin-induced degradation of a prototype substrate, N(alpha)-benzoyl-L-arginine ethyl ester hydrochloride (BAEE). Effect of other reaction variables, such as viscosity and ionic strength of the medium on the trypsin activity, was also analyzed simultaneously. Four concentrations and three commercially available grades of Carbopol were used. The effect of Carbopol was expressed in terms of change in the velocity of degradation reaction. A modified trypsin assay was developed and used for analysis. Up to a concentration of 0.35% w/v, Carbopol 934P showed a concentration-dependent increase in its ability to reduce the rate of enzymatic hydrolysis of BAEE. Similar inhibitory effect was observed with all three grades of Carbopol. The activity of trypsin was unaffected by other reaction variables, suggesting that interaction between the protein and the polymer could be the mechanism responsible for reduced trypsin activity. This study suggests that Carbopol can be a useful excipient for oral delivery of bioactive proteins and peptides, due to its ability to reduce the enzyme-induced degradation of these agents.

Thermodynamic analysis of compact formation; compaction, unloading, and ejection. II. Mechanical energy (work) and thermal energy (heat) determinations of compact unloading and ejection.

A compaction calorimeter, previously described (DeCrosta, M.T., Schwartz, J.B., Wigent, J.B., Marshall, K., 2000. Thermodynamic analysis of compact formation; compaction, unloading, and ejection. I. Design and development of a compaction calorimeter and mechanical and thermal energy determinations of powder compaction. Int. J. Pharm. 198, 113--134), was utilized to evaluate the thermodynamics of the unloading and ejection of compacts of Avicel pH102, Emcompress, Fast-Flo #316, Starch 1500, and acetaminophen (APAP). A constant strain waveform, applied by a compaction simulator, enabled the separate thermodynamic evaluation of unloading from compaction. The brittle materials, Fast-Flo #316 and Emcompress, displayed the most unloading work, and the plastic/self-lubricating materials, Avicel and Starch 1500, displayed the least. Unloading heat values were negative for all materials, except APAP. APAP's positive heat values indicated the breaking of bonds during unloading as a result of its highly elastic nature. Positive internal energy changes of unloading, which indicate the net breaking of bonds, were observed for APAP and Emcompress over the compaction forces tested. Negative energy changes for Starch 1500, Fast-Flo #316, and Avicel became positive with increasing compaction forces. Ejection work increased with increasing compaction force for the brittle materials, whereas smaller ejection work values for Avicel, Starch 1500, and APAP remained constant. Increasing negative heat values as a function of compaction force were observed for Fast-Flo #316 and Emcompress. Negative internal energy values for ejection were observed for Fast-Flo #316 and Emcompress, which indicates net bond formation as a result of high shear of the compact with the die wall. Internal energy changes for Starch 1500, Avicel, and APAP, were approximately zero, indicating the absence of net bonding or bond formation during the process.

Characterization and in vitro release of methotrexate from gelatin/methotrexate conjugates formed using different preparation variables.

The purpose of this study was to evaluate effects of preparation variables on the composition of gelatin-methotrexate conjugates, and to evaluate their in vitro stability. Conjugation variables of pH, amount of conjugating agent 1-ethyl-3-(diaminopropyl)carbodiimide HCl (EDC), and methotrexate (MTX), with unfractionated gelatin were examined. Conjugate composition was determined spectrophotometrically. The molar ratios of MTX to gelatin in the conjugates ranged from 5.9 to 64.9. Molar ratios increased with molecular weight (MW) of gelatin in the conjugate, but the weight ratio was constant. This common conjugating procedure, however, produces by-product crosslinking and produces a mix of covalent MTX binding to carboxyl and amino groups of the gelatin. For release studies, gelatin was fractionated by size exclusion spectra (SEC) into MW of 21, 91, and 195 kDa prior to conjugation. MTX release from conjugates in dialysis cassettes at 25, 37, and 50 degrees C, in isotonic pH 7.4, buffer over 72 h was assayed by high performance liquid chromatography (HPLC). There was no effect of gelatin MW on MTX release. MTX release was approximately linear and attained 2.3, 7.2, and 13% by 72 h at 25, 37, and 50 degrees C, respectively, for the 91 kDa conjugates. First-order release rate constants were 0.23 x 10(-3), 0.95 x 10(-3), and 1.8 x 10(-3) x h(-1), respectively. The calculated activation energy for MTX release was 15.8 kcal/mol. Rate constants and the activation energy for MTX release are consistent with hydrolysis of a peptide bond. Non-degraded MTX species were found in the release medium at amounts similar to free MTX and were attributed to MTX polymers and MTX/gelatin fragments < 10 kDa.

