Stability of lyophilized human growth hormone.

To evaluate relationships between the extent of protein-excipient interactions, structural relaxation of an amorphous matrix, and the physico-chemical stability of a protein, human growth hormone (hGH) was lyophilized with sucrose and trehalose in a 1:2 weight ratio. The protein-excipient interactions were analyzed immediately after lyophilization with isoperibol solution calorimetry (ISC), water sorption analysis (WSA), differential scanning calorimetry (DSC) and Fourier Transform Infrared Spectroscopy (FTIR). The physical and chemical stability of hGH during storage at 50 degrees C was monitored by reverse phase (RP)-HPLC, SEC-HPLC and UV absorption spectroscopy.

Physical characterization of clostridium difficile toxins and toxoids: effect of the formaldehyde crosslinking on thermal stability.

Nosocomial diarrhea and pseudomembranous colitis causing toxins A and B from Clostridium difficile were studied at pH 5-8 and over the temperature range of 10-85 degrees C. The proteins were crosslinked with formaldehyde to inactivate them to toxoid forms and permit their use as vaccines. Structural changes and aggregation behavior were monitored with circular dichroism, intrinsic and extrinsic (ANS) fluorescence spectroscopy, turbidity measurements, high-resolution UV absorbance spectroscopy and dynamic light scattering.

Preformulation studies of Clostridium difficile toxoids A and B.

To enhance the physical stability of Clostridium difficile toxoids A and B, screening for stabilizing compounds was performed. The screening of 30 GRAS compounds at various concentrations and in several combinations was performed in two parts. First, a high-throughput aggregation assay was used to screen for compounds which delayed or prevented aggregation of toxoids under stress conditions (toxoids at pH 5-5.

Spectroscopic evaluation of the stabilization of humanized monoclonal antibodies in amino acid formulations.

The protective effects of amino acids on stabilizing protein secondary structure were evaluated using diffuse reflectance FTIR spectroscopy, and interactions between proteins and arginine were detected using solid-state NMR spectroscopy. Upon freeze-drying, excipient-free anti-CD11a and anti-IgE antibodies underwent significant changes in their secondary structures. For both antibodies, the amount of intermolecular beta-sheet substantially increased and the native conformation of intramolecular beta-sheet content decreased considerably.

Calorimetric investigation of protein/amino acid interactions in the solid state.

Possible protein/amino acid interactions and the physical states of amino acids after freeze-drying have been studied using isoperibol calorimetry and differential scanning calorimetry (DSC). Good linear correlations (R(2) = 0.99) between the enthalpies of solution and the percentage of antibody in all physical mixtures, as well as unchanging melting temperatures of amino acids for physical mixtures demonstrated that there is no interaction between the antibodies and amino acids studied upon physical mixing.

The interaction of the penetration enhancer DDAIP with a phospholipid model membrane.

Differential scanning calorimetry (DSC) was used to investigate the mechanism of action of a proprietary skin penetration enhancer, dodecyl-2-(N,N-dimethylamino)propionate (DDAIP) in dipalmitoylphosphatidylcholine (DPPC) liposomes. Furthermore, the effect of enhancer concentration on lipid thermotropic transitions was investigated. With increasing concentrations of DDAIP (from 5 to 50 mol%), the main transition peak shifted to lower temperatures and became more broad.

Investigation of protein/carbohydrate interactions in the dried state. 2. Diffuse reflectance FTIR studies.

Upon freeze-drying in the absence of lyoprotectants, Fourier transform infrared (FTIR) spectroscopy has detected changes in the secondary structures of proteins. Such FTIR studies have been typically conducted using protein/KBr pellets, where additional protein degradation could potentially occur due to pressure effects and partial dissolution of the chaotropic KBr. Diffuse reflectance FTIR spectroscopy, in which no sample preparation is necessary, was evaluated as an alternative spectroscopic method to examine protein structure upon freeze-drying.

Enhancement of skin permeation of miconazole by phospholipid and dodecyl 2-(N,N-dimethyl amino)propionate (DDAIP).

