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.

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.

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).

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.

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.

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.

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.

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).

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.

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.

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.

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.

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).

Molecular mobility of nifedipine-PVP and phenobarbital-PVP solid dispersions as measured by 13C-NMR spin-lattice relaxation time.

Amorphous nifedipine-PVP and phenobarbital-PVP solid dispersions with various drug contents were prepared by melting and subsequent rapid cooling of mixtures of PVP and nifedipine, or phenobarbital. Chemical shifts and spin-lattice relaxation times (T(1)) of PVP, nifedipine, and phenobarbital carbons were determined by (13)C-CP/MAS NMR to elucidate drug-PVP interactions and the localized molecular mobility of drug and PVP in the solid dispersions. The chemical shift of the PVP carbonyl carbon increased as the drug content increased, appearing to reach a plateau at a molar ratio of drug to PVP monomer unit of approximately 1:1, suggesting hydrogen bond interactions between the PVP carbonyl group and the drugs.

Comparison of the glass transition temperature and fragility parameter of isomalto-olygomer predicted by molecular dynamics simulations with those measured by differential scanning calorimetry.

The purpose of this study is to examine whether molecular dynamics (MD) simulations using a commercially available software for personal computers can estimate the glass transition temperature (Tg) of amorphous systems containing pharmaceutically-relevant excipients. MD simulations were carried out with an amorphous matrix model constructed from isomaltoheptaose, and the Tg estimated from the calculated density versus temperature profile was compared with the Tg measured by differential scanning calorimetry (DSC) for freeze-dried isomalto-oligomer having an average molecular weight close to that of isomaltoheptaose. The Tg values determined by DSC were lower by 10 to 20 K than those extrapolated from the Tg values estimated by MD simulation.

A quantitative assessment of the significance of molecular mobility as a determinant for the stability of lyophilized insulin formulations.

Purpose: The purpose was to explore a method for quantitatively assessing the contribution of molecular mobility to the chemical reactivity of amorphous solids. Degradation of insulin in lyophilized formulations containing trehalose and poly(vinylpyrrolidone)(PVP) was chosen as a model system, and the temperature- and glass transition temperature (Tg)-dependence of the degradation rate was analyzed to obtain the relative contributions of molecular mobility and that of the chemical activational barrier reflected in the energy of activation.

Methods: Insulin degradation and dimerization in lyophilized trehalose and PVP formulations were monitored at various relative humidities (6-60% RH) and temperatures (10-60 degrees C) by reverse-phase high-performance liquid chromatography (HPLC) and high-performance size-exclusion chromatography (HP-SEC), respectively.

Effect of freezing rate on physical stability of lyophilized cationic liposomes.

Factors affecting the storage stability of lyophilized cationic liposomes were investigated using liposomes prepared with various excipients and by different freezing rates, either quick freezing (freezing by immersion into liquid nitrogen) or slow freezing (cooling to -50 degrees C at a rate of -10 degrees C/h). Increases in the particle size of cationic liposomes observed during freeze-drying were inhibited by the addition of sucrose, trehalose and sucrose-dextran mixtures (1 : 1 and 2 : 1 by weight). The storage instability of the formulations, as indicated by changes in particle size, was affected by their glass transition temperature (T(g)).

Glass transition-related changes in molecular mobility below glass transition temperature of freeze-dried formulations, as measured by dielectric spectroscopy and solid state nuclear magnetic resonance.

The purpose of this study was to explore why changes in the molecular mobility associated with glass transition, the timescale of which is on the order of 100 s, can be detected by measuring the nuclear magnetic resonance relaxation times that reflect molecular motions on the order of 10 kHz and 1 MHz. The molecular motions in freeze-dried dextran 40k, dextran 1k, isomaltotriose (IMT), and alpha-glucose comprising a common unit but with different glass transition temperatures, were investigated by dielectric spectroscopy (DES) in the frequency range of 0.01 Hz to 100 kHz and in the temperature range of -20 degrees to 200 degrees C, in order to compare with the molecular motions reflected in nuclear magnetic resonance relaxation times.

Ability of polyvinylpyrrolidone and polyacrylic acid to inhibit the crystallization of amorphous acetaminophen.

The inhibition of crystallization of amorphous acetaminophen (ACTA) by polyvinylpyrrolidone (PVP) and polyacrylic acid (PAA) was studied using amorphous solid dispersions prepared by melt quenching. Co-melting with PVP and PAA decreased the average molecular mobility, as indicated by increases in glass transition temperature and enthalpy relaxation time. The ACTA/PAA dispersion exhibited much slower crystallization than the ACTA/PVP dispersion with a similar glass transition temperature value, indicating that interaction between ACTA and polymers also contributed to the stabilizing effect of these polymers.

