Increase in ceramide level after application of various sizes of sphingomyelin liposomes to a cultured human skin model.

Sphingomyelin-based liposomes (SPM-L) that were sized (or not) by extrusion through a filter with pores of 100, 200, or 400 nm were applied to a three-dimensional cultured human skin model in order to evaluate which size of SPM-L was most effective at increasing its ceramide level. The diameters of the SPM-L in PBS were 102.7, 181.0, 224.0, and 380.1 nm. The diameters of the liposomes in the culture medium were 117.5, 199.2, 242.1, and 749.8 nm. The diameter of the small liposomes (<200 nm in diameter) did not change much, at least for 7 days. SPM-L in saline or culture medium were applied to the basal layer side or stratum corneum side of the cultured skin model, and ceramide II, III, V, and VI were then extracted from it. The extracted ceramide molecules were separated by HPTLC, and the concentration of each type of ceramide was quantified using a densitometer. When the small SPM-L (110 or 190 nm in diameter) were applied to the basal layer side, the levels of ceramide III and V were increased. When they were applied to the stratum corneum side, the levels of ceramide II, III, V, and VI were significantly increased compared to those of the PBS group, especially after the application of the small SPM-L (110 nm in diameter). Thus, the application of small SPM-L was useful for increasing the ceramide II, III, V, and VI levels of a cultured human skin model.

Kinetic analysis on the in vitro cytotoxicity using Living Skin Equivalent for ranking the toxic potential of dermal irritants.

The extent of cytotoxicity injured by several skin irritants was kinetically measured and analyzed in a three-dimensional cultured human skin model, Living Skin Equivalent-002 (LSE). Colorimetric thiazoyl blue (MTT) conversion assay was selected as a cytotoxicity assay, and olive oil (OO), lactic acid (LA), Triton X-100 (TX) and sodium lauryl sulfate (SLS) were evaluated as model irritants. OO had almost no effect on the viability of LSE. When the other irritants were applied on the full-thickness LSE, two first-ordered decreasing phases, initial slow and following rapid phases, were found in the viability of LSE. LA and TX showed a bigger difference between the slow and rapid rates than SLS to show an inflection. The inflection time point from the slow to rapid rate was dependent on the kind and concentration of irritants applied. The higher the concentration of irritants applied, the more rapid the inflection point was observed. When LA and SLS were applied on the stratum corneum-stripped LSE, on the other hand, viability was mono-exponentially decreased with time. LA, TX and SLS probably decrease the barrier function of the stratum corneum to increase the rate of cytotoxicity during the irritant application. Interestingly, the rate of cytotoxicity on the stripped skin was similar to the late rapid rate on the full-thickness skin in LA not in SLS. These results suggest that cytotoxicity of skin irritants on the full-thickness LSE can be represented by two first-order kinetics, and that the skin irritation rate is closely related by the barrier function of skin as well as the application concentration and intrinsic toxicity of irritants.

Estimation of absorption rate of alpha-human atrial natriuretic peptide from the plasma profile and diuretic effect after intranasal administration to rats.

The absorption rate of alpha-human atrial natriuretic peptide (alpha-hANP) after intranasal (i.n.) administration to rats was estimated from the plasma profile and pharmacological effect (diuretic effect) using a pharmacokinetic (PK) model and a PK-pharmacodynamic (PD) model involving data obtained after intravenous (i.v.) bolus injection. The plasma concentrations of alpha-hANP after i.v. administration at different doses were fitted to a two-compartment PK model with zero-order excretion and input of endogenous alpha-rat atrial natriuretic peptide (alpha-rANP) and two elimination processes represented by Michaelis-Menten and first-order kinetics. However, the saturable process was ignored at low doses. The plasma concentrations after low doses via the i.n. route could also be expressed by this model, but with first-order absorption, so that an absorption rate constant was calculated using a deconvolution method. In addition, the diuretic effect plotted against the i.v. dose was represented by the Hill equation and showed an anti-clockwise hysteresis loop versus the plasma concentration. These results suggest that the diuretic effect could be estimated by a PK-PD model having an 'effect' compartment or a homeostatic system. Such a PK-PD model accurately expressed the diuretic effect of alpha-hANP at all doses after i.v. and i.n. administrations. The resulting absorption rate constant calculated using the PK-PD model agreed closely with that obtained by the PK model alone. The absorption rate and simulated diuretic effect suggest that, for i.n. administration of alpha-hANP, a higher absorption rate constant causes a more potent diuretic effect (a dramatic effect over the early period), whereas greater bioavailability is associated with a better hypotensive effect (sustained effect).