Drug release from film-coated chlorpheniramine maleate nonpareil beads: effect of water-soluble polymer, coating level, and soluble core material.

The purpose of this research was to use a new drug release model to study the effects of formulation parameters on drug release from a film-coated chlorpheniramine (CPM) nonpareil system. The film-coated CPM nonpareils were prepared by using a fluid bed apparatus. A hydroxylpropylmethylcellulose (HPMC) solution was blended with an aqueous ethylcellulose dispersion (Surelease) to adjust the permeability of the film. The apparent permeability of samples was obtained from dissolution data using a previously reported drug release equation. The apparent permeability was plotted versus the film coating level or the HPMC concentration in the film. When the natural logarithm of the apparent permeability versus coating level was graphed, a biphasic plot was observed in the group without HPMC in the film, showing the occurrence of a critical coating level. It was suggested that a mechanically formed porous film (due to an incomplete coating) could change to a nonporous film after the bead was completely coated. However, in the group that contained 12% HPMC in the film, the critical coating level was not observed. A porous film, formed by the leaching out of the water-soluble polymer, would not change to a nonporous film even after the bead is completely coated. Through a mathematical derivation, the decrease of apparent permeability versus coating level was related to the reduction of the total hole area. The apparent permeability was found to increase with the HPMC concentration. After a critical concentration was reached, the further addition of HPMC into the film caused a rapid increase in apparent permeability. The critical HPMC concentration was related to a minimum domain formation concentration (MDFC). A rapid increase of the drug release was observed when the dissolution profile of a sample made from a regular sugar nonpareil core (soluble) was compared with the sample made from a precoated nonpareil core (insoluble), which suggests that the drug release can be enhanced by the dissolution of the core. A minimum concentration of the HPMC was required to effectively modify permeability of the film. The critical coating level and critical concentration of HPMC can be determined from the apparent permeability plot using a previously published equation. The dissolution of a soluble core can greatly enhance the release of the drug from the nonpareil system.

Thermodynamic analysis of compact formation; compaction, unloading, and ejection. I. Design and development of a compaction calorimeter and mechanical and thermal energy determinations of powder compaction.

The aim of this investigation was to determine and evaluate the thermodynamic properties, i.e. heat, work, and internal energy change, of the compaction process by developing a 'Compaction Calorimeter'. Compaction of common excipients and acetaminophen was performed by a double-ended, constant-strain tableting waveform utilizing an instrumented 'Compaction Simulator.' A constant-strain waveform provides a specific quantity of applied compaction work. A calorimeter, built around the dies, used a metal oxide thermistor to measure the temperature of the system. A resolution of 0.0001 degrees C with a sampling time of 5 s was used to monitor the temperature. An aluminum die within a plastic insulating die, in conjunction with fiberglass punches, comprised the calorimeter. Mechanical (work) and thermal (heat) calibrations of the elastic punch deformation were performed. An energy correction method was outlined to account for system heat effects and mechanical work of the punches. Compaction simulator transducers measured upper and lower punch forces and displacements. Measurements of the effective heat capacity of the samples were performed utilizing an electrical resistance heater. Specific heat capacities of the samples were determined by differential scanning calorimetry. The calibration techniques were utilized to determine heat, work, and the change in internal energies of powder compaction. Future publications will address the thermodynamic evaluation of the tablet sub-processes of unloading and ejection.

The effect of humidity on samples of microcrystalline cellulose taken from the extrusion/marumerization process.

The purpose of this work was to examine the sorption and desorption of water by various samples of microcrystalline cellulose, MCC (Avicel PH-101), taken from the extrusion/marumerization process, and to provide data that may explain how water affects the MCC polymer matrix during the formation of beads. Two isopiestic (humidity) studies were conducted: the first used samples exposed directly to controlled humidity conditions, whereas the second used samples that were freeze-dried before being exposed to controlled humidity conditions. Water sorption and desorption were determined gravimetrically. When both sets of samples were initially exposed to low-humidity conditions, they reached equilibrium by desorbing water. When these samples were initially exposed to high-humidity conditions, the high moisture content samples desorbed water, whereas the low moisture content and the freeze-dried samples sorbed water to reach equilibrium. When the first set of samples was initially exposed to high- and then to low-humidity conditions, they reached the same water content achieved by being equilibrated directly at the low-humidity condition. However, samples that were initially exposed to low- and then to high-humidity conditions had equilibrium water contents that were lower than those achieved by being equilibrated directly at the high-humidity condition. The original MCC systems exhibit a hysteretic effect above 85%, whereas the freeze-dried systems have a broader range hysteretic effect starting at 20% relative humidity. The results suggest that the internal structure of the MCC polymer fibers must change with the sorption and desorption of water, supporting the autohesion theory.

