Degradation Kinetics of BMP 777, an Elastase Inhibitor

Purpose. The objective was to evaluate the degradation profile of the elastase inhibitor DMP 777 and lay the foundation for formulation development. Methods. The pK_a was determined by potentiometric titration in mixed-aqueous solvents. The degradation kinetics were studied as a function of pH, buffer concentration, ionic strength, methanol concentration and temperature using a stability-indicating HPLC assay. The degradation products were identified by LC-MS, NMR, and by comparison with authentic samples. Results. The pK_a for the protonated piperazine nitrogen was estimated to be 7.04. The pH-rate profile is described by specific acid-, water-, and specific base-catalyzed pathways. The pH of maximum stability is in the range of 4 to 4.5 where water is the principal catalyst in the reaction. Buffer catalysis, primary salt effects and medium effects were observed. The proposed mechanism for acid catalyzed degradation is the rarely observed A_AL1 which involves alkyl-nitrogen heterolysis. The driving force for the reaction appears to lie in the stability of the benzylic carbocation. The proposed mechanism for base catalyzed degradation is B_AC2 which involves -lactam ring opening. The -lactam ring of DMP 777, a monolactam, appears to be as reactive as that in benzylpenicillin in the k _OH controlled region where a similar mechanism of hydrolysis should be operative. A contributing factor to this increased reactivity may lie in the reduced basicity of the -lactam nitrogen making it a good leaving group. Conclusions. The degradation profile indicates that development of a solution dosage form of DMP 777 with adequate shelf-life stability at room temperature is feasible.     

Stability of Alkoxycarbonylamidine Prodrugs

Purpose. Alkoxycarbonylamidine prodrug modification was used to mask the positively-charged amidine moiety of an Arg-Gly-Asp pepti-domimetic and enhance oral bioavailability. The aqueous stability of ethoxycarbonylamidine (EGA), ethanethiocarbonylamidine (ETCA) and phenoxycarbonylamidine (PCA) prodrugs was examined. Methods. Degradation was followed by RP-HPLC and rate constants were determined from a degradation scheme defined by product analysis. Results. EGA gave a pH of maximum stability at pH 7 and was independent of pH below pH 4. A novel degradation pathway of EGA, conversion to ethoxycarbonyl- aminocarbonyl, was observed below pH 7. The relative rates below pH 7 were ECA~ETCAa of the conjugate acid of the substituted amidino group. Base-catalyzed cleavage of EGA to yield the amidine derivative gave the relative rates ECAa of the leaving groups. Conclusions. The observed rate constants at all pHs were small enough that only 5-30% (depending on the substituent) undesirable degradation is predicted during transit time of the gut. The spontaneous post-absorptive conversion to the amidine drugs at neutral pH is predicted to be 6x greater for the PCA than the EGA prodrugs.     

Isothermal and Nonisothermal Decomposition of Thymopentin and Its Analogs in Aqueous Solution

Purpose. The degradation kinetics of thymopentin (RKDVY) and its analogs (RKDVW and RPDVY) in aqueous solution was studied by isothermal and nonisothermal methods. Methods. The isothermal decomposition of thymopentin and its analogs was investigated as a function of pH (2–10), temperature (37, 57, and 80°C) and ionic strength ( = 0.02 to 1). Nonisothermal decomposition studies were performed using a linear temperature programmer. The temperature increasing rate was set to 0.25°C per hour and the temperature interval varied from 40 to 88°C. Results. The decomposition of thymopentin and its analogs followed first order kinetics. The dependence of the rate constant on temperature followed a linear Arrhenius plot. This indicated that the degradation mechanism of thymopentin and its analogs might be the same within the temperature range studied. The energies of activation were found to be in close agreement for the isothermal and nonisothermal studies, suggesting that the nonisothermal studies may save considerable amount of time in the early stages of drug development. The logK-pH profile of thymopentin suggests that maximum stability is achieved in the pH range of 6–8. Conclusions. These results indicate that the nonisothermal methodology provides an attractive alternative to isothermal methods, as it requires a much lower amount of both material and time, to determine the peptide stability and to estimate the shelf-life for peptide pharmaceutical preparations.     