Effect of poly-L-arginine on the nasal absorption of FITC-dextran of different molecular weights and recombinant human granulocyte colony-stimulating factor (rhG-CSF) in rats.

The effect of poly-L-arginine (poly-L-Arg) on the in vivo nasal absorption of FITC-dextrans with a mean molecular weight ranging from 4.3 to 167 kDa and recombinant human granulocyte colony-stimulating factor (rhG-CSF) in rats were studied. When FITC-dextrans were co-administered intranasally with 1.0 w/v% poly-L-Args of different molecular weight (MW, ca. 45.5 and 92 kDa, poly-L-Arg (50) and poly-L-Arg (100)), the bioavailability (F(infinity)) increased markedly compared with that after administration of FITC-dextran alone. However, the F(infinity) decreased exponentially with the increasing molecular weight of FITC-dextrans. There was no significant difference between the enhanced nasal absorption of FITC-dextrans achieved by the co-administration of poly-L-Arg (50) and poly-L-Arg (100). Moreover, the relationship between the F(infinity) and the molecular weight of FITC-dextrans indicated that the molecular weight of protein drugs, which exhibited efficient absorption with poly-L-Arg, was about 20 kDa, when the lower limit of bioavailability for developing a potent transnasal delivery system was assumed to be about 10%. Indeed, the nasal absorption of rhG-CSF, which has a molecular weight of 18.8 kDa, was also increased after co-administration of 1.0 w/v% poly-L-Arg (50) and the F(infinity) was about 11%. It seems likely that poly-L-Arg can be used to provide adequate nasal absorption of various protein drugs which have a molecular weight of about 20 kDa, thereby allowing the successful development of a variety of transnasal drug delivery systems.

Potential usefulness of solubility index for prediction of the skin permeation rate of 5-ISMN from pressure-sensitive adhesive tape.

Skin permeation of 5-ISMN from pressure sensitive adhesive (PSA) tape was evaluated using thermodynamic activity of the drug in PSA. Three acrylic adhesives (Gelva 737, Gelva 1430 and Gelva 1753) were used as PSA. Since the drug activity in PSA is difficult to determine, however, a solubility index was defined. Several PSA tapes containing different amounts of 5-ISMN were prepared, and heat of fusion at the dissolution of 5-ISMN in each PSA was determined by DSC. No exothermic peak was found when the drug concentration was less than the solubility in PSA, whereas the heat of fusion increased proportionally with amount of solid drug in the PSA when the drug concentration was above the solubility. The bending point in the profile of heat of fusion versus 5-ISMN content in PSA was defined as the solubility index. In vitro skin permeation was determined using excised hairless rat skin from 5-ISMN-saturated PSA tapes. The obtained skin permeation of the drug decreased with increases in the solubility index. These profiles were confirmed by a theoretical approach using the differential equation corresponding to Fick's second law of diffusion. These results suggested that the solubility index can be utilized for prediction of the skin permeability of drugs from PSA tape.

Improved nasal absorption of drugs using poly-L-arginine: effects of concentration and molecular weight of poly-L-arginine on the nasal absorption of fluorescein isothiocyanate-dextran in rats.

