Quantitative determination of lattice fluoride effects on the solubility and crystallinity of carbonated apatites with incorporated fluoride.

The purpose of this study was to evaluate quantitatively the effects of fluoride on the solubility and crystallinity of carbonated apatites (CAPs) after its incorporation into the crystal lattice using the metastable equilibrium solubility (MES) distribution method. Fluoride-incorporated CAPs (F-CAPs) of two different carbonate levels (3 and 5%) and fluoride contents from 0 to 20,000 µg/g were synthesized. X-ray diffraction experiments and Rietveld analysis were conducted to obtain crystallite microstrain and unit cell parameters. Acetate buffer MES solution media were prepared at two solution fluoride concentrations (0.2 and 2.0 mg/l) and at two pHs (5.0 and 5.7). The unit cell a-axis values of the F-CAPs were found to decrease as the fluoride content increased, consistent with the fluoride being incorporated into the crystal lattice. The fluoride concentrations in the MES solution media were high enough to provide a 'swamping' effect such that the fluoride released from the F-CAPs during dissolution was minimal in changing the solution fluoride concentration. Employing the MES distribution superposition method, it was shown that the surface complex possessing the fluorapatite (FAP) stoichiometry [Ca10(PO4)6F2] accounted for the MES distribution behavior of all experiments. In addition, the mean pIFAP [the value of -log(aCa(10)aPO4(6)aF(2)) calculated from the ionic activity product based on FAP stoichiometry of the MES dissolution media in which 50% of the F-CAPs had dissolved] correlated well with the crystallite microstrain parameters of the F-CAPs. The incorporated fluoride in the F-CAPs showed only modest effects on F-CAP crystallinity and solubility.

Noninvasive measurement of phenylalanine by iontophoretic extraction in patients with phenylketonuria.

Phenylketonuria is an autosomal recessive disorder characterized by elevated concentrations of phenylalanine. Elevated phenylalanine concentrations can impair intellectual functions and the disease is treated with a lifelong diet and frequent monitoring of plasma phenylalanine concentrations. Previous in vitro studies have demonstrated the feasibility of iontophoretically enhanced transdermal transport of phenylalanine. Here we evaluate the feasibility of transdermal iontophoretic extraction of phenylalanine in vivo. Phenylalanine was iontophoretically extracted from the skin of healthy volunteers and of patients with phenylketonuria for up to 6 h and concentrations were compared with those measured in plasma. The amount of phenylalanine iontophoretically extracted from the skin declined over time, suggesting contribution of phenylalanine from the skin in the initial extraction. Phenylalanine iontophoretically extracted from skin correlated with plasma phenylalanine levels at plasma levels above 300 micromol/L. This correlation supports the feasibility of iontophoretic phenylalanine extraction for monitoring phenylketonuria.

A comparative study of the metastable equilibrium solubility behavior of high-crystallinity and low-crystallinity carbonated apatites using pH and solution strontium as independent variables.

UNLABELLED: Using solution strontium and pH as independent variables, the metastable equilibrium solubility (MES) behavior of two carbonated apatite (CAP) samples has been examined, a high-crystallinity CAP (properties expected to be similar to dental enamel) and a low-crystallinity CAP (properties expected to be similar to bone mineral). CAP samples were prepared by precipitation/digestion: (CAP A: high-crystallinity, 1.3 wt% CO3, synthesized at 85 degrees C; CAP B: low-crystallinity, 6.4 wt% CO3, synthesized at 50 degrees C). Baseline MES distributions were determined in a series of 0.1 M acetate buffers containing only calcium and phosphate (no strontium) over a broad range of solution conditions. To assess the influence of strontium, MES profiles were determined in a similar fashion with 20, 40, 60, and 80% of the solution calcium being replaced on an equal molar basis by solution strontium. To determine the correct function governing CAP dissolution, ion activity products (IAPs) were calculated from the compositions of buffer solutions based on the hydroxyapatite template (Ca(10-n)Sr(n)(PO4)6(OH)2 (n = 0-10)) and the calcium/hydroxide deficient hydroxyapatite template (Ca(9-n)Sr(n)(HPO4)(PO4)5OH (n = 0-9)). FINDINGS: (a) for CAP A, at high solution strontium/calcium ratios, the MES profiles were essentially superimposable when the solution IAPs were calculated using the stoichiometry of Ca6Sr4(PO4)6(OH)2 and for CAP B by a stoichiometry of Ca7Sr2(HPO4)(PO4)5OH; (b) for CAP A, at low strontium/calcium ratios, the stoichiometry yielding MES data superpositioning was found to be that of hydroxyapatite and for CAP B, that of calcium/hydroxide deficient hydroxyapatite. When other stoichiometries were assumed, good superpositioning of the data was not possible.