Solubilization and Stabilization of a Benzylpenicillin Chemical Delivery System by 2-Hydroxypropyl-β-cyclodextrin

A dihydropyridine pyridinium salt redox carrier-based chemical delivery system for benzylpenicillin (1) was complexed with 2-hydroxypropyl--cyclodextrin (HPCD). The solubility of the lipophilic 1, which is incompatible with aqueous formulations, was dramatically increased and showed a linear dependency on the HPCD concentration. The degree of incorporation was 20 mg of 1 per g of complex. The stability study of 1 in various pH buffers indicated the base-catalyzed hydrolysis of the acyloxyalkyl linkage and the hydration of the 5,6 double bond of the dihydropyridine as the main degradation processes. The overall loss of 1, which follows first-order kinetics, was not influenced by changes in ionic strength and elimination of oxygen from the reaction medium. The HPCD complex of 1, which has a stability constant of 720–940 M ^–1, stabilized the chemical delivery system. The influence of the temperature on the stability of 1 is also discussed.     

Glycine Crystallization During Freezing: The Effects of Salt Form,pH, and Ionic Strength

Purpose. The purpose of the study is to characterize glycine crystallization during freezing of aqueous solutions as a function of the glycine salt form (i.e., neutral glycine, glycine hydrochloride, and sodium glycinate), pH, and ionic strength. Methods. Crystallization was studied by thermal analysis, microscopy, x-ray diffraction, and pulsed Fourier transform nmr spectroscopy. Results. A solution of neutral glycine with no additives undergoes rapid secondary crystallization during freezing, forming the polymorph, with a eutectic melting temperature of –3.4°C. Glycine hydrochloride solutions undergo secondary crystallization relatively slowly, and the eutectic melting temperature is –28°C. Sodium glycinate crystallizes from frozen solution at an intermediate rate, forming a eutectic mixture with a melting temperature of –17.8°C. Where secondary crystallization does not occur rapidly, a complex glass transition is observed in the –70° to – 85°C temperature range in the DSC thermograms of all systems studied. Rates of secondary crystallization and the type of crystal formed are influenced by solution pH relative the the pKs of glycine, and also by the change in ionic strength caused by adjustment of pH. Increased ionic strength significantly slows the crystallization of neutral glycine and promotes formation of the polymorph. Thermal treatment or extended holding times during the freezing process may be necessary in order to promote secondary crystallization and prevent collapse during freeze drying. Conclusions. The results underscore the importance of recognizing that seemingly minor changes in formulation conditions can have profound effects on the physical chemistry of freezing and freeze drying.     

Interleukin-1 Receptor (IL-1R) Liquid Formulation Development Using Differential Scanning Calorimetry

Purpose. To elucidate the solution conditions that confer stability of aqueous IL-1R using differential scanning calorimetry (DSC). Methods. Optimal pH conditions were determined by monitoring degradation products encountered during accelerated studies (at elevated temperatures) using SDS-PAGE. At the pH optimum, DSC screened for excipients that enhanced thermal stability by shifting the Tm to higher values. Using SEC the relationship between thermal unfolding and stability was investigated by considering if lower Tm's in the presence of preservatives correlated with degradation products at 37°C over time. The degree of aggregation relative to that of a control determined the level of stability achieved. Results. Circular dichroism (CD) measurements confirmed molecular modeling studies showing IL-1R to be about 39% -sheet. Two major transitions characterized the DSC data with Tm's observed near 47°C and 66°C. Among 21 excipients screened, NaCl exhibited the greatest stabilizing influences based on shifting the low temperature transition to 53°C. The low temperature transition was later found to comprise two transitions, yielding a total of three melting transitions for IL-1R. High Tm's arising from the presence of preservatives correlated with the order of stability (i.e., 0.065% phenol > 0.1% m-Cresol > 0.9% benzyl alcohol). Conclusions. The three melting transitions are consistent in origin with the cooperative unfolding of three unique immunoglobulin-like domains of IL-1R. Optimal stability was achieved in 20 mM sodium citrate at pH 6 with sufficient NaCl to attain the tonicity of human serum. A correlation between the predicted ranking of stability and the extent of aggregation was demonstrated using DSC.     