The effects of the concentration and molecular weight of poly-L-arginine (poly-L-Arg) on the in vivo nasal absorption of fluorescein isothiocyanate-labeled dextran (MW, 4 kDa, FD-4) in rats were studied. When poly-L-Arg with a range of different molecular weights (MW, 8.9, 45.5 and 92.0 kDa) was applied intranasally at various concentrations, the bioavailability (F(0-9 h)) of FD-4 increased with the increasing concentration of poly-L-Arg. The enhanced absorption was also dependent on the molar concentration, in that the poly-L-Arg with a higher molecular weight increased F(0-9 h) at a lower molar concentration. In addition, for each applied concentration, the poly-L-Arg exhibited a molecular weight-dependence as far as the enhancement of FD-4 absorption was concerned. On the other hand, the maximum absorption rate (MAR) of FD-4, calculated by means of a deconvolution method, tended to reach a maximum plateau level at a lower applied concentration for the poly-L-Arg with the highest molecular weight, but this plateau level was almost the same for poly-L-Arg with molecular weights of 45.5 and 92.0 kDa. Moreover, the simulated absorption profiles of FD-4 indicate that the degree of enhancement (the level of MAR and the subsequent reduction in the absorption rate) was dependent on the molecular weight of poly-L-Arg, while the effect of poly-L-Arg was maintained for a longer period, depending on the applied concentration, although the MAR was relatively similar. These results indicate that the molecular weight of poly-L-Arg appears to affect both the enhancing efficiency (absorption rate) and the time-frame of this enhancing effect, whereas the concentrations of each poly-L-Arg system applied only have an effect on the time-frame. These effects may also be associated with the charge density of a poly-L-Arg molecule.

Electric field analysis on the improved skin concentration of benzoate by electroporation.

The objective in the present study was to understand the relationship between the increased skin concentration of benzoate as a model drug after topical application of its sodium salt and the electric field intensity produced in the skin barrier, the stratum corneum, by electroporation. A piece of excised abdominal hairless rat skin was set in a Franz type diffusion cell, and 0.5% sodium benzoate and physiological saline were applied to the stratum corneum and dermis sides, respectively. Two needle electrodes made of Ag were connected to an electrical power source, which produced exponentially decaying pulses. The electrodes were placed on the skin surface with a distance of 0.5 cm between both electrodes. After the 4 h passive permeation experiment, an electrical pulse was applied to the rat skin at 300 V every minute for 10 min. The skin was then removed from the diffusion cell, and the amounts of benzoate in different positions of the skin specimen were measured. Field intensity generated in the stratum corneum by electroporation was determined by a finite element method using a computer program. The amounts of benzoate at different sites in the skin were almost proportional to the mean field intensity in the corresponding stratum corneum. These results suggested that the enhancing effect of electroporation can be evaluated by the field intensity more directly than the application voltage.

[Pharmaceutical evaluation of fast-disintegrant tablet containing nicorandil-loaded particles].

AIM: To improve the bioavailability and taste of fast-disintegrating tablet (FD tablet) containing nicorandil-loaded particles. METHODS: A FD tablet containing nicorandil-loaded particles with 1%-4% croscarmellose sodium in addition of D-mannitol and lactose (9:1) was prepared and the dissolution and absorption characteristics were examined, in comparison with FD tablet and commercial tablets of nicorandil. In vivo absorption of nicorandil from FD tablet was evaluated in beagle dogs. RESULTS: The disintegration time of FD tablets containing 1% croscarmellose sodium with 6 mm and 10 mm in diameter were about 12 and 23 seconds, respectively. When nicorandil-loaded particles consist of myristyl alcohol and stearyl alcohol were put in FD tablet, nicorandil release from FD tablet continued until 6 h while nicorandil release from Sigmart and FD tablet containing nicorandil crystals finished within 5 min. In vivo absorption of nicorandil from Sigmart and FD tablet containing nicorandil crystals was very similar after oral administration in beagle dogs and no statistic difference in AUC, Tmax, Cmax was observed between these tablets. However pharmacokinetics parameters of nicorandil after oral administration of FD tablet containing nicorandil-loaded particles showed that nicorandil was delivered into the body at a suitable absorption rate with similar AUC, delayed Tmax and lower Cmax. CONCLUSION: The reports suggest that the modification of properties of myristyl alcohol and stearyl alcohol released from the drug-loaded particles system would lead to more acceptable bioavailability of the system. However, The formulation of particles and may have a masking effect against the bitter taste and irritation of the drug.

