Aluminum elution and precipitation in glass vials: effect of pH and buffer species.

Inorganic extractables from glass vials may cause particle formation in the drug solution. In this study, the ability of eluting Al ion from borosilicate glass vials, and tendencies of precipitation containing Al were investigated using various pHs of phosphate, citrate, acetate and histidine buffer. Through heating, all of the buffers showed that Si and Al were eluted from glass vials in ratios almost the same as the composition of borosilicate glass, and the amounts of Al and Si from various buffer solutions at pH 7 were in the following order: citrate > phosphate > acetate > histidine. In addition, during storage after heating, the Al concentration at certain pHs of phosphate and acetate buffer solution decreased, suggesting the formation of particles containing Al. In citrate buffer, Al did not decrease in spite of the high elution amount. Considering that the solubility profile of aluminum oxide and the Al eluting profile of borosilicate glass were different, it is speculated that Al ion may be forced to leach into the buffer solution according to Si elution on the surface of glass vials. When Al ions were added to the buffer solutions, phosphate, acetate and histidine buffer showed a decrease of Al concentration during storage at a neutral range of pHs, indicating the formation of particles containing Al. In conclusion, it is suggested that phosphate buffer solution has higher possibility of forming particles containing Al than other buffer solutions.

Optimization of primary drying condition for pharmaceutical lyophilization using a novel simulation program with a predictive model for dry layer resistance.

The purpose of this study was to develop a novel simulation program to accurately predict the maximum product temperature and the primary drying time in lyophilization using the predictive model for dry layer resistance, which is the resistance of dried cake against water vapor flow. Ten percent sucrose aqueous solution was selected as a model formulation. It was demonstrated that the deviations between the predicted and measured maximum product temperature were attributed to the error of dry layer resistance at a given drying condition, which was different from the measured dry layer resistance in a preliminary lyophilization run for the simulation program. However, when the predictive model of dry layer resistance was used for the simulation program, the model remarkably enhanced the accuracy of the simulation program to predict the maximum product temperature and primary drying time under various operating conditions. Furthermore, the primary drying condition required for minimized drying at a close collapse temperature was successfully discovered through one preliminary run. Therefore, it is expected that the developed simulation program is useful for designing the lyophilization cycle without a trial and error approach.

Effects of phosphate buffer in parenteral drugs on particle formation from glass vials.

The characteristics of inorganic particles generated in glass vials filled with phosphate buffer solutions were investigated. During storage, particles were visually detected in the phosphate buffer solution in particular glass vials which pass compendial tests of containers for injectable drugs. These particles were considered to be different from ordinal glass delamination, which has been reported in a number of papers because the particles were mainly composed of Al, P and O, but not Si. The formation of the particles accelerated at higher storage temperatures. Among the surface treatments tested for the glass vials, sulfur treatment showed a protective effect on the particle formation in the vials, whereas the SiO(2) coating did not have any protective effects. It was found that the elution ratio of Al and Si in the solution stored in the glass vials after the heating was similar to the ratio of Al and Si in borosilicate glass. However, the Al concentration decreased during storage (5°C, 6 months), and consequently, particle formation was observed in the solution. Adding citrate, which is a chelating agent for Al, effectively suppressed the particle formation in the heated solution. When 50 ppb and higher concentrations of Al ion were added to the phosphate buffer solution, the formation of white particles containing Al, P and O was detected. It is suggested that a phosphate buffer solution in a borosilicate glass vial has the ability to form particles due to interactions with the Al that is eluted from the glass during storage.

[Multicenter surveillance of Pseudomonas aeruginosa strains for antimicrobials in Aichi prefecture in 2009].