Reduced UV-Induced Degradation of Doxorubicin Encapsulated in Polyethyleneglycol-Coated Liposomes

Purpose. The aim of this study was to investigate the stability of doxorubicin encapsulated in polyethyleneglycol-coated liposomes (Doxil) under UV-A light. Methods. High performance liquid chromatography and a fluorimetric method were used to quantify doxorubicin in bulk solution and doxorubicin in Doxil formulation. Results. The photodegradation of Doxil was significantly lower in comparison to the photodegradation of the free drug and showed no concentration dependency at the measured concentration range of 5–50 g/ml. During and after UV-A irradiation, there was no leakage of the drug from liposomes to the medium. After induced leakage of doxorubicin from the liposomes by the ionophore nigericin, the degradation kinetics of Doxil were identical to that of free doxorubicin. Conclusions. High intraliposomal doxorubicin concentration and intraliposomal acidic pH are the two critical factors that protect DXR in Doxil from UV-A degradation.     

Development of a Stable Freeze-dried Formulation of Recombinant Human Interleukin-1 Receptor Antagonist

Purpose. A formulation of recombinant human interleukin-1 receptor antagonist (rhIL-1ra) was developed that provided both acute protection during lyophilization and storage stability in the dried solid. Methods. The formulation was optimized by monitoring the impact of excipients on protein degradation which was analyzed by turbidimetry and cation-exchange HPLC. Results. The most appropriate pH was 6.5. Sodium citrate buffer provided better stability than sodium phosphate buffer. Glycine was selected as a bulking agent because the greatest protein stability was noted when this bulking agent was used in combination with an amorphous protein stabilizer. Among the amorphous stabilizers tested, sucrose protected rhIL-lra best in the presence of glycine. When the protein was freeze-dried in the presence of an inadequate mass ratio of sucrose/protein (< 0.3), the rate of degradation of rhIL-lra increased. For a formulation containing 100 mg/ml of rhIL-lra, increasing the sucrose/protein mass ratio to 0.3 greatly increased storage stability. The moisture content of the dried solid affected the storage stability to a minor degree. Three different stoppers obtained from the WEST Company did not affect the stability of rhIL-lra. Conclusions. An optimized formulation could be reconstituted without precipitation after 14 months at 30 or 50°C. At 30°C, there was no loss of native protein due to deamidation, and only a 4% loss at 50°C. These results indicated that the optimized formulation could be stored at ambient temperatures for long periods, without damage to the protein.     

Stabilized polymeric micelles by electrostatic interactions for drug delivery system

Methoxy poly(ethylene glycol)-block-oligo(l-aspartic acid)-block-poly(-caprolactone) with four aspartic acid groups was synthesized with a molecular weight and M_w/M_n of 8930 and 1.22. Polymeric micelles were formed by dialysis and stabilized by electrostatic interactions between the carboxylic acid groups and calcium cations. The critical micelle concentration of mPEG–Asp–PCL was determined to be 0.078 mg/mL. At 0.02 mg/mL, the dissociation of micelles without ionic stabilization formed an opaque, phase-separated solution, while the stabilized micelles under the same conditions showed structural stability through ionic stabilization. The paclitaxel-loading and efficiency were 8.7% and 47.6%, respectively, and the drug loading increased the mean diameter from 73.0 nm to 87 nm, which was increased further to 96 nm after ionic fixation. Rapid releases of approximately 65% of the encapsulated paclitaxel from a non-stabilized micelle and 45% from a stabilized micelle were observed in the first 24 h at pH 7.4 in a PBS solution containing 0.1 wt% Tween 80. The stabilized micelles then showed a sustained, slow release pattern over a couple of weeks, while the profile from the non-stabilized micelles reached a plateau at approximately 75% after 50 h. The enhanced micelle stability independent of concentration through ionic stabilization opens a way for preparing long circulating delivery systems encapsulating water-insoluble drugs.     

