[Detection and Analysis of Drug Crystals in Medical Transdermal Patches by Using X-ray Diffraction Measurement].

The crystallization of active pharmaceutical ingredients (APIs) in matrix-type transdermal patches has implications for the rate of drug absorption through the skin and patch adhesion strength. Therefore, the presence or absence and the degree of API crystallinity must be controlled to guarantee the quality of patches. In this study, the utility of laboratory-level X-ray diffractometers for the detection and analysis of crystalline APIs in transdermal patches was investigated using medical patches of tulobuterol and isosorbide dinitrate. Several matrix-type patches employ a controlled drug delivery system containing intentionally crystallized API. Both benchtop and high-resolution laboratory X-ray diffractometers can detect several characteristic peaks of the APIs in these patches even if the patches are wrapped in an outer bag, although a benchtop model provides peak heights one-seventh to one-fifth that of a high-resolution instrument. An isosorbide dinitrate patch containing an unintentionally crystallized spot was wrapped in an outer bag, followed by measurements using both X-ray diffractometers. For both instruments, several isosorbide dinitrate-derived peaks were detected only at the crystallized spot, although the signal-to-noise ratio was poorer for the benchtop model. These results show that a high-resolution X-ray diffractometer is advantageous for high-detection sensitivity and offers a high degree of freedom of the measurement position on the sample. It was concluded that a laboratory-level high-resolution X-ray diffractometer can be used to examine the crystalline state of APIs in patches inside an unopened outer bag.

[Comparison of Adhesive Properties of Transdermal Patches Distributing in Japan-Tack and Peel Strength].

Forty-four brands of transdermal patches for twelve kinds of active pharmaceutical ingredients (APIs) are available in Japan as of April 30, 2018. Although approximately one-third of the corresponding pharmaceutical interview forms lack information on how to evaluate the adhesive properties of the patches, the peel test, probe tack test, or inclined ball tack test have generally been adopted. This means that it might be difficult to simply compare the adhesive properties among the patches because the testing methods are not unified in some cases. In this study, measurements of the adhesive properties of 38 transdermal patches of ten different APIs were performed using several unified testing methods (180° peel test, 90° peel test, self-adhesion test, and probe tack test) under unified experimental conditions. The adhesive properties were found to be quite different among the patches, even for the same API, dose, and size. For example, the ratios of the maximum to minimum measured values of tack and 180° peel strength for tulobuterol patches were 5 and 29, respectively. In the case of generic products for which the bioequivalence to a brand-name product is assured, the variation in adhesive properties can extend the range of choices for patients, which is advantageous. Providing information to medical experts on adhesive properties through, for example, pharmaceutical interview forms and package inserts, is considered to be useful for helping patients to make better choices.

[Ion-pair HPLC analysis of B vitamins in syrup products in Indonesia].

A training course for analysis of B vitamins in syrup products was undertaken at the National Agency of Drug and Food Control at Jakarta as part of the project to deliver safe drugs to people in Indonesia by Japan International Cooperation Agency. Analytical methods have been developed for quantitative determination of B vitamins by ion-pair high-performance liquid chromatography using 1-hexanesulfonic acid sodium salt. Measurements were performed for two syrup products removed from a drug store in Jakarta to determine the amount of each vitamin B. The measured values of riboflavin 5'-phosphate sodium, nicotinamide and pyridoxine hydrochloride were almost the same with those of nominal content for both products. While the measured values of thiamine hydrochloride, pantothenol and cyanocobalamin were approximately twice the amount of nominal contents.

[Studies on the food allergenic proteins contained in pharmaceutical excipients].