Metastable equilibrium solubility behavior of carbonated apatite in the presence of solution strontium.

The purpose of this study was to use the concept of metastable equilibrium solubility (MES) to describe the anomalous solubility behavior of carbonated apatite (CAP) in the presence of solution strontium. A CAP sample (4.8 wt% CO(3), synthesized at 70 degrees C) was prepared by precipitation. Baseline MES distributions were determined in a series of 0.1 M acetate buffers containing only calcium and phosphate (no strontium) over a broad range of solution conditions. In order to assess the influence of strontium, MES profiles were then determined in a similar fashion with 20, 30, 40, 50, 60, 70, and 80% of the solution calcium being replaced on an equal molar basis by solution strontium. From the compositions of the equilibrating buffer solutions, ion activity products (IAPs) of the form Ca(10-n)Sr(n)(PO(4))(6)(OH)(2) (n = 0-10) were calculated in an attempt to determine the correct function governing the dissolution of the CAP preparation. The results demonstrate the following important findings: (a) at high solution strontium/calcium ratios (i.e., when 60% or more of the solution calcium was replaced by strontium), the MES profiles in all the experiments were found to be essentially superimposable when the solution IAPs were calculated using the stoichiometry of Ca(6)Sr(4)(PO(4))(6)(OH)(2), and (b), at low solution strontium/calcium ratios (i.e., when 40% or less of the solution calcium was replaced by strontium), the stoichiometry yielding MES data superpositioning was found to be that of hydroxyapatite. When other stoichiometries were assumed, good superpositioning of the data was not possible.

Mechanistic studies of the effect of hydroxypropyl-beta-cyclodextrin on in vitro transdermal permeation of corticosterone through hairless mouse skin.

Literature reports reveal that the issue of whether cyclodextrins may act as skin permeation enhancers has not been resolved. Accordingly, in vitro skin transport studies were conducted to address this question. Corticosterone (3H-CS and/or non-radiolabeled CS) was chosen as the model permeant for transport experiments with hairless mouse skin (HMS) and with a synthetic cellulose membrane of 500 molecular weight cut off (MWCO), the latter to help establish baseline behavior. Hydroxypropyl-beta-cyclodextrin (HPbetaCD) was selected as the representative cyclodextrin. The CS/HPbetaCD complexation constant was determined both from solubility data (saturation conditions) in phosphate buffered saline (PBS), pH 7.4 and with data obtained from PBS/silicone polymer partitioning experiments, the latter experiments permitting the determination of the complexation constant at low CS concentrations. These results were used in the calculations of the free CS concentrations in the donor chamber of the transport experiments. The CS transport experiments were conducted at CS solubility saturation and under supersaturation (resulting from autoclaving at 121 degrees C) conditions as well at very low (tracer level) concentrations. The effect of polyvinylpyrrolidone as a solution additive was also evaluated. The following were the key outcomes of this study. Contrary to literature reports, there was no evidence that HPbetaCD is an enhancer for CS transport through HMS. The CS permeability coefficient values obtained with HMS in all of the experiments were found to be the same within experimental error when calculated on the basis of the free CS concentration as the driving force for permeation. The constancy of the permeability coefficient in the presence and absence of HPbetaCD is interpreted to mean that, in these experiments, HPbetaCD did not alter the barrier properties of HMS stratum corneum to any significant extent nor did it enhance CS transport in any other manner such as by a carrier mechanism involving the aqueous boundary layer or by a carrier mechanism within the stratum corneum.

Effect of fluoride pretreatment on the solubility of synthetic carbonated apatite.