We investigated the susceptibility to antimicrobials of 204 Pseudomonas aeruginosa strains isolated from 21 hospitals in Aichi prefecture from September to November 2009. MIC distributions of various antimicrobials were analyzed in terms of geographic region of isolation, patient status (outpatient or inpatient), and type of specimens that the strain was isolated from. The results were as follows. 1. Although more than 90% of strains were susceptible to all aminoglycosides and colistin, 80-90% of them were susceptible to beta-lactams and fluoroquinolones. MIC distributions of all antimicrobials measured were not significantly different between regions. 2. Only 1 strain (0.5%) was multi-drug resistant Pseudomonas aeruginosa (MDRP). Thirteen strains (6.4%) showed imipenem MIC > or = 16 microg/mL, and 16 strains (7.8%) showed ciprofloxacin MIC > or = 4 microg/mL. These strains tended to be more isolated from urine, respiratory tract specimens, or surgical specimens. 3. The MICs of tazobactam/piperacillin, panipenem, meropenem, doripenem, biapenem, sulbactam/cefoperazone, cefepime, and aztreonam were significantly higher in strains isolated from inpatients than in those from outpatients. MIC distributions of antimicrobials other than beta-lactams were not significantly different between situations where strains were isolated. 4. MIC distributions of piperacillin, all carbapenems, cefepime, gentamicin, and all fluoroquinolones were significantly different among samples from which strains were isolated. The strains isolated from blood showed lower MICs against all antimicrobials than those from other samples. No difference was found in MIC distributions when categorized according to bacteremic origin. The MICs were apparently elevated against beta-lactams, fluoroquinolones, and gentamicin in strains isolated from respiratory tract specimens, and against beta-lactams, and fluoroquinolones in strains isolated from urine. It was suggested that in P. aeruginosa surveillance, the results should be reported by stratifying with patient status, and type of specimens that the strain was isolated from and that regional surveillance should be useful with such stratification to establish antibiograms for empirical antimicrobial choice.

Using terahertz reflectance spectroscopy to quantify drug substance in tablets.

The purpose of this research is to investigate the applicability of terahertz (THz) reflectance spectroscopy for quantification of drug substance in tablets, so as to demonstrate the feasibility for applying this technique to tableting process monitoring. In order to acquire a suitable absorbance spectrum for this purpose, it was necessary to enhance the reflection intensity. By using an aluminum plate as a mirror at the opposite surface of the tablet, a reasonable absorbance spectrum could be acquired to reflect the bulk information of the tablet. To assess the limit of tablet thickness, linearity between the tablet thickness and the absorbance value was investigated using lactose and mannitol tablets. Since linearity was found within 0.75-5.0 mm for both tablets using 0.4 and 0.8 THz region, it was confirmed that THz reflectance spectroscopy is applicable to tablets within at least 5.0 mm thickness. Mannitol tablets containing sodium salicylate as the model drug substance were used to investigate the quantitative performance of this technique. It was confirmed that the established calibration model was acceptable for this quantification because of the root-mean-squared error of cross-validation (RMSECV) being 1.95%. In order to evaluate the applicability of this technique, content quantitative performance in double layered tablets having active and placebo layers were assessed. Since this calibration model achieved the root-mean-squared error of prediction (RMSEP) of 1.42% for the double layered tablets, this technique was considered feasible even if the drug substance is localized in the tablets.

Chemical mapping of hydration and dehydration process of theophylline in tablets using terahertz pulsed imaging.

The purpose of this research is to investigate the topographical pattern of hydration and dehydration (also known as pseudo-polymorphic change) of drug substance in drug product using terahertz (THz) pulsed imaging. Emphasis is placed on (1) applicability of THz pulsed imaging and (2) kinetic analysis in the pseudo-polymorphic change. Either anhydrous or monohydrated form of theophylline was used as the drug substance, leading to initially anhydrous or monohydrated tablets. These tablets were stored at 25°C to keep the relative humidity constant at 84% (anhydrous tablets) or 45% (monohydrated tablets), respectively. The THz pulsed imaging was confirmed to enable visualization that the hydration of the anhydrous form or the dehydration of the monohydrated form began on the surface of the tablets and gradually progressed to the core side in the tablets with storage. Kinetic studies indicated that these pseudo-polymorphic changes followed the phase boundary mechanism. Since the other imaging techniques has been scarcely achieved to show the topographical pattern of pseudo-polymorphic change of drug substances in drug products directly and visually, it is considered that THz pulsed imaging has a potential ability to solve complicated issues in pharmaceutical development.

Prevalence of a streptococcal inhibitor of a complement-mediated cell lysis-like gene (sicG) in Streptococcus dysgalactiae subsp. equisimilis.

Streptococcus dysgalactiae subsp. equisimilis isolates (n = 110) were analyzed by PCR to determine whether the gene encoding SICG, a homolog of Streptococcus pyogenes SIC, was present. Nineteen strains (17%) had this gene of which 11 (55%) were isolated from patients with invasive disease. All 19 strains possessed group G carbohydrate. Molecular characterization of emm type revealed that the majority of emm sequences were stG643 and stG2078. Only the N-terminal sequence of SICG was similar to that of SIC in S. pyogenes. Although we found no significant relationship between pathogenic severity and sicG possession, further investigation into the mechanism of SICG may elucidate the virulence in S. dysgalactiae subsp. equisimilis infection.

Development of a rapid process monitoring method for dry-coated tableting process by using near-infrared spectroscopy.