[Transdermal microparticle delivery by a supersonic-Helios gun system].

AIM: To investigate the effect of particle size and high speed flow of helium gas on the systemic absorption of indomethacin using a needle-less injection system. METHODS: Poly-L-lactic acid microspheres containing indomethacin was prepared by the o/w solvent evaporation technique. After anesthetizing the male hairless rat, microspheres filled in the tube cartridge was accelerated by a stream of helium gas at various velocity in the Helios gun system, and then was introduced to the abdominal skin. RESULTS: Introduction of indomethacin to the hairless rat skin was proportionally increased with enhancing the helium pressure (supersonic flow). Bioavailability and Cmax were also dependent on the helium pressure. CONCLUSION: This method can be used to deliver the powered drug and/or microparticulate systems into the skin tissues and the systemic circulation.

Synergistic effects of iontophoresis and jet injector pretreatment on the in-vitro skin permeation of diclofenac and angiotensin II.

A non-needle syringe (jet injector) was utilized to increase skin permeation of drugs by iontophoresis. Briefly, physiological saline was initially flushed by the injector to make a pore in the stratum corneum of excised hairless rat skin, and the iontophoretic skin permeation of two model compounds, sodium diclofenac and angiotensin II, was followed using a 2-chamber diffusion cell. Constant voltage and constant current iontophoresis treatments were evaluated. Pretreatment using the jet injector alone resulted in about 13- and 22-fold increases in the steady-state flux of diclofenac and angiotensin II, respectively, through the skin, compared with non-treated controls. Jet injector pretreatment with constant voltage iontophoresis further enhanced skin permeation of diclofenac and angiotensin II, and the enhancement was also greater than that by constant voltage iontophoresis alone. Thus, a synergistic effect was observed. The ratio of enhancement was greater compared with the control. Jet injector pretreatment with constant current iontophoresis, however, did not always yield higher skin permeation of the drugs than injector pretreatment alone, although the lag time was shortened. The difference in the enhancement between the constant voltage- and constant current iontophoresis can be explained by the electric current through the excised skin. Constant current iontophoresis after a short period of constant voltage iontophoresis with multiple jet injector pretreatments may be the best way to increase drug permeability while preventing severe skin damage.

Effect of l-menthol-ethanol-water system on the systemic absorption of flurbiprofen after repeated topical applications in rabbits.

The effect of the l-menthol-ethanol-water system (MEW system), a skin penetration enhancer, on the systemic absorption of flurbiprofen (FP) after repeated topical applications was investigated. FP (1%) gel containing ethanol (25%) and l-menthol (3%) as penetration enhancers was applied to rabbit dorsal skin and the in vivo absorption rate of FP was compared with the in vitro penetration rate through excised skin. In vivo absorption rate of FP was initially high and decreased with time to a value approximately equal to the in vitro rate. The remaining FP in the gel 6 h after the application was 60% of the initial loading and the systemic bioavailability over the 6 h application was about 10%, suggesting that the rest (30%) had accumulated in the skin tissues. The gel was applied for 6 h on the same site or on a new site after the first 6 h-application to learn the effect of repeated applications on FP absorption. The maximum FP concentration after the second application on the virgin skin was slightly higher than that after the first application, as expected in a typical pharmacokinetic process. In contrast, the same site application induced remarkably lower plasma concentration and area under the curve (AUC). A drug-free gel was also utilized to evaluate the effects of the enhancer system. Pretreatment of the drug-free gel on the same site also decreased the FP absorption, whereas post-treatment increased the plasma level of FP, in spite of the removal of the drug gel. These phenomena could be explained by ethanol in the MEW system acting a local irritant and a drug carrier.