Miconazole (MCZ) has very low solubility in both water and oil. Permeation rates through shed snakeskin from an aqueous suspension and a mineral oil suspension were 0.5 microg/cm(2)/h and almost none, respectively.

Investigation of protein/carbohydrate interactions in the dried state. 1. Calorimetric studies.

Isoperibol calorimetry was used to evaluate protein/carbohydrate interactions after freeze drying. rh-DNase, rh-GH, rh-MetGH, and rh-IGF-I were freeze dried with either mannitol, sucrose, trehalose, or dextran at concentrations ranging from 0% to 100% (w/w). Enthalpies of solution for both freeze-dried and physical mixtures were measured in water at 25 degrees C.

The use of solution calorimetry with micellar solvent systems for the detection of polymorphism.

The presence of multiple polymorphic forms in seven batches of raw material of a model compound having poor wettability properties (cimetidine) was studied by solution calorimetry. Due to the large number of polymorphic forms described in the literature ('Gazz. Chim.

Novel approach to improve permeation of ondansetron across shed snake skin as a model membrane.

The purpose of this study was to investigate the feasibility of transdermal drug delivery of ondansetron, an antagonist of the 5-HT3 receptor, used for the treatment of chemotherapy-induced emesis. The permeability of ondansetron from an aqueous suspension through shed snake skin as a model membrane was very low and in order to improve it, several enhancers were tested. Ethanol increased the flux at a concentration of 40% or more.

Influence of pH on the permeability of p-toluidine and aminopyrine through shed snake skin as a model membrane.

The influence of pH on the permeability of p-toluidine (pKa, 5.3) and aminopyrine (pKa, 5.0) through shed snake skin as a model membrane was studied.

Enhanced in vitro percutaneous penetration of salicylate by ion pair formation with alkylamines.

The apparent octanol/water partition coefficient (APC) of salicylate (SA) increased as the concentration of alkylamine (amyl, hexyl, heptyl, octyl and nonylamine) in aqueous phase increased, presumably through intermolecular ion pair formation between the negatively charged SA moiety and the alkylamine cation. The true partition coefficient (TPC) and the formation constant (Kf) of the ion pair were calculated from the partition data. The skin permeability of SA increased as the APC of SA increased, when 20-fold molar excess of alkylamine was added to the donor compartment.

In vitro controlled release kinetics of local anaesthetics from poly(D,L-lactide) and poly(lactide-co-glycolide) microspheres.

Poly(D,L)lactide and polylactide-co-glycolide drug-loaded microspheres were prepared with lipopholic (bupivacaine and etidocaine) and hydrophilic (mepivacine and lidocaine) local anaesthetics. Formulations of drug-loaded microspheres were characterized by the drug content, the in-vitro release kinetics and by the physical state of the drug within the microspheres. Release rates of the local anaesthetics from the microspheres were different and could not be accounted for by the intrinsic dissolution rates of the drugs.

Effect of some penetration enhancers on epithelial membrane lipid domains: evidence from fluorescence spectroscopy studies.

The effect of the penetration enhancers Azone, oleic acid, 1-dodecanol, dodecyl N,N-dimethylaminoacetate (DDAA), and dodecyl N,N-dimethylaminoisopropionate (DDAIP) on epithelial membrane lipids was examined using human buccal cell membranes as a model for epithelial lipid bilayer. Buccal epithelial cells (BEC) were labeled with 1,6-diphenyl-1,3,5-hexatriene (DPH), 1-(4-(trimethylammonio)phenyl)-6- phenyl-1,3,5-hexatriene (TMA-DPH), and 8-anilino-1-naphthalene sulphonic acid (ANS) fluorophores to characterize enhancer-induced changes in the hydrophobic core, in the superficial polar head region, and on the exterior surface, respectively, with fluorescence anisotropy and fluorescence lifetimes. All the enhancers studied were found to decrease the BEC membrane lipid packing order in a concentration-dependent and time-dependent manner in the deep bilayer region, as shown by a 37-66% decrease in anisotropy.

Synthesis and enhancing effect of dodecyl 2-(N,N-dimethylamino)propionate on the transepidermal delivery of indomethacin, clonidine, and hydrocortisone.