The purpose of this research was to address the following question: How is the solubility of fluoride-pretreated carbonated apatite (CAP) in aqueous acidic media related to the equilibrium solution fluoride and/or the CAP adsorbed fluoride levels? A CAP sample prepared by a precipitation method at 70 degrees C containing approximately 6% carbonate was fluoride-treated (F adsorption from neutral aqueous solutions) to yield a approximately 1000 ppm F CAP and a approximately 3300 ppm F CAP. Metastable equilibrium solubility distributions were determined in acetate buffers at pH 5.0. Solution fluoride, calcium, phosphate, and pH were determined from the equilibrated solutions. The equilibrium solution fluoride levels were extremely low, e.g., as low as approximately 0.10 ppb to approximately 0.30 ppb at 50% dissolved for the two CAP preparations. The approximately 3300 ppm F CAP yielded a lower solubility than the approximately 1000 ppm F CAP (shift in the mean pKHAP value of 1.5-2 units). This can be attributed to the lower solution F(-) for the sample containing approximately 1000 ppm fluoride compared with the approximately 3300 ppm fluoride-containing CAP. These important findings suggest that a fluoride treatment simply may provide an adsorption fluoride depot for subsequent release, providing a solution fluoride effect upon the CAP solubility and not necessarily any intrinsic alteration of the mineral solubility.

Effect of age on rat bone solubility and crystallinity.

It has been shown that biominerals such as dental enamel and bone demonstrate a non-thermodynamic equilibrium state following initial dissolution where no further mineral dissolution or precipitation occurs; this state is termed metastable equilibrium solubility (MES). Furthermore, these minerals are composed of a distribution of domains each with their own MES. Recent studies have also demonstrated a linear relationship between crystallinity and the mean MES of these minerals, with solubility decreasing with increasing crystallinity. This study investigates the effect of age on the MES and crystallinity of rat bone mineral. The bone mineral samples were prepared by protein extraction with a series of hydrazine and alcohol solutions. The MES distributions of the bone mineral were measured by exposure to 0.1 M acetate buffers containing a range of calculated amounts of calcium (Ca) and phosphate for 48 h. The amount of mineral dissolved in each solution was determined from Ca and phosphate analyses of the undissolved residue. The full width of the half maximum (FWHM) of the 002 reflection of the X-ray powder diffraction was used as an indicator of crystallinity. The MES of mineral from bone of rats of different ages (1-25 months) were compared. Results of this study indicate that (l) there is a difference in the mean MES of bone mineral from rats of different ages, with older bone mineral being less soluble and more crystalline than bone mineral from younger rats; (2) the nature of the solubility distribution changes from a narrow to a broader distribution with age; and (3) all of the bone samples demonstrated an inverse-linear correlation between crystallinity and mean MES value consistent with previous results obtained with synthetic apatites and dental enamel.

Influences of alkyl group chain length and polar head group on chemical skin permeation enhancement.

Previous investigations in our laboratory on the influence of the n-alkanols and the 1-alkyl-2-pyrrolidones as skin permeation enhancers for steroid molecules as permeants demonstrated that the enhancer potencies (based on aqueous concentration values) of these two homologous series were the same when compared at the same alkyl chain length; that is, the contribution of the hydroxyl group and that of the pyrrolidone group to enhancer potency were the same. The purpose of the present study was to further investigate what was believed to be a somewhat surprising finding, and two additional homologous series, the 1,2-alkanediols and N,N-dimethylalkanamides, were selected for study as enhancers. Corticosterone (CS) flux enhancement along the lipoidal pathway of hairless mouse skin stratum corneum was determined with 1,2-hexane-, 1,2-octane-, and 1,2-decanediol and with N,N-dimethylhexanamide, N,N- dimethylheptanamide, N,N-dimethyloctanamide, and N,N-dimethylnonanamide as enhancers. The enhancement factor (E) for the lipoidal pathway was calculated from the CS permeability coefficient and the CS solubility data over a 4 to 100 range of E values. Comparisons of the enhancer potencies of all four homologous series revealed that the enhancer potencies of all were very nearly the same when compared at equal alkyl group chain length. Moreover, the contribution of each of the polar head groups toward the enhancer potency was essentially constant, independent of the alkyl group chain length. It was reasoned that this outcome was either the result of the random selection of four polar head groups making the same contribution to enhancer potency or the result of these particular polar head groups not contributing to enhancer potency. To test the hypothesis that the former was more likely than the latter and that a suitable semipolar organic phase may mimic the microenvironment of the polar head group at the site of enhancer action, n-octanol-phosphate buffered saline (PBS) and n-hexane-PBS partition coefficients were determined for all the enhancers. The n-octanol-PBS partition coefficients for the enhancers, but not the n-hexane-PBS partition coefficients, were very nearly the same when compared at equal alkyl group chain lengths; this result supports the hypothesis that each of the four polar head groups likely contributes the same toward the enhancer potency and locates in the semipolar region of the hairless mouse skin stratum corneum lipid bilayers, which is well-approximated by water-saturated n-octanol.