Most drugs contain pharmaceutical excipients. These are pharmacologically inactive substances used as vehicles for the active ingredients of a medication. Some of these pharmaceutical excipients are produced from allergenic foods (e.g., milk, egg, peanut, soybean, and sesame) and removing proteins completely from such excipients is difficult. Therefore, if individuals with food allergy consume drugs containing allergenic food-derived excipients, eliminating the risk of developing specific allergic symptoms induced by them may not be possible. We determined the levels of proteins in pharmaceutical excipients and ethical drugs (inhalants and injections) by spectrophotometric analyses. The level of protein in the pharmaceutical excipient lactose in each sample was approximately 1 mg/g. In the case of oils from soybeans, peanuts, and sesame in pharmaceutical excipients, proteins were detected in the range 7-9 microg/g sample. We also determined levels of allergenic proteins in pharmaceutical excipients and ethical drugs using commercial enzyme-linked immunosorbent assay systems. The milk proteins in lactose were detected in the range 1.39-13.07 microg/g. The results of this study suggest that physicians, patients with food allergies, pharmacists, and healthcare providers must pay attention to presence of potential impurities those may cause allergic symptoms in pharmaceutical products.

Hydration of surfactant-modified and PEGylated cationic cholesterol-based liposomes and corresponding lipoplexes by monitoring a fluorescent probe and the dielectric relaxation time.

For the optimization of plasmid DNA (pDNA)-cationic lipid complexes and lipoplex delivery, proper indexes of the physicochemical properties of lipoplexes are required. In general, the characteristics of lipoplexes are defined by particle size and zeta-potential at various mixing ratios of cationic liposomes and pDNA. In this study, we characterized the hydration level of surfactant-modified and PEGylated cationic cholesterol-based (OH-Chol) liposomes and their lipoplexes by monitoring both the fluorescent probe laurdan and the dielectric relaxation time. Fluorescence measurement using laurdan detected hydration of the headgroup of lipids in surfactant-modified liposomes and PEGylated DOTAP-liposomes, but hardly any fluorescence was detected in PEGylated OH-Chol-liposomes because the PEG layers may extend and cover the fluorescent maker. On the other hand, the measurement of dielectric relaxation time of water molecules revealed total hydration, including hydration of the PEG layer and the headgroup of cationic lipids. Furthermore, we found an inverse correlation between hydration level and cellular uptake of PEGylated lipoplexes (R=0.946). This finding indicated that the dielectric relaxation time of water molecules provides an important indicator of hydration of liposome and lipoplexes along with the fluorescence intensity of laurdan.

Effect of sugars on the molecular motion of freeze-dried protein formulations reflected by NMR relaxation times.

PURPOSE: To relate NMR relaxation times to instability-related molecular motions of freeze-dried protein formulations and to examine the effect of sugars on these motions. METHODS: Rotating-frame spin-lattice relaxation time (T(1ρ)) was determined for both protein and sugar carbons in freeze-dried lysozyme-sugar (trehalose, sucrose and isomaltose) formulations using solid-state (13)C NMR. RESULTS: The temperature dependence of T(1ρ) for the lysozyme carbonyl carbons in lysozyme with and without sugars was describable with a model that includes two different types of molecular motion with different correlation times (τ(c)) for the carbon with each τ(c) showing Arrhenius temperature dependence. Both relaxation modes have much smaller relaxation time constant (τ(c)) and temperature coefficient (Ea) than structural relaxation and may be classified as ß-relaxation and γ-relaxation. The τ(c) and Ea for γ-relaxation were not affected by sugars, but those for ß-relaxation were increased by sucrose, changed little by trehalose, and decreased by isomaltose, suggesting that the ß-mobility of the lysozyme carbonyl carbons is decreased by sucrose and increased by isomaltose. CONCLUSION: T(1ρ) determined for the lysozyme carbonyl carbons can reflect the effect of sugars on molecular mobility in lysozyme. However, interpretation of relaxation time data is complex and may demand data over an extended temperature range.

Feasibility of atomic force microscopy for determining crystal growth rates of nifedipine at the surface of amorphous solids with and without polymers.