Chemical Stability of Esters of Nicotinic Acid Intended for Pulmonary Administration by Liquid Ventilation

Purpose. It has been suggested that fluorocarbon liquid may be a unique vehicle for the delivery of drugs directly to the acutely injured lung. A prodrug approach was used as a means of enhancing the solubility of a model drug (nicotinic acid) in the fluorocarbon. The solubility, the chemical stability of the putative prodrugs, and the sensitivity to enzymatic hydrolysis was investigated. Methods. The solubility of each nicotinic acid ester was determined in buffer as a function of pH and in perflubron. The octanol/buffer partition coefficient was determined at pH 7.4. The chemical stability of the putative prodrugs was determined as a function of pH, temperature, buffer content, and ionic strength. In addition, sensitivity of the esters to enzymatic degradation was evaluated. Results. Compared with nicotinic acid, the solubility in perflubron of the esters was significantly enhanced. In aqueous buffers, the esters exhibited pseudo-first order degradation kinetics, with both acid and base catalyzed loss. Studies of the fluorobutyl ester indicate quantitative loss of the putative prodrug and release of the parent nicotinic acid. Porcine esterase accelerated the loss of fluorobutyl ester by a factor of over 200 compared with chemical hydrolysis at pH 7.4. Conclusions. The properties of the fluorinated esters suggest that they may be suitable candidates for further testing as possible prodrugs of nicotinic acid based upon higher solubility in perflubron, rapid release of the parent drug after simple hydrolysis, and sensitivity to the presence of a model esterase enzyme.     

Stability of Gonadorelin and Triptorelin in Aqueous Solution

The influence of pH, temperature, various buffer species at different concentrations, and ionic strength on the stability of gonadorelin and triptorelin in aqueous solution has been studied using stability-indicating high-performance liquid chromatographic methods. The degradation behavior of both peptides is similar. The maximum stability of both peptides was shown to be at an approximate pH of 5.0. Acetate has the most favorable effect on stability, while phosphate causes higher degradation. Varying the concentration of acetate buffer does not affect the degradation behavior of the peptides. A higher phosphate concentration in buffer solutions causes higher degradation, however. The ionic strength of buffer solutions has no significant influence on stability. Solutions of gonadorelin and triptorelin, respectively, buffered with acetate (0.1 M, pH 5.0) with 3% (w/v) mannitol as an additive show a predicted t _90% of 9.0 years and 7.7 years at 20°C, respectively.     

Effect of pH on the micellar properties of amphiphilic drugs in aqueous solution

The influence of pH on the micellar properties of several amphiphilic drugs under conditions of constant ionic strength has been investigated. No significant effect of pH on the critical micelle concentration or micellar size of chlorpromazine hydrochloride was noted over a pH range well below the pK_a. The micellar properties of opipramol, thiopropazate, flupenthixol, clopenthixol, and trifluoperazine, which contain a piperazine moiety showed considerable pH dependence. The concentration dependence of the pK_a in these micellar systems was taken into consideration in the selection of pH values representative of complete protonation of either one or both of the piperazine N atoms. A lower aggregation number and higher critical micelle concentration was observed at a low pH corresponding to complete protonation of both charge centres. Mepyramine maleate exhibited a non-micellar mode of association at pH 5·5 which could be described by a stepwise association model in which association constants, K_N, increased sequentially with aggregation number, N, according to the relationship, K_N = K(N ? 1)/N where K = 31·3 dm³ mol^?1. No significant association could be detected at pH 2 when the pyridine ring N was fully protonated.     