Iontophoretic transport of oligonucleotides across human epidermal membrane: a study of the Nernst-Planck model.

The objective of this study was to investigate the transport behavior of a series of oligonucleotides with human epidermal membrane (HEM) and to examine the applicability of the modified NERNST-PLANCK model to transdermal iontophoresis of these macromolecules. Iontophoretic transport experiments were first carried out in a synthetic model membrane system (Nuclepore membranes) with a four-electrode potentiostat to examine the baseline modified NERNST-PLANCK model. The modified NERNST-PLANCK model derived from the Einstein relation and the Stokes-Einstein equation taken from previous work did not hold for the oligonucleotides. Results obtained in the Nuclepore studies were, however, consistent with predictions of the modified NERNST-PLANCK model using the experimentally determined electromobilities and diffusion coefficients. The electromobilities of the oligonucleotides (determined by capillary electrophoresis) were found to be more than a factor of two smaller than expected from the Einstein relation between electromobilities and diffusion coefficients (the latter determined in diffusion cell experiments). A correlation between these electromobilities and the theoretical electromobilities estimated by considering the effects of counterion binding and the effects of mobility reduction according to colloid theory was also observed. These results suggest that the modified NERNST-PLANCK model predictions are satisfactory only when the electromobilities and the effective molecular size of the oligonucleotides are known and are used directly to predict the iontophoretically enhanced transport. Results with the HEM experiments generally agreed with model predictions based on the experimental electromobilities. The oligonucleotide HEM flux data also suggest the existence of pores with effective pore radii greater than the effective radii estimated in previous studies with small molecular weight model permeants.

Quantification of pore induction in human epidermal membrane during iontophoresis: the importance of background electrolyte selection.

It has been shown that significant pore induction (electroporation) occurs in human epidermal membrane (HEM) during iontophoresis even at moderate applied voltages (1-10 V). Recent efforts in our laboratory have been aimed at quantifying HEM electroporation by examining the proportionality between flux enhancement due to electroporation and electrical conductance changes during iontophoresis. The specific purpose of the present study was to test the hypothesis that by matching the background electrolyte ion sizes with the permeant ion sizes, the flux enhancement due to electroporation can be quantified by the change in HEM electrical conductance. In this study, radiolabeled tetraethylammonium (TEA(+)), methylammonium (MA(+)), and mannitol were the permeants. Potassium chloride (KCl), tetraethylammonium bromide (TEAB), tetraethylammonium pivalate (TEAP), and sodium fluoride (NaF) were the background electrolytes. Iontophoresis experiments were carried out over an applied voltage range of 1 to 3 V. The experimental flux enhancement results were compared with the theoretical predictions from the Nernst-Planck model after corrections were made: (a) for HEM pore induction during iontophoresis based on electrical conductance changes and (b) for electroosmosis employing mannitol as the neutral probe permeant. In experiments where the ion sizes of the background electrolyte and permeant were closely matched (e.g., TEA(+) as the permeant and TEAP as the background electrolyte), there was excellent agreement between experimental results and theoretical predictions of the modified Nernst-Planck model, with only modest data scatter. When the electrolyte and permeant sizes were quite different (e.g., TEA(+)/KCl and MA(+)/TEAP), the experimental flux data were inconsistent with model predictions and there were large variations in the experimental results. The results of the present study illustrate that permeant flux enhancement can be predicted by the modified Nernst-Planck model even during moderate voltage iontophoresis when electroporation is operative.

Relationships involving metastable equilibrium solubility, surface complexes, and crystallite disorder with carbonated apatites.