Analysis of skin permeation-enhancing mechanism of iontophoresis using hydrodynamic pore theory.

The effects of constant DC iontophoresis (0-1.5 mA/0.966 cm(2)) on the permeation of three hydrophilic compounds, antipyrine (ANP, M.W. 188.23), sucrose (SR, M.W. 342.30) and 1-kestose (KT, M.W. 506.73), through excised hairless rat skin were evaluated using hydrodynamic pore theory. The electro-osmotic flow caused by iontophoresis was measured using deuterium oxide (D(2)O). The penetration-enhancing mechanism of iontophoresis was found to increase solvent flow through electro-osmosis and pore enlargement and/or new pore production in the skin barrier, together with enhancement of electrochemical potential difference across the skin. These effects were closely related to the strength of the current applied. The electro-osmotic flow of D(2)O (J(D(2)O)) greatly enhanced the skin permeation clearance of all hydrophilic penetrants (CL(drug)). Pore production was classified into reversible and irreversible processes, which resulted from lower (0-0.5 mA/0.966 cm(2)) and higher (0.5-1. 5 mA/0.966 cm(2)) currents, respectively. Thus, the enhancing effects of iontophoresis on skin permeation of nonionic hydrophilic compounds can be explained by increase in pore size and higher solvent flow.

Transdermal delivery of the potent analgesic dihydroetorphine: kinetic analysis of skin permeation and analgesic effect in the hairless rat.

Dihydroetorphine is an extraordinarily strong opioid analgesic. To assess its effectiveness after topical application in hairless rats we have examined the kinetic analysis of skin permeation through excised skin and the in-vitro reservoir effect of skin, and have investigated the predictability of plasma concentration and analgesic effect following in-vivo transdermal application. Dihydroetorphine was moderately permeable from an aqueous suspension through excised hairless rat skin. Dihydroetorphine flux from drug-dispersed pressure-sensitive adhesive tape was threefold that from the applied aqueous suspension. The fluxes through the abdominal and the dorsal skin during tape application fitted the Fickian diffusion equation well after the tape was removed peeling off the outer layer of the stratum corneum. The relationship between the plasma concentration and the analgesic effect was examined for four different rates of infusion of dihydroetorphine. A non-linear pharmacokinetic disposition was observed. Following abdominal (0.28 cm2, 20 microg) and dorsal (0.50 cm2, 35 microg) applications of the dihydroetorphine tape, plasma concentration (0.2-0.8 ng mL(-1)) and analgesic effect were maintained at a suitable level, for more than 8 h, until removal of the tape. These profiles were predictable using the combined equation for percutaneous absorption, disposition and the analgesic effect, but the analgesic effect was slightly lower than the predicted value. The results show that it was possible to control the plasma concentration and the analgesic effect of dihydroetorphine by topical application of the analgesic using pressure-sensitive adhesive tape in the hairless rat. It was possible to predict the result using mathematical modelling.

Evaluation of bile acids and fusidate derivative as nasal absorption enhancers using an electrophysiological technique.