Drug release from film-coated chlorpheniramine maleate nonpareil beads: water influx and development of a new drug release model.

The purpose of this work was to investigate drug release from film-coated chlorpheniramine maleate (CPM) nonpareils (sugar spheres) and the effect of water influx on the drug release mechanism. The methods used in the study involved the layering of CPM onto nonpareil cores using a fluid-bed apparatus. These CPM cores were then coated with an aqueous ethylcellulose dispersion, which was blended with a solution of hydroxylpropylmethylcellulose (HPMC) at different concentrations. The net water influx was determined by measuring water uptake during dissolution. The film surface area was calculated from bead diameters measured with an optical microscope. Drug release profiles were measured using USP dissolution method I (basket). The results showed that significant water influx occurred, which produced an internal liquid phase ranging from 0 to 1.8 x 10(3) mm3/g of sample. As a result of the water uptake, an increase in bead size was observed. The bead surface area varied over the range of 40-80 x 10(3) mm2/g sample because of a combined effect of the water uptake and the release of the bead contents. A bead geometry parameter was proposed as the ratio of the bead surface area to the volume of the internal liquid phase. This bead geometry parameter was measured as a function of time and fit to an equation using a computer curve-fitting technique. This equation was substituted into an existing drug release model to give a more appropriate mathematical model describing drug release from this system. The conclusion drawn from these results is that the influx of water during drug dissolution creates a progressive increase in the liquid phase within the nonpareil bead; this causes a corresponding increase in the bead surface area which influences the drug release rate.

Differences in the mechanical strength of dried microcrystalline cellulose pellets are not due to significant changes in the degree of hydrogen bonding.

The mechanical strengths of oven-dried pellets of microcrystalline cellulose, MCC, prepared by extruder/marumerizer technology are weaker when ethanol/water mixtures are used as granulating solutions than when granulated with water. Previously, the difference in the strengths of these pellet systems were thought to be due to changes in the degree of hydrogen bonding within these systems. This work reports the results of studies using magic-angle-spin nuclear magnetic resonance (NMR), x-ray diffraction and degree of crystallinity, and oxygen combustion calorimetry studies of various MCC systems developed to test this hypothesis. Carbon-13 cross polarization magic-angle spin NMR studies of MCC pellets with and without 10% theophylline showed no differences in the spectra of these systems. X-ray diffraction studies and heats of combustion data obtained from oxygen combustion calorimetry of oven-dried pellets of MCC granulated with either water, deuterated water, or a 70/30 ethanol/water mixture were found to have no significant differences in their diffraction patterns, degree of crystallinity, or internal energies. None of these results provide any evidence that a significant net change in the degree of hydrogen bonding is responsible for the observed changes in the strengths of these systems. It is hypothesized that the strength of these dried pellets may, in part, be due to the conversion of some of the intramolecular hydrogen bonded amorphous fibrils at the surface of the MCC particles to intermolecular hydrogen bonded fibrils with other MCC particles.

Charge-transfer complexes of iodine and nonionic surfactants: interpretation and use in the Winkler method.

Formation constants (Kc) and molar absorption coefficients (epsilon c) of complexes of iodine and various nonionic surfactants were determined, providing a basis for selection of a surfactant for use in a spectrophotometric modification of the Winkler method. The method of calculation of Kc and epsilon c was extended to include absorption by triiodide at the wavelength of maximum absorbance of the complex. Because the molar absorption coefficients of polyoxyethylene 10 oleyl ether (oleth 10) and polyoxyethylene 23 oleyl ether were significantly greater than those of other surfactants, they are superior candidates for use in the Winkler method. Formation constants could not be correlated with molecular characteristics of the surfactants such as alkyl chain or polyoxyethylene chain length, nor with physical characteristics of iodine-surfactant solutions such as reduction of iodine loss due to volatilization.