Previous studies have shown that the metastable equilibrium solubility (MES) behavior of carbonated apatites (CAPs) may be described by a surface complex with the hydroxyapatite (HAP) stoichiometry in the absence of solution fluoride and by that with the fluorapatite (FAP) stoichiometry when appreciable solution fluoride is present. Studies have also shown that the magnitude of the MES is directly related to the crystallinity of the CAP. The aim of the present investigation was to examine the relationship between the CAP MES determined in the presence of solution fluoride and CAP crystallinity and to examine the effect of the change in the stoichiometry of the surface complex (from that of HAP to that of FAP) upon the relationship of the CAP MES to crystallinity. CAP samples were prepared by methods based on the precipitation of CAP from calcium phosphate solutions and the hydrolysis of dicalcium phosphate dihydrate in bicarbonate solutions. From X-ray diffraction experiments, the crystallite microstrain, and the full width at half maximum (FWHM) of the 002 reflection were determined for the CAPs. From CAP MES experiments conducted in the presence of solu tion fluoride, linear plots of the mean MES (i.e., mean pK(FAP)) values vs the crystallinity parameter (i.e., microstrain and FWHM) were obtained that yielded slopes that were essentially the same as those obtained in the absence of solution fluoride (i.e., mean pK(HAP) values vs crystallinity). This parallel finding suggests that the CAP crystallite disorder affects the energetics of the two surface complexes essentially to the same extent and provides new insight into the nature of CAP surface complexes.

Calcium-level responsive controlled drug delivery from implant dosage forms to treat osteoporosis in an animal model.

The effects of plasma calcium levels on estradiol release from a self-setting apatite bone cement containing 0.5% estradiol and on the bone mineral density (BMD) of ovariectomized rats were investigated. Apatite cement consisting of an equimolar mixture of tetracalcium phosphate, dicalcium phosphate dihydrate and 0.5% beta-estradiol was prepared. The in vitro release profiles from the cements in simulated body fluid containing 0, 5 and 10 mg/100 ml calcium indicated that estradiol release rate decreased with increasing calcium concentration in the dissolution medium. After subcutaneous implantation of the cement, in vivo estradiol release in diseased rats (ovariectomized rats on a low calcium diet) was significantly higher than that in normal rats. The diseased rats maintained a low calcium level during drug release. The bone mass of the recovery model rat was greater after the experiment than before. The results suggested that the severity of osteoporosis in this animals can be reduced by the implantation of this estradiol-loaded apatite cement.

Mechanistic aspects of iontophoresis in human epidermal membrane.

A large number of factors are involved in the movement of ions and molecules across human epidermal membrane (HEM) under the influence of an electric field. These factors and their interplay need to be understood if our knowledge of iontophoretic transport of drugs across HEM is to reach a point where physical models and strategies may be employed for useful quantitative predictions. In a typical in vitro experiment, the fully hydrated HEM is positioned between aqueous compartments of a two-chamber diffusion cell. When a low electric field is applied across the HEM under these conditions, the transport enhancement of ions in the pre-existing pores of the stratum corneum is the result of, (a) the direct interaction of the electric field with the charge of the ion in question, and (b) convective solvent flow (electroosmosis); in the case where the permeant is non-ionic under these circumstances, transport enhancement is by convective solvent flow only. At moderate-to-high voltage iontophoresis (> or = around 1.0 V applied across a single HEM), in addition to the direct field effect and convective solvent flow in the pre-existing pores, there can generally be a significant (e.g. 10- to 100-fold enhancement) contribution to transport enhancement arising from new pore induction (electroporation). Much of the recent work in our laboratory has been devoted to defining and quantifying HEM electroporation, and an especially difficult aspect has been that of dealing with the large HEM membrane-to-membrane variabilities with regard to, (a) the extent of new pore induction, and (b) the characteristics of the newly induced pores. Recently we discovered that the extent of relevant (i.e. permeant accessible) pore induction may be correlated to the change in HEM electrical conductance (and quantifiable) if an appropriate matching background electrolyte can be selected having ion sizes comparable to that of the permeant. For example, employing tetraethylammonium (TEA) pivalate (PIV) for which the ion sizes are approximately 3.5 A, but not KCl (ion sizes approximately 1.9 A), as the background electrolyte for TEA (as the permeant) gave very good results; in this example, the sizable contribution of pore induction to iontophoresis was quantitatively factored out from the total iontophoretic enhancement. Experiments with a large number of HEM samples gave good agreement with the Nernst-Planck (N-P) predictions of the direct field effect when TEA-PIV was used as the background electrolyte for TEA transport, but large variations (up to 300%) between N-P predictions and experimental results were observed with KCl as the background electrolyte. Another area of recent effort has been HEM pore size determinations, both at low voltages (i.e. for pre-existing pores) and at voltages where the newly induced pores dominate HEM permeability. The sizes of pre-existing pores of HEM have been determined with the hindered diffusion theory (using experimental fluxes of several probe permeants of different known molecular sizes) to be generally in the range, 10-20 A, by a number of investigators in our laboratory for a large number HEM samples. Deducing pore sizes of electric field induced pores under steady electroporation conditions has been a more challenging task. We succeeded recently in developing a novel method for 'passively' determining pore sizes (i.e. by passive diffusion with hindered diffusion theory) under steady electroporation conditions: by using low frequency (12.5 Hz) a.c. at 2-5 V. We have been able to sustain electroporation at a nearly constant state of electroporation long enough to carry out a set of 'passive' diffusion experiments with relatively good precision to obtain the sizes of the newly induced pores. Studies to date have revealed that the sizes of pores induced with 2-5 V are of the same order of magnitude as those of the pre-existing pores (i.e. 10-20 A). Finally, another research question of interest has been that of pore charge