Temperature- and glass transition temperature-dependence of bimolecular reaction rates in lyophilized formulations described by the Adam-Gibbs-Vogel equation.

Bimolecular reaction rates in lyophilized aspirin-sulfadiazine formulations containing poly(vinylpyrrolidone), dextran, and isomalto-oligomers of different molecular weights were determined in the presence of various water contents, and their temperature- and glass transition temperature (Tg)-dependence was compared with that of structural relaxation time (tau calculated according to the Adam-Gibbs-Vogel equation, in order to understand how chemical degradation rates of drugs in lyophilized formulations are affected by molecular mobility. The rate of acetyl transfer in poly(vinylpyrrolidone) K30 and dextran 40k formulations with a constant Tg, observed at various temperatures, exhibited a temperature dependence similar to that of tau at temperatures below Tg. Furthermore, the rates of acetyl transfer and the Maillard reaction in formulations containing alpha-glucose polymers and oligomers increased, as the Tg of formulations decreased, either associated with decreases in molecular weight of excipient or with increases in water content.

Molecular mobility-based estimation of the crystallization rates of amorphous nifedipine and phenobarbital in poly(vinylpyrrolidone) solid dispersions.

The overall crystallization rates and mean relaxation times of amorphous nifedipine and phenobarbital in the presence of poly(vinylpyrrolidone) (PVP) were determined at various temperatures to gain further insight into the effect of molecular mobility on the crystallization rates of amorphous drugs and the possibility of predicting stability from their molecular mobility. Nifedipine-PVP (9:1 w/w) and phenobarbital-PVP (95:5 w/w) solid dispersions were prepared by melting and rapidly cooling mixtures of each drug and PVP. The amount of amorphous nifedipine remaining in the solid dispersion was calculated from the heat of crystallization,which was obtained by differential scanning calorimetry.

Inactivation and aggregation of beta-galactosidase in lyophilized formulation described by Kohlrausch-Williams-Watts stretched exponential function.

Purpose: To examine whether the empirical Kohlrausch-Williams-Watts (KWW) equation is applicable not only to protein aggregation but also to protein denaturation in lyophilized formulations. Lyophilized beta-galactosidase (beta-GA) formulations containing polyvinylalcohol and methylcellulose were used as model formulations. The possibility of predicting storage stability based on the temperature dependence of the estimated parameters of inactivation/aggregation--time constant (tau) and its distribution (beta) is discussed.

Molecular mobility of lyophilized poly(vinylpyrrolidone) and methylcellulose as determined by the laboratory and rotating frame spin-lattice relaxation times of 1H and 13C.

Laboratory- and rotating- frame spin-lattice relaxation times (T(1) and T(1rho)) of (1)H and (13)C in lyophilized poly(vinylpyrrolidone) (PVP) and methylcellulose (MC) are determined to examine feasibility of using T(1) and T(1rho) as a measure of molecular motions on large time scales related to the storage stability of lyophilized formulations. The T(1rho) of proton and carbon was found to reflect the mobility of PVP and MC backbones, indicating that it is useful as a measure of large-time-scale molecular motions. In contrast to the T(1rho), the T(1) of proton measured in the same temperature range reflected the mobility of PVP and MC side chains.

Prediction of glass transition temperature of freeze-dried formulations by molecular dynamics simulation.

Purpose: To examine whether the glass transition temperature (Tg) of freeze-dried formulations containing polymer excipients can be accurately predicted by molecular dynamics simulation using software currently available on the market. Molecular dynamics simulations were carried out for isomaltodecaose, a fragment of dextran, and alpha-glucose, the repeated unit of dextran. in the presence or absence of water molecules.

Different molecular motions in lyophilized protein formulations as determined by laboratory and rotating frame spin-lattice relaxation times.

The spin-lattice relaxation times in the laboratory and rotating frame (T(1) and T(1rho)) of protons and carbons in lyophilized bovine serum gamma-globulin formulation containing dextran were determined by (1)H solid-state pulsed nuclear magnetic resonance (NMR) and high-resolution (13)C solid-state NMR. The temperature dependence of T(1) and T(1rho) of dextran protons in the lyophilized formulation suggests that the correlation time, tau(c), of the methylene protons in dextran is approximately 10(-6) s at -100 degrees C and 60% relative humidity, and decreases to 10(-7) s at 0 degrees C. When temperature is increased from 0 degrees C, the increased motion of the methylene groups is reflected in T(1), but is too fast to be observed by changes in T(1rho).