The present study was carried out to investigate the reversibility of the action of two nasal absorption enhancers, bile acids and fusidate derivative, on nasal membrane resistance. The nasal mucosa was isolated from rabbit nasal septum and mounted in a Ussing-type chamber to allow the monitoring of the membrane resistance and flux of fluorescein isothiocyanate-labeled dextran (FD10, M.W. 9400). Membrane resistance was reduced by 46% following treatment with 0.5% (w/v) sodium taurodihydrofusidate (STDHF) for 10 min and then gradually returned to the control level after being wash. The resistance was restored to 76% of the control level following a 30 min treatment with 0.5% (w/v) STDHF. However, there was no recovery of resistance following treatment with 0.5% (w/v) STDHF for 120 min or 1% (w/v) STDHF for 10in. Concurrently, FD10 transport was enhanced while membrane resistance was reduced. Treatment with 0.5% (w/v) sodium deoxycholate (DC) for more than 10 min showed no reversible action and marked FD10 transport enhancement, whereas a 10-30 min treatment with 0.5% (w/v) sodium glycocholate (GC) or sodium taurocholate (TC) resulted in the rapid recovery of membrane resistance without any enhancement of FD10 permeation. STDHF transport across the nasal mucosa was approximately 2-fold faster than that of DC, GC, and TC. The accumulation of STDHF in the nasal mucosa was 2-fold lower than that of DC and 1.7-fold higher than that of GC and TC after a 30 min treatment. The rank order of hydrophobicity determined by reverse-phase HPLC was: DC>STDHF>GC>TC. These results suggest that the reduction in membrane resistance and its reversibility appear to be due to a balance between the accumulation and clearance of STDHF.

Analysis of skin disposition of flurbiprofen after topical application in hairless rats.

Cutaneous disposition of topically applied flurbiprofen (FP) was evaluated using a new in situ experimental model in hairless rats. A disk-shaped agar gel (3.85 cm in diameter and 0.5 cm in thickness) was subcutaneously implanted in the abdominal region of rats as a drug receptor, and a drug donor cell was subsequently placed above this agar gel. No significant pharmacokinetic modification of FP was observed as a result of this experimental procedure. A bolus injection and a constant intravenous infusion of FP were applied to the rats, followed by an analysis of FP levels in the plasma and agar gels. Using these results, the clearance rate of FP from the systemic circulation to the gel could be calculated. FP (1% gel formulation, 1.0 g/3.14 cm(2)) was then topically applied to the skin of these rats. From these experiments, the amount of FP that migrated from the formulation to the systemic circulation and the amount of FP that migrated directly to the agar gel across the skin, over 10 h, were separately evaluated to be 235.4 and 2.0 microg, respectively. Thus, most of the FP was absorbed into the systemic circulation. The effect of endogeneous vasoactive compounds and penetration enhancers on the FP disposition within skin was also determined. Epinephrine and bradykinin were used as vasoactive compounds that were entrapped in agar gel, and an isopropyl myristate system (IPM system) and a l-menthol-ethanol-glycerin-water system (MEGW system) were used as enhancers in the formulation. Epinephrine enhanced the direct delivery of FP into the agar gel to more than ten times its former level, in spite of the fact that it had no effect on systemic delivery. Bradykinin strengthened systemic delivery slightly, without changing the quantity of FP in the gel. IPM increased only the systemic delivery of FP, as was the case with bradykinin, whereas the MEGW system markedly increased both the blood concentration and the quantity of FP in the gel (13 and 200 times, respectively). This technique has proven to be an effective tool for the quantitative evaluation of cutaneous disposition of a topically applied drug.

Simultaneous transport and metabolism of ethyl nicotinate in hairless rat skin after its topical application: the effect of enzyme distribution in skin.