Amorphous nifedipine (NFD), which has a smooth surface immediately after preparation, was shown to have structures resembling clusters of curling and branching fibers approximately 1 µm wide by atomic force microscopy (AFM) after storage at 25°C. The size of the cluster-like structures increased with storage over time, implying crystal growth. The average elongation rate of the fibers determined by AFM at ambient room temperature was 1.1 × 10(-9) m/s, and this agreed well with the crystal growth rate of 1.6 × 10(-9) m/s determined by polarized light microscopy. The crystal growth rate of NFD in solid dispersions with 5% polyethylene glycol (PEG) was found to be 5.0 × 10(-8) m/s by AFM. Although this value was approximately the same as that obtained by polarized light microscopy, three-dimensional information obtained by AFM for the crystallization of NFD in a solid dispersion with PEG revealed that the changes in topography were not a consequence of surface crystal growth, but rather attributable to the growth of crystals formed in the amorphous bulk. For solid dispersions with α,ß-poly(N-5-hydroxypentyl)-l-aspartamide, acceleration of NFD crystallization by tapping with an AFM probe was observed. The present study has demonstrated the feasibility and application of AFM for interpretation of surface crystallization data.

Differences in crystallization rate of nitrendipine enantiomers in amorphous solid dispersions with HPMC and HPMCP.

To clarify the contribution of drug-polymer interaction to the physical stability of amorphous solid dispersions, we studied the crystallization rates of nitrendipine (NTR) enantiomers with identical physicochemical properties in the presence of hydroxypropylmethylcellulose (HPMC), hydroxypropylmethylcellulose phthalate (HPMCP) and polyvinylpyrrolidone (PVP). The overall crystallization rate at 60°C and the nucleation rate at 50-70°C of (+)-NTR were lower than those of (-)-NTR in the presence of 10-20% HPMC or HPMCP. In contrast, similar crystallization profiles were observed for the NTR enantiomers in solid dispersions containing PVP. The similar glass transition temperatures for solid dispersions of (-)-NTR and (+)-NTR suggested that the molecular mobility of the amorphous matrix did not differ between the enantiomers. These results indicate that the interaction between the NTR enantiomers and HPMC or HPMCP is stereoselective, and that differences in the stereoselective interaction create differences in physical stability between (-)-NTR and (+)-NTR at 50-70°C. However, no difference in physical stability between the enantiomers was obvious at 40°C. Loss of the difference in physical stability between the NTR enantiomers suggests that the stereoselective interaction between NTR and the polymers may not contribute significantly to the physical stabilization of amorphous NTR at 40°C.

The impact of thermal treatment on the stability of freeze-dried amorphous pharmaceuticals: II. Aggregation in an IgG1 fusion protein.

The objective of this research was to investigate the impact of thermal treatment on storage stability of an IgG1 fusion protein. IgG1 protein formulations were prepared by freeze-drying the protein with sucrose. Some samples were used as controls, and others were subjected to a further heat treatment (annealing). The protein structure was investigated with Fourier transform infrared spectroscopy (FTIR), and protein aggregation was monitored with size exclusion HPLC. Enthalpy recovery was studied using DSC, and global mobility represented by the structural relaxation time constant (tau(beta)) was characterized by a thermal activity monitor (TAM). The local mobility of the protein system was monitored by both (13)C solid-state NMR and neutron backscattering. Annealing increased the storage stability of the protein, as shown by the smaller aggregation rate and less total aggregation at the end of a storage period. The structural relaxation time constant of an annealed sample was significantly higher than the unannealed control sample, suggesting a decrease in global mobility of the protein system upon annealing. However, annealing does not significantly impact the protein secondary structure or the local mobility. Given the similar protein native structure and specific surface area, the improved stability upon annealing is mainly a result of reduced global molecular mobility.

Feasibility of 19F-NMR for assessing the molecular mobility of flufenamic acid in solid dispersions.