Attenuation of Kupffer Cell Activation in Cold-Preserved Livers After Pretreatment of Rats with Methylprednisolone or Its Macromolecular Prodrug

Purpose. Activation of hepatic Kupffer cells (KCs) during organ preservation and subsequent reperfusion causes release of proinflammatory mediators and is responsible, at least in part, for rejection of transplanted livers. Our hypothesis was that donor pretreatment, before liver harvest, with methylprednisolone (MP) or its dextran prodrug (DMP) would reduce KC activation. Methods. Adult donor rats were administered a single 5-mg/kg (MP equivalent) IV dose of MP or DMP or saline 2 h before liver harvest. The livers were then stored in University of Wisconsin solution for 24, 48, or 96 h (n = 4/treatment/time). A recirculating perfusion model was used to study, for 180 min, the release of KC activation markers, tumor necrosis factor (TNF)- and acid phosphatase, and other biochemical indices from the cold-preserved livers. Results. Cold ischemia-reperfusion resulted in release of substantial levels of TNF- in untreated groups. Pretreatment of rats with MP or DMP caused a significant (p < 0.0001) reduction in TNF- AUC in the perfusate, with no significant differences between MP and DMP. The maximum inhibitory effect of MP (77.5 ± 10.2%) was observed after 48 h of preservation, whereas DMP showed maximal inhibition of TNF- AUC at both 24 (74.5 ± 15.8%) and 48 (74.8 ± 12.6%) h of preservation. Similarly, both MP and DMP resulted in a significant (p < 0.0004) decrease in acid phosphatase levels of cold-preserved livers. However, neither pretreatment had any substantial effect on the levels of other biochemical markers. Conclusions. Both MP and DMP pretreatments decreased the release of TNF- and acid phosphatase from livers subjected to cold ischemia preservation. Therefore, pretreatment of liver donors with MP or its prodrug decreases KC activation by cold ischemia-reperfusion.     

The Stability of Insulin in Crystalline and Amorphous Solids: Observation of Greater Stability for the Amorphous Form

Purpose. Generalizations based upon behavior of small molecules have established that a crystalline solid is generally much more stable toward chemical degradation than is the amorphous solid. This study examines the validity of this generalization for proteins using biosynthetic human insulin as the model protein. Methods. Amorphous insulin was prepared by freeze drying the supernate from a suspension of zinc insulin crystals adjusted to pH 7.1. Storage stability at 25°C and 40°C were compared for the freeze dried material, the dried suspended crystals, and the starting batch of crystals. Samples were equilibrated at selected relative humidities between zero and 75% to obtain samples at various water contents. Assays for dimer formation were performed by size exclusion HPLC and assays for deamidated product were carried out by reverse phase HPLC. Degradation was found to be linear in square root of time, and the slopes from % degradation vs. square root of time were used to define the rate constants for degradation. Differential scanning calorimetry (DSC) and Fourier-transform infrared spectroscopy (FTIR) were used to characterize the state of the protein in the solids. Results. As expected based upon previous results, the primary degradation pathways involve deamidation at the Asn^A21 site and co-valent dimer formation, presumably involving the A-21 site. Contrary to expectations, amorphous insulin is far more stable than crystalline insulin under all conditions investigated. While increasing water content increases the rate of degradation of crystalline insulin, rate constants for degradation in the amorphous solid are essentially independent of water content up to the maximum water content studied (15%). Conclusions. Based upon the FTIR and DSC data, both crystalline and amorphous insulin retain some higher order structure when dried, but the secondary structure is significantly perturbed from that characteristic of the native solution state. However, neither DSC nor FTIR data provide a clear interpretation of the difference in stability between the amorphous and crystalline solids. The mechanism responsible for the superior stability of amorphous insulin remains obscure.     

A Central Composite Design to Investigate the Thermal Stabilization of Lysozyme

Purpose. The formulation and processing of protein drugs requires the stabilization of the native, biologically active structure. Our aim was to investigate the thermal stability of a model protein, lysozyme, in the presence of two model excipients, sucrose and hydroxypropyl--cyclodextrin (HP--CD). Methods. We used high sensitivity differential scanning calorimetry (HSDSC) in combination with a central composite design (CCD). As indicators of protein thermal stability, the measured responses were the unfolding transition temperature (T_m), the onset temperature of the denaturation (T₀), and the extrapolated onset temperature (T_o,e). Results. A highly significant (F probability <0.001) statistical model resulted from analysis of the data. The largest effect was due to pH (over the range 3.2-7.2), and the pH value that maximized T_m was 4.8. Several minor but significant effects were detected that were useful for mechanistic understanding. In particular, the effects of protein concentration and cyclodextrin concentration on T_m and T_o,e were found to be pH-dependent. This was indicative of the partially hydrophilic nature of protein-protein interactions and protein-cyclodextrin interactions, respectively. Conclusions. Response surface methodology (RSM) proved efficient for the modeling and optimization of lysozyme thermal stability as well as for the physical understanding of the protein-sugar-cyclodextrin system in aqueous solution.     