Pore charge distribution considerations in human epidermal membrane electroosmosis.

The aim of this study was to assess the extent to which a model with pores having only net negative charges would adequately describe transdermal electroosmosis in human epidermal membrane (HEM) at neutral pH. Such information would enhance the predictive value of the modified Nernst-Planck model for transdermal iontophoresis, in addition to providing insights regarding the likelihood of significant pore charge distribution in HEM. Baseline results (the control) obtained from 0.1 to 0.4 V anodal and cathodal electroosmosis experiments with synthetic polycarbonate membranes (Nuclepore membranes), using radiolabeled urea and mannitol as the model permeants, demonstrated that such a membrane system can be modeled by the electrokinetic (electroosmosis) theory with the assumption of the pores possessing only negative charges. The studies with HEM were carried out at low voltage (/approximately = 1.0 V) where significant field-induced pore formation in HEM occurred. In both the low and high voltage studies, radiolabeled urea, mannitol, and water were employed as permeants in cathodal and anodal iontophoresis experiments. The results of the low voltage iontophoresis experiments suggest significant pore charge distribution in HEM (a significant deviation between the predictions from the single pore charge type assumption and the experimental data). Under the higher applied voltage conditions (>/approximately = 1.0 V), results from anodal and cathodal electroosmosis studies were consistent with the model in which the HEM has only pores that are net negatively charged.

Effect of plasma-calcium-level-responsive oestradiol release from apatitic bone cement on bone mineral density in ovariectomized rats.

The effects of plasma calcium levels on oestradiol release from apatite bone cement and on the bone mineral density of ovariectomized rats have been investigated. Apatite cement was prepared from an equimolar mixture of tetracalcium phosphate, dicalcium phosphate dihydrate and 0.5% beta-oestradiol bulk powder. After subcutaneous implantation of the cement, oestradiol release in diseased rats (ovariectomized rats on a low-calcium diet) was significantly higher than in normal rats. The drug levels of recovery-model rats (ovariectomized, but on a high-calcium diet) were significantly lower than those of the diseased rats. Calcium levels in diseased rats remained low during drug release but the plasma calcium levels of the recovery-model rats increased. The areas under the plasma calcium concentration-time curves (Ca-AUCs) for the recovery-model rats were higher than those for the diseased-model rats. The plasma oestradiol concentration AUCs and the Ca-AUCs were linearly related. The body weight of the recovery-model rats increased after five days, but that of the diseased-model rats did not. The bone mass of the recovery-model rats was greater after the experiment than before. The relationship between the bone mineral density and Ca-AUC of the diseased rats suggested that bone mineral density increased with increasing Ca-AUC. The results suggest that the severity of osteoporosis in this animal model is reduced by implantation of the oestradiol-loaded apatite cement.

Influence of the treatment protocol upon the in vivo efficacy of cidofovir (HPMPC) and of acyclovir (ACV) formulations in topical treatment of cutaneous HSV-1 infection in hairless mice.