The purpose of the present study was to clarify the feasibility of 19F-NMR for assessing the molecular mobility of flufenamic acid (FLF) in solid dispersions. Amorphous solid dispersions of FLF containing poly(vinylpyrrolidone) (PVP) or hydroxypropylmethylcellulose (HPMC) were prepared by melting and rapid cooling. Spin-lattice relaxation times (T1 and T(1rho)) of FLF fluorine atoms in the solid dispersions were determined at various temperatures (-20 to 150 degrees C). Correlation time (tauc), which is a measure of rotational molecular mobility, was calculated from the observed T1 or T1rho value and that of the T1 or T1rho minimum, assuming that the relaxation mechanism of spin-lattice relaxation of FLF fluorine atoms does not change with temperature. The tauc value for solid dispersions containing 20% PVP was 2-3 times longer than that for solid dispersions containing 20% HPMC at 50 degrees C, indicating that the molecular mobility of FLF in solid dispersions containing 20% PVP was lower than that in solid dispersions containing 20% HPMC. The amount of amorphous FLF remaining in the solid dispersions stored at 60 degrees C was successfully estimated by analyzing the solid echo signals of FLF fluorine atoms, and it was possible to follow the overall crystallization of amorphous FLF in the solid dispersions. The solid dispersion containing 20% PVP was more stable than that containing 20% HPMC. The difference in stability between solid dispersions containing PVP and HPMC is considered due to the difference in molecular mobility as determined by tauc. The molecular mobility determined by 19F-NMR seems to be a useful measure for assessing the stability of drugs containing fluorine atoms in amorphous solid dispersions.

Effect of sugars on storage stability of lyophilized liposome/DNA complexes with high transfection efficiency.

Cationic lipid-based gene delivery systems have shown promise in transfecting cells in vitro and in vivo. However, liposome/DNA complexes tend to form aggregates after preparation. Lyophilization of these systems, therefore, has become of increasing interest. In this study, we investigated the feasibility of preserving complexes as a dried preparation using a modified dehydration rehydration vesicle (DRV) method as a convenient and reliable procedure. We also studied storage stability of a lyophilized novel cationic gene delivery system incorporating sucrose, isomaltose and isomaltotriose. Liposomes were composed of 3beta-[N-(N',N'-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and L-dioleoylphosphatidylethanolamine (DOPE), plus sucrose, isomaltose or isomaltotriose. Lyophilized liposome/DNA complexes were stored at -20, 25, 40 and 50 degrees C and their stability was followed for 50 days. Liposome/DNA complexes with sucrose could be stored even at 50 degrees C without large loss of transfection efficiency. The transfection efficiency of formulations stored at various temperatures indicated that the stabilizing effect of sugars on plasmid DNA was higher in the following order: isomaltotriose

Wide-ranging molecular mobilities of water in active pharmaceutical ingredient (API) hydrates as determined by NMR relaxation times.

In order to examine the possibility of determining the molecular mobility of hydration water in active pharmaceutical ingredient (API) hydrates by NMR relaxation measurement, spin-spin relaxation and spin-lattice relaxation were measured for the 11 API hydrates listed in the Japanese Pharmacopeia using pulsed (1)H-NMR. For hydration water that has relatively high mobility and shows Lorentzian decay, molecular mobility as determined by spin-spin relaxation time (T(2)) was correlated with ease of evaporation under both nonisothermal and isothermal conditions, as determined by DSC and water vapor sorption isotherm analysis, respectively. Thus, T(2) may be considered a useful parameter which indicates the molecular mobility of hydration water. In contrast, for hydration water that has low mobility and shows Gaussian decay, T(2) was found not to correlate with ease of evaporation under nonisothermal conditions, which suggests that in this case, the molecular mobility of hydration water was too low to be determined by T(2). A wide range of water mobilities was found among API hydrates, from low mobility that could not be evaluated by NMR relaxation time, such as that of the water molecules in pipemidic acid hydrate, to high mobility that could be evaluated by this method, such as that of the water molecules in ceftazidime hydrate.

Miscibility of nifedipine and hydrophilic polymers as measured by (1)H-NMR spin-lattice relaxation.