An in vitro permeation study of ethyl nicotinate (EN) was carried out using excised hairless rat skin, and simultaneous skin transport and metabolism of the drug were kinetically followed. Fairly good steady-state fluxes of EN and its metabolite nicotinic acid (NA) through the skin were obtained after a short lag time for all the concentrations of EN applied. These steady-state fluxes were not proportional to the initial donor concentration of EN: EN and NA curves were concave and convex, respectively, which suggests that metabolic saturation from EN to NA takes place in the viable skin at higher EN application. Further permeation studies of EN or NA were then carried out on full-thickness skin or stripped skin with an esterase inhibitor to measure their permeation parameters, such as partition coefficient of EN from the donor solution to the stratum corneum and diffusion coefficients of EN and NA in the stratum corneum and the viable epidermis and dermis. Separately, enzymatic parameters (Michaelis constant K(m) and maximum metabolism rate V(max)) were obtained from the production rate of NA from different concentrations of EN in the skin homogenate. The obtained permeation and enzymatic parameters were then introduced to differential equations showing Fick's second law of diffusion in the stratum corneum and the law with Michaelis-Menten metabolism in the viable epidermis and dermis. The calculated steady-state fluxes of EN and NA by the equations were very close to the obtained data. We then measured the esterase distribution in skin microphotographically using fluorescein-5-isothiocyanate diacetate. A higher enzyme concentration was observed in the epidermal cells and near hair follicles than in the dermis. Simulation studies using the even and the partial enzyme distribution models suggested that no significant difference between the models was observed in the skin permeations of EN and NA, whereas concentration-distance profiles of EN and NA were very different. This finding suggests that the total amount of enzyme in skin which converts EN to NA is a determinant of the metabolic rate of EN in skin. The present approach is a useful tool for analyzing simultaneous transport and metabolism of many drugs, especially those showing Michaelis-Menten type-metabolic saturation in skin.

Relationship between tyrosinase inhibitory action and oxidation-reduction potential of cosmetic whitening ingredients and phenol derivatives.

The oxidation-reduction potentials of cosmetic raw materials, showing tyrosinase inhibitory action, and phenolic compounds structurally similar to L-tyrosine were determined by cyclic voltammetry. The voltammograms obtained could be classified into 4 patterns (patterns 1-4). Pattern 1, characterized by oxidation and reduction peaks as a pair, was observed with catechol, hydroquinone or phenol, and pattern 2 exhibiting another oxidation peak in addition to oxidation and reduction peaks as a pair was found with arbutin, kojic acid, resorcinol, methyl p-hydroxybenzoate and L-tyrosine as the substrate of tyrosinase. Pattern 3 with an independent oxidation peak only was expressed by L-ascorbic acid, and pattern 4 with a reduction peak only at high potentials, by hinokitiol. The tyrosinase inhibitory activity of these compounds was also evaluated using the 50% inhibitory concentration (IC50) and the inhibition constant (Ki) as parameters. Hinokitiol, classified as pattern 4, showed the highest inhibitory activity (lowest IC50 and Ki). Hydroquinone showing the second highest activity belonged to pattern 1, which also included compounds showing no inhibition of tyrosinase activity. The inhibitory activity of compounds exhibiting pattern 2 was relatively low with Ki values being in the order of 10(-4) M. Although there was no consistent relationship between oxidation-reduction potentials and tyrosinase inhibitory action, the voltammetry data can be used as an additional index to establish the relationship between the structure and the tyrosine inhibitory activity.

Skin disposition of drugs after topical application in hairless rats.

Drug fraction transported from a topical formulation on skin to subcutaneous tissues or muscles is dependent on the physicochemical properties of the entrapped drug. Cutaneous disposition of model drugs, antipyrine (ANP), lidocaine (LC) and piroxicam (PXC) as well as flurbiprofen (FP) was thus evaluated in hairless rats in which an agar gel disc was subcutaneously inserted into the abdominal region as a drug receptor and a drug donor cell was placed above it. Time courses of plasma level and agar gel amount were measured after topical application of 50% ANP, 3% LC, 1% PXC and 1% FP in hydroxypropylcellulose gel. Percutaneous absorption clearance of unionized form, CLab* was proportional to true octanol/water distribution coefficient and the order of FP > PXC > LC > ANP, suggesting that skin permeation of the drug was determined mainly by its distribution from the formulation to the skin barrier. PXC, however, had a relatively low flux compared to the other three drugs, probably due to its high molecular weight and melting point. Migration clearance of unionized form from systemic circulation to the subcutaneous agar gel, CLg* was also influenced by the lipophilicity of drugs. On the other hand, fraction from the formulation to the systemic circulation was in the order of PXC > FP > ANP > LC. This fraction was much higher than the direct migration fraction from the formulation to the subcutaneous agar gel. Factors determining for these fractions are still unclear. A drug having a low lipophilicity and a low protein binding, however, had a tendency to have a great targeting ability to the subcutaneous agar gel. In addition, most of the drug in the agar gel was contributed by the direct flow from formulation, not from the systemic circulation. The present in situ experimental method is a useful tool to evaluate skin disposition of drugs. Detailed understanding of the skin disposition of drugs from several formulations will enable the findings of a good drug and formulation candidates.