The biodegradable transdermal penetration enhancer, dodecyl 2-(N,N-dimethylamino)propionate (II; DDAIP), was prepared by reacting dodecyl 2-bromopropionate (I), obtained by reaction of n-dodecanol with 2-bromopropionyl halogenide, with dimethylamine. The penetration enhancing effects of DDAIP on the transport of indomethacin, clonidine, and hydrocortisone across shed snake skin (Elaphe obsoleta) were evaluated. Azone and lauryl alcohol, a possible decomposition product of DDAIP, were used as standard enhancers for comparison.

Percutaneous permeation of basic compounds through shed snake skin as a model membrane.

Relationships between the in-vitro permeability of basic compounds through shed snake skin as a suitable model membrane for human stratum corneum and their physiochemical properties were investigated. Compounds with low pKa values were selected to compare the permeabilities of non-ionized forms of the compounds. Steady-state penetration was achieved immediately without a lag time for all compounds.

Effects of transdermal penetration enhancers on the permeability of shed snakeskin.

The effects of Azone and lauryl alcohol on the permeability of shed snakeskin were examined. Permeability of a variety of compounds through shed snakeskin was increased after Azone or lauryl alcohol pretreatment but the magnitude of the enhancement varied depending on the lipophilicity and the molecular size of the permeant. It was found that the shed snakeskin became more permeable after Azone or lauryl alcohol pretreatment, with a greater permeability increase for more hydrophilic and larger-molecular size permeants.

Dodecyl N,N-dimethylamino acetate and azone enhance drug penetration across human, snake, and rabbit skin.

The effectiveness of the penetration enhancers, dodecyl N,N-dimethylamino acetate (DDAA) and Azone, on pretreated human epidermis for the permeation of model drugs, indomethacin, 5-fluorouracil, and propranolol-HCl, was studied in in vitro diffusion cells. Snakeskin (Elaphe obsoleta) and rabbit pinna skin were compared as possible models for human skin. The drug concentrations were analyzed by HPLC.

In vitro percutaneous transport of sodium diclofenac and diclofenac from oleaginous vehicle.

The penetration enhancement of sodium diclofenac and diclofenac by alcohols with various alkyl chains (C8 to C14) was evaluated by the steady state flux of diclofenac through rat abdominal skin. Decanol showed the greatest effect in this series. A more remarkable enhancing effect of the alcohols was observed in sodium diclofenac than in diclofenac.

In vitro transport of sodium diclofenac across rat abdominal skin: effect of selection of oleaginous component and the addition of alcohols to the vehicle.

The in vitro percutaneous transport of sodium diclofenac from various oil vehicles was examined using rat abdominal skin as a model skin membrane. The overall transport of diclofenac through the skin from the oleaginous vehicles was very poor because of a poor solubility of sodium diclofenac in nonpolar oils. To increase the solubility and the permeability of sodium diclofenac, ethanol and n-octanol were added to each oil (designated as the formulated vehicles).

A method to predict the percutaneous permeability of various compounds: shed snake skin as a model membrane.

Penetration of various compounds through shed snake skin was measured in vitro to examine the effect of lipophilicity and molecular size of a compound on permeability through this model membrane. The permeabilities were found to be controlled by the lipophilicity and the molecular size of the permeant. The smaller and the more lipophilic the compound, the greater the permeability.

Use of shed snake skin as a model membrane for in vitro percutaneous penetration studies: comparison with human skin.

The potential usefulness of shed snake skin as a model membrane for transdermal research was examined. There are similarities between shed snake skin and human stratum corneum in terms of structure, composition, lipid content, water permeability, etc. The permeability of various compounds and the contribution of several functional groups to the permeability were also found to be similar between shed snake skin and human skin.

Combined effect of alcohol and urea on the in vitro transport of indomethacin across rat dorsal skin.

An aqueous gel, prepared with hydrogenated soya phospholipid, increased the in vitro transport of indomethacin across rat dorsal skin. The addition of various alkanols further accelerated the transport, with an increasing effect as the chain length of the alkanol increased. The addition of urea alone did not significantly affect the transport of indomethacin.