The miscibility of a drug with excipients in solid dispersions is considered to be one of the most important factors for preparation of stable amorphous solid dispersions. The purpose of the present study was to elucidate the feasibility of (1)H-NMR spin-lattice relaxation measurements to assess the miscibility of a drug with excipients. Solid dispersions of nifedipine with the hydrophilic polymers poly(vinylpyrrolidone) (PVP), hydroxypropylmethylcellulose (HPMC) and alpha,beta-poly(N-5-hydroxypentyl)-L-aspartamide (PHPA) with various weight ratios were prepared by spray drying, and the spin-lattice relaxation decay of the solid dispersions in a laboratory frame (T(1) decay) and in a rotating frame (T(1rho) decay) were measured. T(1rho) decay of nifedipine-PVP solid dispersions (3 : 7, 5 : 5 and 7 : 3) was describable with a mono-exponential equation, whereas T(1rho) decay of nifedipine-PHPA solid dispersions (3 : 7, 4 : 6 and 5 : 5) was describable with a bi-exponential equation. Because a mono-exponential T(1rho) decay indicates that the domain sizes of nifedipine and polymer in solid dispersion are less than several nm, it is speculated that nifedipine is miscible with PVP but not miscible with PHPA. All the nifedipine-PVP solid dispersions studied showed a single glass transition temperature (T(g)), whereas two glass transitions were observed for the nifedipine-PHPA solid dispersion (3 : 7), thus supporting the above speculation. For nifedipine-HPMC solid dispersions (3 : 7 and 5 : 5), the miscibility of nifedipine and HPMC could not be determined by DSC measurements due to the lack of obviously evident T(g). In contrast, (1)H-NMR spin-lattice relaxation measurements showed that nifedipine and HPMC are miscible, since T(1rho) decay of the solid dispersions (3 : 7, 5 : 5 and 7 : 3) was describable with a mono-exponential equation. These results indicate that (1)H-NMR spin-lattice relaxation measurements are useful for assessing the miscibility of a drug and an excipient in solid dispersions.

Significance of local mobility in aggregation of beta-galactosidase lyophilized with trehalose, sucrose or stachyose.

PURPOSE: The purpose of this study is to compare the effects of global mobility, as reflected by glass transition temperature (T(g)) and local mobility, as reflected by rotating-frame spin-lattice relaxation time (T(1rho)) on aggregation during storage of lyophilized beta-galactosidase (beta-GA). MATERIALS AND METHODS: The storage stability of beta-GA lyophilized with sucrose, trehalose or stachyose was investigated at 12% relative humidity and various temperatures (40-90 degrees C). beta-GA aggregation was monitored by size exclusion chromatography (SEC). Furthermore, the T(1rho) of the beta-GA carbonyl carbon was measured by (13)C solid-state NMR, and T(g) was measured by modulated temperature differential scanning calorimetry. Changes in protein structure during freeze drying were measured by solid-state FT-IR. RESULTS: The aggregation rate of beta-GA in lyophilized formulations exhibited a change in slope at around T(g), indicating the effect of molecular mobility on the aggregation rate. Although the T(g) rank order of beta-GA formulations was sucrose < trehalose < stachyose, the rank order of beta-GA aggregation rate at temperatures below and above T(g) was also sucrose < trehalose < stachyose, thus suggesting that beta-GA aggregation rate is not related to (T-T(g)). The local mobility of beta-GA, as determined by the T(1rho) of the beta-GA carbonyl carbon, was more markedly decreased by the addition of sucrose than by the addition of stachyose. The effect of trehalose on T(1rho) was intermediate when compared to those for sucrose and stachyose. These findings suggest that beta-GA aggregation rate is primarily related to local mobility. Significant differences in the second derivative FT-IR spectra were not observed between the excipients, and the differences in beta-GA aggregation rate observed between the excipients could not be attributed to differences in protein secondary structure. CONCLUSIONS: The aggregation rate of beta-GA in lyophilized formulations unexpectedly correlated with the local mobility of beta-GA, as indicated by T(1rho), rather than with (T-T(g)). Sucrose exhibited the most intense stabilizing effect due to the most intense ability to inhibit local protein mobility during storage.

Correlations between molecular mobility and chemical stability during storage of amorphous pharmaceuticals.