A nondestructive transmittance near-infrared (NIR) method for detecting off-centered cores in dry-coated (DC) tablets was developed as a monitoring system in the DC tableting process. Caffeine anhydrate was used as a core active pharmaceutical ingredient (API), and DC tablets were made by the direct compression method. NIR spectra were obtained from these intact DC tablets using the transmittance method. The reference assay was performed with HPLC. Calibration models were generated by partial least squares (PLS) regression and principal component regression (PCR) utilizing external validations. Hierarchical cluster analysis (HCA) of the results confirmed that NIR spectroscopy correctly detected off-centered cores in DC tablets. We formulated and used the Centering Index (CI) to evaluate the precision of core alignment and generated an NIR calibration model that could correctly predict this index. The principal component (PC) 1 loading vector of the final calibration model indicated that it could specifically detect the misalignment of tablet cores. The model also had good linearity and accuracy. The CIs of unknown sample tablets predicted by the final calibration model and those calculated through the HPLC analysis were closely parallel with each other. These results demonstrate the validity of the final calibration model and the utility of the transmittance NIR spectroscopic method developed in this study as a monitoring system in DC tableting process.

Do amorphous troglitazones prepared from two diastereomer-pairs have the same molecular mobility and crystallization rate at the surface?

The surface of amorphous compounds crystallizes faster compared to the bulk. This suggests that molecules at the surface have high molecular mobility. Crystallization behavior is affected by various factors including molecular weight and glass transition temperature (T(g)). In this study, we focus on troglitazone which is composed of diastereomers, RR/SS and RS/SR, as model compound, because each diastereomer has the same molecular weight and similar chemical structure. Troglitazone is isolated into each diastereomer, and both amorphous prepared from RR/SS and RS/SR showed similar T(g) (around 60°C). The surface relaxation of each amorphous troglitazone prepared from two diastereomers, RR/SS and RS/SR, was determined to compare surface molecular mobility, using inverse gas chromatography under dry conditions. As a result, amorphous prepared from RS/SR, showed the shorter surface relaxation time at 40°C (temperature below T(g)), which means it has higher molecular mobility than that from RR/SS at the surface although both have the same molecular weight and similar T(g). Microscopy analysis was conducted to observe the crstallization behavior at the surface of amorphous troglitazone in conditions of high temperature and humidity. Micrographs showed that crystallization area at the surface of amorphous prepared from RS/SR, which showed the shorter surface relaxation time, increased faster than that of the amorphous prepared from RR/SS. Although the reason for the difference in the surface relaxation time of each amorphous troglitazone could not be determined, factors such as the difference of configuration might affect the difference.

A retrospective study of the epidemiology of Clostridium difficile infection at a University Hospital in Japan: genotypic features of the isolates and clinical characteristics of the patients.

Clostridium difficile is a major cause of antibiotic-associated diarrhea and frequently results in healthcare-associated infections. The epidemiology of C. difficile infection (CDI), including the prevalent polymerase chain reaction (PCR) ribotypes and the clinical characteristics of the patients, is not well known in Japan, compared to the situation in the United States and Europe. We performed PCR ribotyping of C. difficile isolates from 71 consecutive patients with CDI at a University Hospital over a 3-year period and investigated the clinical features of those patients. CDI was diagnosed when a patient with diarrhea or colitis was found to have toxin B-positive C. difficile with no other enteropathogenic microorganisms. Toxin A-positive, toxin B-positive, binary toxin-positive (A(+)B(+)CDT(+)) strains; toxin A-positive, toxin B-positive, binary toxin-negative (A(+)B(+)CDT(-)) strains; and toxin A-negative, toxin B-positive, binary toxin-negative (A(-)B(+)CDT(-)) strains were isolated from 4, 58, and 9 patients, respectively, indicating that infections with binary toxin-positive strains were uncommon (5.6%). PCR ribotyping of the isolates demonstrated that among the 71 strains, 20 different PCR ribotypes were identified and that types smz, yok, and hr were predominant (19, 14, and 13 isolates, respectively), all of which were A(+)B(+)CDT(-). No specific time periods or wards were found to be associated with the three types; PCR ribotyping analysis clearly showed that the three types spread almost evenly in all wards for the 3 years studied. Comparative analysis of the clinical characteristics of patients harboring the three C. difficile types indicated that the duration of CDI was longer in the yok group than in the hr group. PCR ribotyping, which is easy to perform, appears to give us useful information to trace CDI cases in clinical settings. Further, the analysis of a large number of CDI cases may allow evaluation of the possible relationship between specific C. difficile types and the clinical features of patients.