In recent studies we found that the topical effectiveness of acyclovir (ACV) formulations was a single-valued function of C-the target site free drug concentration. The topical efficacy was the same when the therapy was initiated 0, 1, or 2 days after intracutaneous herpes simplex virus type-1 (HSV-1) inoculation in hairless mice. The purpose of the present study was to examine the hypothesis that the topical effectiveness of cidofovir (HPMPC) would not be a single valued function of C and that it would be dependent upon when the therapy was initiated relative to the time of viral infection. Formulations of HPMPC and ACV in 95% DMSO as a vehicle were used. Hairless mice intracutaneously infected with HSV-1 were used, and 20 microL of the test formulation was topically applied twice a day. In protocol A, the treatment was continued until the fourth day after virus inoculation, whereas in protocol B the treatment was terminated on the day of virus inoculation. Treatment was initiated on various days ranging from day -6 to day 4, and the lesions were scored on day 5. Treatment of ACV according to protocol A proved efficacious whether started as early as 6 days before virus inoculation or later, whereas the efficacy of ACV was annihilated if applied following protocol B. For HPMPC, on the other hand, the in vivo efficacies were found to be strongly dependent on how early the therapy was initiated, and significant efficacy was observed even when the treatment was terminated on the day of virus inoculation. This difference was attributed to the virus-independent intracellular phosphorylation of HPMPC and slow clearance of its metabolites from the cell. It was also noted that, similar to ACV, for HPMPC the topical efficacy is likely to be a function of C for a fixed protocol. However, unlike for ACV, for HPMPC the efficacy was not a single-valued function of C.

Relationships among carbonated apatite solubility, crystallite size, and microstrain parameters.

The use of the metastable equilibrium solubility (MES) concept to describe the solubility properties of carbonated apatites (CAPs) and human dental enamel (HE) has been well established in previous studies using a range of CAPs with varying carbonate contents and crystallinities. It was shown in these studies that the mean value of the CAP MES is directly related to the broadening parameter full width at half maximum (FWHM) of the 002 reflection of the X-ray diffraction profile. The apparent solubility of the CAPs increased monotonically with an increase in the broadening of the diffraction peaks, and when this peak broadening was taken into account, carbonate had no additional effect upon the MES. The broadening of the diffraction peaks has been used as an indicator of crystallinity, and is generally influenced by both crystallite size and microstrain. The purpose of the present study was to extract the crystallite size and microstrain parameters separately from the X-ray diffraction peaks and then to determine their relationships to the corresponding MES values. The samples studied were CAPs synthesized by precipitation from Ca(NO3)2 and NaH2PO4 solutions in carbonate containing media at temperatures of 95, 80, and 70 degrees C, and powdered HE. The crystallite size and microstrain parameters were determined simultaneously with the refinement of the structural parameters with the Rietveld method of whole-pattern-fitting structure-refinement. A modified pseudo-Voigt function was used to model the observed peak profiles. The MES distributions for the CAPs and HE were determined by a previously described method. The results of this study showed that the CAPs possessed an MES distribution and therefore provided further support that MES distribution is a common phenomenon, regardless of the method of CAP synthesis. The crystallite size decreased and the microstrain increased with increasing carbonate content and decreasing temperature of synthesis of the CAPs. A plot of the mean of the MES distribution versus the microstrain parameter showed that the apparent solubility of the CAPs and HE correlated very well with the microstrain parameter. On the other hand, a plot of the mean of the MES distribution versus the crystallite size parameter showed a poor correlation between MES and crystallite size. These findings support a view that microstrain, rather than crystallite size, is the dominant factor governing the effective solubility of the CAPs and dental enamel.

Pore induction in human epidermal membrane during low to moderate voltage iontophoresis: A study using AC iontophoresis.