Recent studies have demonstrated that molecular mobility is an important factor affecting the chemical stability of amorphous pharmaceuticals, including small-molecular-weight drugs, peptides and proteins. However, quantitative correlations between molecular mobility and chemical stability have not yet been elucidated. The purpose of this article is to review literature describing the effect of molecular mobility on chemical stability during storage of amorphous pharmaceuticals, and to seek a better understanding of the relative significance of molecular mobility and other factors for chemical reactivity. We first consider the feature of chemical stability often observed for amorphous pharmaceuticals; changes in temperature dependence of chemical stability around matrix glass transition temperature (Tg), and greater stability associated with higher Tg. Secondly, we review papers which quantitatively studied the effects of the global mobility (often referred to as structural relaxation or -relaxation) of amorphous pharmaceuticals on chemical stability, and discuss correlations between chemical stability and global mobility using various equations that have thus far been proposed. Thirdly, the significance of local mobility of drug and excipient molecules in chemical reactivity is discussed in comparison with that of global mobility. Furthermore, we review literature reports which show no relationship between chemical stability and molecular mobility. The lack of apparent relationship is discussed in terms of the effects of the contribution of excipient molecules as reactants, the specific effects of water molecules, the heterogeneity of the matrix, and so on. The following summary has been obtained; the chemical stability of amorphous pharmaceuticals is affected by global mobility and/or local mobility, depending on the length scale of molecular mobility responsible for the chemical reactivity. In some cases, when activation energy for degradation processes is high and when other factors such as the specific effects of water and/or excipients contribute the degradation rate, stability seems to be largely independent of molecular mobility.

Crystallization rate of amorphous nifedipine analogues unrelated to the glass transition temperature.

To examine the relative contributions of molecular mobility and thermodynamic factor, the relationship between glass transition temperature (T(g)) and the crystallization rate was examined using amorphous dihydropyridines (nifedipine (NFD), m-nifedipine (m-NFD), nitrendipine (NTR) and nilvadipine (NLV)) with differing T(g) values. The time required for 10% crystallization, t(90), was calculated from the time course of decreases in the heat capacity change at T(g). The t(90) of NLV and NTR decreased with decreases in T(g) associated with water sorption. The t(90) versus T(g)/T plots almost overlapped for samples of differing water contents, indicating that the crystallization rate is determined by molecular mobility as indicated by T(g). In contrast, differences in the crystallization rate between these four drugs cannot be explained only by molecular mobility, since the t(90) values at a given T(g)/T were in the order: NLV>NTR>NFD approximately m-NFD. A lower rate was obtained for amorphous drugs with lower structural symmetry and more bulky functional groups, suggesting that these factors are also important. Furthermore, the crystallization rate of NTR in solid dispersions with poly(vinylpyrrolidone) (PVP) and hydroxypropyl methylcellulose (HPMC) decreased to a greater extent than expected from the increased T(g). This also suggests that factors other than molecular mobility affect the crystallization rate.

Physical stability of amorphous acetanilide derivatives improved by polymer excipients.

Crystallization rates of drug-polymer solid dispersions prepared with acetaminophen (ACA) and p-aminoacetanilide (AAA) as model drugs, and polyvinylpyrrolidone and polyacrylic acid (PAA) as model polymers were measured in order to further examine the significance of drug-polymer interactions. The crystallization of AAA and ACA was inhibited by mixing those polymers. The most effective inhibition was observed with solid dispersions of AAA and PAA. The combination of AAA and PAA showed a markedly longer enthalpy relaxation time relative to drug alone as well as a higher T(g) than predicted by the Gordon-Taylor equation, indicating the existence of a strong interaction between the two components. These observations suggest that crystallization is effectively inhibited by combinations of drug and polymer that show a strong intermolecular interaction due to proton transfer between acidic and basic functional groups.

Degradation rate of lyophilized insulin, exhibiting an apparent Arrhenius behavior around glass transition temperature regardless of significant contribution of molecular mobility.