Stabilization of amorphous indomethacin by co-grinding in a ternary mixture.

Mechanochemical amorphization of indomethacin (IM) was substantially enhanced by grinding with SiO(2), talc and a Mg(OH)(2)-SiO(2) mixture. The rates of the mechanochemical amorphization were in the order of Mg(OH)(2)-SiO(2) mixture>talc>SiO(2). Amorphous state stability of IM compounded with the carrier was examined by crystallization behavior under the condition of 30 degrees C and 11% relative humidity. Superiority of the binary mixture as a carrier was explained in terms of the mechanically induced strong acidic sites of the carrier.

Prediction of apparent equilibrium solubility of indomethacin compounded with silica by 13C solid state NMR.

The apparent equilibrium solubility (AES) of indomethacin increased by co-grinding with silica. Change in the long- and short range disorder of indomethacin by co-grinding was examined by X-ray powder diffraction and 13C solid state NMR, respectively, to elucidate the increased AES. Since the increase in AES was particularly marked after complete disappearance of X-ray diffraction peaks, we attributed the enhanced AES primarily to the short range disorder on the molecular basis. This was confirmed by a high correlation between the standardized full width at half maximum (SFWHM) of the specific peaks observed by 13C solid state NMR and log (AES). The correlation enables the prediction of AES as well.

Comparison between polyvinylpyrrolidone and silica nanoparticles as carriers for indomethacin in a solid state dispersion.

States of interaction between indomethacin (IM) and polyvinylpyrrolidone (PVP) in an amorphous solid dispersion prepared by co-grinding were compared with those between IM and silica nanoparticles. Changes in the carbon chemical states of the solid dispersions were evaluated based on the chemical shift in the 13C-CP/MAS-NMR. Hydrogen bonds between the amide carbonyl of PVP particles and the carboxyl groups of IM molecules were formed by co-grinding. Despite the wide difference in carrier properties, the apparent equilibrium solubility (AES) of IM in the ground IM-PVP mixture was predicted by solid state NMR on the basis of the relationship previously established for IM with SiO(2). This indicates that AES is affected solely by the state of IM, irrespective of the carrier species, and despite carrier-dependent chemical interactions.

Solid phase transition of CS-891 enantiotropes during grinding.

The physical properties of N-[1-(4-methoxyphenyl)-1-methylethyl]-3-oxo-4-aza-5a-androst-1-ene-17beta-carboxamide (CS-891), a novel and orally effective testosterone 5-reductase inhibitor, were investigated by differential scanning calorimetry, powder X-ray diffraction at elevated temperature and single crystal X-ray crystallography. CS-891 was revealed to exist as two enantiotropic forms, a low-temperature stable form (Form A) and a high-temperature stable form (Form B) which reversibly transforms to Form A at around 58 degrees C. The effect of grinding temperature on the transition of CS-891 between the amorphous and the crystalline state during grinding of the eantiotropes was examined. Form A transformed into an amorphous form during the grinding process while the product temperature was kept below the transition temperature. On the other hand, when the product temperature during grinding reached above the transition temperature, Form A transformed into an amorphous form and some of the amorphous form converted back to Form B. Form B crystallized from the amorphous form was physically stable even at below the transition temperature. The amorphous form in equilibrium with Form B exhibited remarkable physical stability in comparison with the amorphous form obtained by continued grinding below the transition temperature.

Optimization of secondary drying condition for desired residual water content in a lyophilized product using a novel simulation program for pharmaceutical lyophilization.

The aim of this study was to optimize the shelf temperature and the drying time, mainly dependent on the residual water content of a lyophilized product using a novel simulation program for the secondary drying of lyophilization. The simulation program was developed based upon heat transfer formulas, two empirical formulas, and a modified Fick's second law. When a preliminary lyophilization run of secondary drying was carried out, the equilibrium product temperature at the end of secondary drying under various shelf temperatures was accurately predicted by the heat transfer formulas. The apparent diffusion coefficient of water, Deff, and the apparent equilibrium residual water content, We, under the predicted equilibrium product temperature were estimated by two empirical formulas. These estimated Deff and We allow the modified Fick's second law to predict the residual water content in the lyophilized product. Using the developed simulation program, it was verified that the secondary drying condition to achieve the desired residual water content in the lyophilized product was successfully predicted. Therefore, the simulation program can be used to effectively design the secondary drying condition of lyophilization cycles without a trial and error approach.

Verification of model development technique for NIR-based real-time monitoring of ingredient concentration during blending.