Phagocytosis and Phagosomal Fate of Surface-Modified Microparticles in Dendritic Cells and Macrophages

Purpose. We compared cationic, polyamine-coated microparticles (MPs) and anionic, protein-coated MPs with respect to their phagocytosis and phagosomal fate in dendritic cells (DCs) and macrophages (M). Methods. Polystyrene MPs were surface modified by covalent coupling with fluorescein isothiocyanate-labeled polyamines or proteins. Phagocytosis of MP and the pH of their intracellular microenvironment was assessed in human-derived DCs and M in a fluorescence plate reader. Visualization of MP phagocytosis in DCs was performed by transmission electron microscopy. Results. Phagocytosis of bovine serum albumin-coated MPs was low with significant differences between DC and M, whereas phagocytosis of IgG-coated MPs was significantly enhanced in both cell types. Phagocytosis of both particle types resulted in an acidified phagosomal microenvironment (pH 4.6-5.1). In contrast, cationic, polyamine-coated MPs were equally phagocytosed by DCs and M to a high extent and showed lower degrees of acidification (pH 6.0-6.8) in the phagosomal microenvironment. Transmission electron microscopy examination demonstrated all phagocytosed particles to be surrounded by a phagosomal membrane, which was more tightly apposed to the surface of cationic MPs and more loosely to bovine serum albumin-coated MPs. Conclusion. Phagocytosis of cationic, polyamine-coated MPs is suggested to lead to diminished phagosomal acidification. Thus, cationic MP are potential carriers that may display beneficial features for the intracellular delivery of immunomodulating therapeutics and their protection against lysosomal degradation.     

The Effect of Benzyl Alcohol on Recombinant Human Interferon-γ

Purpose. The goal of this study was to investigate the conformational change and aggregation of recombinant human interferon-gamma (rhIFN-) as a result of interaction between benzyl alcohol and the protein. The effects of buffer concentration, buffer species, ionic strength, rhIFN- and benzyl alcohol concentrations on the dynamics of the interaction in liquid formulations were also examined. Methods. The effect of benzyl alcohol on the secondary and tertiary structure of rhIFN- in succinate and acetate buffers was studied using far-UV and near-UV circular dichroism spectrophotometry, respectively. Dynamic light scattering was employed to detect aggregate formation due to the interaction of benzyl alcohol with rhIFN-. Results. The addition of benzyl alcohol at 0.9% (w/v) in various liquid rhIFN- formulations induced changes in circular dichroism (CD) spectra of the protein in the near-UV region, while the CD spectra in the far-UV region remained unaltered. There were gradual decreases in ellipticity with time throughout the near-UV CD spectra. The decreases in near-UV ellipticity induced by benzyl alcohol were accompanied by the formation of high molecular weight aggregates as measured by dynamic light scattering. Loss in near-UV ellipticity was accelerated at lower protein concentration and by increasing buffer or benzyl alcohol concentration. It was also faster in succinate than in acetate buffer. Formulation ionic strength did not affect the CD spectral changes in both the near- and far-UV regions. Conclusions. Interaction between benzyl alcohol and rhIFN- is formulation dependent. Protein concentration, buffer species, buffer concentration, and preservative concentration play a significant role in determining the extent of the interaction and consequently the stability of the product.     