The relative influences of chemical activation energy and molecular mobility in determining chemical reactivity were evaluated for insulin lyophilized with alpha,beta-poly(N-hydroxyethyl)-L-aspartamide (PHEA), and compared with that for insulin lyophilized with trehalose, which had been found to have the ability to decrease the molecular mobility of insulin at low humidity. The ratio of the observed rate constant k(obs) to the chemical activation energy-controlled rate constant k(act) (k(obs)/k(act)) at glass transition temperature (T(g)) was estimated to be approximately 0.6 and 0.8 at 6% RH and 12% RH, respectively, indicating that the degradation rate is significantly affected by molecular mobility at lower humidity conditions. However, these k(obs)/k(act) values at T(g) were larger than those for the insulin-trehalose system, and changes in the temperature-dependent slope around T(g) were less obvious than those for the insulin-trehalose system. Thus, the contribution of molecular mobility to the degradation rate in the insulin-PHEA system appeared to be less intense than that in the insulin-trehalose system. The subtle change in the temperature-dependent slope around T(g) observed in the insulin-PHEA system brought about a significant bias in shelf-life estimation when the reaction rate was extrapolated from temperatures above T(g) according to the Arrhenius equation.

Beta-relaxation of insulin molecule in lyophilized formulations containing trehalose or dextran as a determinant of chemical reactivity.

PURPOSE: The purpose of this study was to elucidate whether the degradation rate of insulin in lyophilized formulations is determined by matrix mobility, as reflected in glass transition temperature (Tg), or by beta-relaxation, as reflected in rotating-frame spin-lattice relaxation time (T1rho). METHODS: The storage stability of insulin lyophilized with dextran was investigated at various relative humidities (RH; 12-60%) and temperatures (40-90 degrees C) and was compared with previously reported data for insulin lyophilized with trehalose. Insulin degradation was monitored by reverse-phase high-performance liquid chromatography. Furthermore, the T1rho of the insulin carbonyl carbon in the lyophilized insulin-dextran and insulin-trehalose systems was measured at 25 degrees C by 13C solid-state NMR, and the effect of trehalose and dextran on T1rho was compared at various humidities. RESULTS: The degradation rate of insulin lyophilized with dextran was not significantly affected by the Tg of the matrix, even at low humidity (12% RH), in contrast to that of insulin lyophilized with trehalose. The insulin-dextran system exhibited a substantially greater degradation rate than the insulin-trehalose system at a given temperature below the Tg. The difference in degradation rate between the insulin-dextran and insulin-trehalose systems observed at 12% RH was eliminated at 43% RH. In addition, the T1rho of the insulin carbonyl carbon at low humidity (12% RH) was prolonged by the addition of trehalose, but not by the addition of dextran. This difference was eliminated at 23% RH, at which point the solid remained in the glassy state. These findings suggest that the beta-relaxation of insulin is inhibited by trehalose at low humidity, presumably as a result of insulin-trehalose interaction, and thus becomes a rate determinant. In contrast, dextran, whose ability to interact with insulin is thought to be less than that of trehalose, did not inhibit the beta-relaxation of insulin, and thus, the chemical activational barrier (activation energy) rather than beta-relaxation becomes the major rate determinant. CONCLUSIONS: Beta-relaxation rather than matrix mobility seems to be more important in determining the stability of insulin in the glassy state in lyophilized formulations containing trehalose and dextran.

Negligible contribution of molecular mobility to the degradation rate of insulin lyophilized with poly(vinylpyrrolidone).

The purpose of this study is to confirm the speculation which arose in our previous study that the degradation rate of insulin lyophilized with poly(vinylpyrrolidone) is mainly governed by the chemical activational barrier rather than molecular mobility. This speculation was based on the degradation data of insulin lyophilized with poly(vinylpyrrolidone) K-30 (PVP K-30), which was obtained at temperatures well below the glass transition temperature (T(g)). In this study, the degradation rate of insulin at temperatures below and above T(g) was determined using PVP 10k as an excipient, instead of PVP K-30, in order to examine whether or not the temperature dependence of the degradation rate changes around T(g). The relative contributions of molecular mobility and the activational barrier, calculated from the temperature- and T(g)-dependence of the degradation rate, indicated that the contribution of molecular mobility to the degradation rate was negligible. Furthermore, the negligible contribution of molecular mobility was confirmed by the lack of significant change observed in the temperature- and T(g)-dependence of the rate around T(g).