There has been a considerable research on the process analytical technology (PAT) and real-time monitoring based on NIR, but the model development is still an important issue and persons in charge have difficulty in building good models. In this study, to realize efficient NIR-based real-time monitoring of ingredient concentration and establish a model development method, we investigated the effect of a calibration set, spectral preprocessing, wavelengths, and other factors on the prediction error through pilot and commercial scale blending experiments. The results confirmed that the small prediction error was realized by a calibration set, including dynamic measurement spectra acquired with the target blender. In addition, the results demonstrated that locally weighted partial least squares (LW-PLS) achieved the smaller prediction error than conventional PLS. The present study has also clarified that spectral preprocessing methods and wavelengths selected to build a model affect the prediction error of ingredient concentration interactively. A wide wavelength range should be selected when the spectral preprocessing does not lessen the effect of baseline variation, while a narrow wavelength range should be selected when it strongly decreases the effect.

Real-time monitoring of lubrication properties of magnesium stearate using NIR spectrometer and thermal effusivity sensor.

Real-time monitoring techniques based on an NIR spectrometer and a thermal effusivity sensor for lubrication properties of magnesium stearate (Mg-St) are proposed. The lubrication properties of Mg-St are defined by its concentration distribution and flatting state in a mixture. The concentration distribution of Mg-St significantly affects the absorbance in the NIR wavelength region between 1128 nm and 1240 nm. Thus, the absorbance area in this region after baseline correction was selected as a monitoring index (MI). In laboratory-scale experiments, the difference of Mg-St concentration distribution could be detected by the proposed MI with high sensitivity. In addition, experimental results using spherical mannitol granules confirmed that the changes of the flatting state of Mg-St could also be detected by the proposed MI. Similar experiments with spherical mannitol granules and the thermal effusivity sensor confirmed that effusivity could also be used to detect the changes of the flatting state of Mg-St. The applicability of these monitoring techniques was verified using a 2000 L commercial-scale blender equipped with the NIR spectrometer and the thermal effusivity sensor. The results showed that both lubrication properties could be monitored by the proposed MI, and that the flatting state of Mg-St could be monitored more sensitively by using the effusivity.

Determination for dry layer resistance of sucrose under various primary drying conditions using a novel simulation program for designing pharmaceutical lyophilization cycle.

Dry layer resistance, which is the resistance of dried cake against water vapor flow generated from sublimation, is one of the important parameters to predict maximum product temperature and drying time during primary drying in lyophilization. The purpose of this study was to develop the predictive model of dry layer resistance under various primary drying conditions using the dry layer resistance obtained from a preliminary lyophilization run. When the maximum dry layer resistance was modified under the assumption that the chamber pressure is zero, the modified dry layer resistance, which is defined as specific dry layer resistance, correlated well with the sublimation rate. From this correlation, the novel predictive model including the empirical formula of sublimation rate and specific dry layer resistance is proposed. In this model, the dry layer resistance under various conditions of shelf temperature and chamber pressure was successfully predicted based on the relationship of the sublimation rate and specific dry layer resistance of the edge and center vials obtained from the product temperature in one preliminary cycle run. It is expected that this predictive model could be a practical and useful tool to predict product temperature during primary drying.

Fc domain mediated self-association of an IgG1 monoclonal antibody under a low ionic strength condition.

Recently, we reported that IgG1 monoclonal antibody A (MAb A) underwent liquid-liquid phase separation and separated into light and heavy phases under a low ionic strength condition. The liquid-liquid phase separation was induced due to self-association of MAb A in the heavy phase when the initial concentration of MAb A was between the two critical concentrations [Nishi et al., Pharm. Res., 27, 1348-1360 (2010)]. Here, we determined the interaction site of MAb A by using proteolytic Fab and Fc fragments of MAb A. The mean hydrodynamic diameter of the Fc fragment increased in a low ionic strength buffer, and furthermore the SPR measurement detected interactions of the Fc fragment with both whole MAb A and the Fc fragment, whereas the Fab fragment interacted with neither whole MAb A nor the Fc fragment. No binding was detected under an isotonic ionic strength condition. Zeta potential of MAb A was significant positive below pH 5.5 and negative above pH 6.5. Between pH 5.5 and 6.5 where the phase separation is significantly induced, MAb A had only a small positive or negative net charge. The isothermal titration calorimetry dilution method revealed that dissociation of MAb A accompanied endothermic heat changes, suggesting that intermolecular interactions among MAb A molecules were attributed to the enthalpically driven process. These results suggest that liquid-liquid phase separation of MAb A is mediated by a weak electrostatic intermolecular interaction among MAb A molecules mainly at Fc portions.