Effect of Cyclodextrin Charge on Complexation of Neutral and Charged Substrates: Comparison of (SBE)7M-β-CD to HP-β-CD

Purpose. To understand the role of charge in substrate/cyclodextrin complexation by comparing the binding of neutral and charged substrates to a neutral cyclodextrin, such as hydroxypropyl –CD (HP––CD) with 3.5 degrees of substitution, and an anionically charged cyclodextrin, such as sulfobutyl ether –CD ((SBE)_7M––CD) with 6.8 degrees of substitution. Method. HP––CD and (SBE)_7M––CD were evaluated in their ability to form inclusion complexes with neutral compounds, as well as to cationic and anionic substrates in their charged and uncharged forms. The complexation constants (K_c) were determined via a UV spectrophotometric technique, by monitoring the change in substrate absorbance upon incremental addition of a concentrated cyclodextrin solution. The role of electrostatic interaction was probed by observing K_c as a function of solution ionic strength. Results. Neutral molecules displayed a stronger interaction with (SBE)_7M––CD compared to HP––CD. In those cases where the guest possessed a charge (positive or negative), HP––CD/substrate complexes exhibited a decrease in complexation strength (2 to 31 times lower) compared to the neutral forms of the same substrate. The same was true (but to a larger extent, 41 times lower) for negatively charged molecules binding to (SBE)_7M––CD due to charge–charge repulsion. However, positively charged molecules interacting with the negatively charged (SBE)_7M––CD displayed a similar binding capability as their neutral counterpart, due to charge–charge attraction. Further evaluation through manipulation of solution ionic strength revealed strong electrostatic interactions between substrate and cyclodextrin charges. In addition, the studies suggested that on average two sulfonates out of seven may be involved in forming ionic attraction or repulsion effects with the positive charges on prazosin and papaverine, or negative charges of ionized naproxen and warfarin. Conclusions. Presence of charge on the cyclodextrin structure provides an additional site of interaction compared to neutral cyclodextrins, which may be modified using solution ionic strength.     

Theory and Computer Programs for Calculating Solution pH,Buffer Formula,and Buffer Capacity for Multiple Component System at a Given Ionic Strength and Temperature

Purpose. A theory and computer programs running on Microsoft^® Excel for Windows for calculation of solution pH, buffer formula, and buffer capacity at a given ionic strength and temperature were developed. The theory does not limit the category of buffer components, the number of buffer components, or the number of ionizations for each buffer component. The usefulness of the programs was examined. Methods. The formulas for 7 single component buffer solutions and 2 multiple component buffer solutions composed of citrate, phosphate, Tris, borate, and glycine were calculated. The solution pH values were measured at 25, 40, 55 and 70°C for comparison with the calculated pH values. Results. Of the 108 predictions made, 96 were of pH values within ± 0.1 pH unit of the measured values, at temperatures ranging from 25°C to 70°C and at ionic strengths ranging from 0.1 M to 0.5 M. Conclusions. These programs will be useful for identifying appropriate buffer solutions at various temperatures and/or ionic strengths.     

Response Surface Method: A Novel Strategy to Optimize lontophoretic Transdermal Delivery of Thyrotropin-releasing Hormone

Purpose. To maximize the iontophoretic transdermal delivery rate of thyrotropin-releasing hormone (TRH) facilitated by periodically monophase-pulsed current across excised skin. Methods. The pH of the buffer, the ionic strength in the solution, the frequency of the periodically monophase-pulsed current and the current on/off ratio were chosen as the key variables. A response surface method was applied to optimize the transdermal delivery rate of TRH under different operational conditions. Results. The optimum operating conditions were achieved via experimentation based on the response surface method by systematically adjusting the pH of the buffer, the ionic strength in the solution, the current amplitude, frequency and the active temporal ratio of the pulsed current. The rate of permeation of TRH crossing the skin during iontophoresis varied from two to ten-fold, depending on operating conditions. Conclusions. Only a few steps, two in this work, were needed to reach the optimal. The response surface near the region of the maximal point was thoroughly described with a quadratic function. A maximal transdermal rate of permeation of TRH, 103.2 µg h^–1 cm^–2, was obtained when the donor solution was at pH = 7.0, ionic strength = 0.037, and with a periodically monophase-pulsed current iontophoresis with duty cycle = 75%. The effect of pulse frequency was not statistically significant.