The present study aimed to investigate new pore induction as a flux-enhancing mechanism in human epidermal membrane (HEM) with low to moderate voltage electric fields. The extent of pore induction and the effective pore sizes of these induced pores were to be assessed using a low frequency (12.5 Hz) low to moderate voltage (2. 0 to 4.0 V) square-wave alternating current (ac) "passive" permeation method (ac iontophoresis). This ac approach was to allow for inducing and sustaining a state of pore induction in HEM while permitting no significant transport enhancement via electroosmosis; thus, transport enhancement entirely due to new pore induction (enhanced passive permeation) was to be assessed without any contributions from electroosmosis. Good proportionality between the increase in HEM permeability and its electrical conductance was found with the "passive" transport data obtained during square-wave ac iontophoresis using urea as the model permeant. Typically, at 3.0 to 4.0 V, HEM conductance increases (and permeability increases) ranged from around 3- to 30-fold. These results appear to be the first direct evidence that new pore induction in HEM is a significant flux enhancing mechanism under moderate voltage conditions. The extents of pore induction in HEM under low frequency moderate voltage (2.0 to 3.0 V) ac, pulsed direct current (dc), and continuous dc were also compared. The extents of pore induction from square-wave ac and pulsed dc were generally of the same order of magnitude but somewhat less than that observed during continuous dc iontophoresis at the same applied voltage and duration, suggesting less extent of pore induction with reversing polarity or when a brief delay is provided between pulses to allow for membrane depolarization. The average effective pore sizes calculated for the induced pores from the experimental data with urea and mannitol as probe permeants and the hindered transport theory were 12 +/- 2 A, which are of the same order of magnitude as those of preexisting pores determined from conventional passive diffusion experiments.

Metastable equilibrium solubility behavior of bone mineral.

Previous studies have shown that carbonated apatites with a range of carbonate contents and crystallinities exhibit the phenomenon of metastable equilibrium solubility (MES) distributions. The purpose of the present study was to investigate the solubility behavior of bone mineral using the concepts of MES and MES distributions and, together with crystallinity and chemical composition data, examine the similarity of bone mineral to carbonated apatite (CAP). Bone samples were harvested from 1-, 5-, and 8-month-old rats. The organic components of the bone samples were removed by hydrazine deproteination. Carbonated apatite was synthesized by the hydrolysis of dicalcium phosphate dihydrate (DCPD) in a NaHCO3-containing media at 50 degrees C. The MES distributions of bone mineral and CAP were determined by equilibrating predetermined amounts of CAP or bone mineral in a series of 0.1 M acetate buffers containing calculated levels of calcium and phosphate and maintained at essentially constant pHs of 5.0, 5.3, 5.7, and 6.5. From the compositions of the equilibrating buffer solutions, ion activity products based upon the stoichiometries of octacalcium phosphate, hydroxyapatite, and carbonated apatite were calculated in an attempt to determine the function governing the dissolution of CAP and bone mineral. The results of this study demonstrated that the MES distribution phenomenon appeared to hold for bone mineral and that the changes in crystallinity of bone mineral with age correlated well with changes in the MES values. A CAP sample was prepared that was found to be an excellent synthetic prototype closely mimicking the physicochemical behavior of bone mineral from an 8-month-old rat. Another finding of this study was that the ion activity product function based upon the hydroxyapatite stoichiometry well described the MES results obtained with both CAP and bone mineral. The interpretation that a surface complex with hydroxyapatite stoichiometry governs the solubility behavior of bone mineral is, therefore, consistent with the experimental data. Other calcium phosphate stoichiometries for the surface complex showed systematic variations in the MES profiles when the pH of the equilibrating solution was varied.

Effect of geometrical cement size on in vitro and in vivo indomethacin release from self-setting apatite cement.

The relationship between in vitro and in vivo indomethacin (IMC) release from a self-setting bioactive apatite cement and cement size were investigated. Differently sized apatite cements (total weight, 500 mg); either 64 of the small size (2 mm diameter x 2 mm thickness), sixteen of the medium size (4 mm x 2 mm) or one of the large size (15 mm x 2 mm) were obtained from cement bulk powder containing tetracalcium phosphate, dicalcium phosphate dihydrate and hydroxyapatite. In vitro IMC release from the 1, 2 and 5% drug-loaded apatite cement systems in simulated body fluid (SBF) (pH 7.25) at 37 degrees C increased with increasing concentrations of IMC and with decreasing geometrical size of the cement. The plots of in vitro IMC release per unit area against the square root of time increased with increasing IMC concentrations, but not with decreasing geometrical size of the cement. After subcutaneous (s.c.) implantation of differently sized 1% IMC-loaded cements in male Wistar rats, the plasma IMC concentration and the area under the curve increased with decreasing cement diameter. The in vivo IMC release profiles of the cement were deconvoluted from the plasma IMC profiles after s.c. administration of IMC solution. The plots of in vivo IMC release per unit area against the square root of time suggested that the initial release from all 1% drug-loaded cements was very rapid, slowed after one day, but continued for over two weeks. The relationship between the in vitro release in SBF and the in vivo release in rats of IMC-loaded cements was linear.