Screening of cationic compounds as an absorption enhancer for nasal drug delivery.

Several cationic compounds were screened as potential nasal absorption enhancers to increase intranasal absorption of a model drug, fluorescein isothiocyanate labeled dextran (MW 4.4 kDa, FD-4), without nasal membrane damage in rats. Their effects were compared with those of classical enhancers. Various cationic compounds (poly-L-arginines with different molecular weights (MW 8.9, 45.5 and 92.0 kDa, poly-L-Arg (10), (50) and (100), respectively), L-arginine (L-Arg), L-lysine (L-Lys), and cetylpyridinium chloride (CPCL) were evaluated. Of the cationic compounds, poly-L-Arg and CPCL greatly enhanced the intranasal absorption of FD-4, as did chitosan, a cationic polysaccharide which has been reported to show a great effect on the transnasal delivery of peptide and protein drugs. The enhancing intensity by poly-L-Arg was dependent on its molecular weight. Rank order of the enhancing ratio, calculated from the AUC ratio for the enhancer treatment against the untreatment, was 0.5% poly-L-Arg (100) congruent with0.5% sodium dodecylsulfate congruent with0.5% CPCL?0.5% poly-L-Arg (50)?0.5% sodium deoxycholate congruent with0.5% sodium taurodihydrofusidate?0.5% polyoxyethylene-9-lauryl ether congruent with0.5% lysophosphatidylcholine?0.5% chitosan congruent with0.5% poly-L-Arg (10)>/=10% L-Arg congruent with10% L-Lys?0.5% sodium glycocholate congruent with0.5% sodium taurocholate congruent with0.5% EDTA. Only the poly-L-Args represented almost the same degree of hemolysis of cationic compounds compared with pH 7.0 phosphate buffered saline in the rat erythrocyte lysis experiment. The enhancing ratio by classical enhancers correlated with leaching of protein, phospholipids and LDH from isolated rabbit nasal mucosa. CPCL also fell on the regression lines between the enhancing ratio and their degree of leaching from classical enhancers. In contrast, the enhancing intensities by poly-L-Arg (10), (50) and (100) were greatly shifted from the regression line: the amount of leaching was markedly low in spite of a great enhancement of FD-4 absorption. These findings suggest that of the assessed enhancers only the poly-L-Args enhance the transnasal delivery of high molecular substances without severe damage to the nasal mucosal membrane. Poly-L-Arg is therefore a promising candidate having a good balance between enhancing activity and safety for nasal peptide and protein delivery.

Optimum threshold values of 201Tl SPET in the determination of the left ventricular border and extent of myocardial infarction.

The aim of this study was to determine the optimum threshold value of the left ventricular border and the extent of myocardial infarction using quantitative 201Tl single photon emission tomography (SPET). We used the unfolded map method to determine the size of the left ventricle and the extent of myocardial infarction in 10 patients, because it has been shown to be superior to the conventional polar map method. The relative differences in the size of the left ventricle and extent of infarction between 201Tl SPET and post-mortem examination were calculated. The optimum threshold value was determined when the relative difference = 0%. There was an excellent correlation between scintigraphic and post-mortem left ventricular size at a threshold value of 53% (r = 0.91, P < 0.001); an excellent correlation was also observed between scintigraphic and post-mortem infarct size at a threshold value of 55% (r = 0.93, P < 0.03). The optimum threshold value in determining left ventricular size using 201Tl SPET is 53% and that in determining infarct size is 55%.