Comparison of the conventional and immersing powdered crude drugs (IPCD) methods for color and extraction of quantitative indicator ingredients in the Kampo formula decoction of daiokanzoto.

AIM: The immersing powdered crude drugs (IPCD) method is a quick and simple method for preparing decoctions. Here, the conventional and IPCD methods were compared for the color and extraction of quantitative indicator ingredients in the daiokanzoto decoction solution, and the suitability of the IPCD method was assessed. METHODS: The color of decoction solutions was visually observed, and the Commission Internationale de L'éclairage (CIE) L*a*b*color parameters were measured using conventional and IPCD methods. The extracted amounts of sennoside A and glycyrrhizic acid, which are quantitative indicator ingredients of rhubarb and glycyrrhiza, respectively, were quantified. RESULTS: Using both methods, the decoction solution colors were strong for rhubarb alone and daiokanzoto but weak for glycyrrhiza alone. The color change of daiokanzoto was thought to be primarily caused by rhubarb alone. The L*a*b* values of the decoction solution determined by the IPCD method were comparable to those determined by the conventional method (60 min). Using the conventional method, sennoside A and glycyrrhizic acid were mostly extracted in 10 and 30 min, respectively. Using the IPCD method, both sennoside A and glycyrrhizic acid were fully extracted in 2 min. The IPCD method yielded significantly more sennoside A and glycyrrhizic acid (2 times and 1.5 times, respectively) than the conventional method (60 min). CONCLUSION: The IPCD method was found to be comparable to the conventional method in terms of the color, and using IPCD method, the same or greater amounts of quantitative indicator ingredients of crude drugs in the decoction of daiokanzoto compared to the conventional method. It was suggested that there are limitations to assessing the equivalence of decoctions from decoction color. The IPCD method may be a useful method although it is prudent to use the IPCD method for Kampo formula decoction in clinical practice with a certain degree of caution.

An attempt at administering a transdermal formulation of bisoprolol in patients with acute cardiac symptoms.

BACKGROUND: Bisono® is the world's first transdermal formulation of a bisoprolol, which is approved for the treatment of hypertension in Japan. We aimed to investigate the usefulness of this formulation in patients who were admitted to our hospital with cardiac symptoms suggestive of acute coronary syndrome or an acute exacerbation of heart failure. METHODS: This study involved a retrospective survey of medical records from September 1, 2017 to April 30, 2018 obtained from the Cardiovascular Center of Kyoto Katsura Hospital. The clinical data of patients on admission who had received a transdermal formula of bisoprolol (Bisono® tape) were retrieved; their blood pressure and heart rate data were analyzed in relation to the doses of Bisono® tape administered. RESULTS: Sixty-three patients received the Bisono® tape. Their final diagnoses included acute myocardial infarction, an exacerbation of heart failure, and atrial fibrillation. While there was no significant correlation observed between the administered doses of the drug and reduction in blood pressure achieved within 24 h after admission, there was a significant (p < 0.05) correlation between the doses of Bisono®tnd reduction in the heart rate within 24 h after admission (ΔHR0-24 h). Only one patient who received 8 mg of Bisono® exhibited temporal bradycardia (heart rate < 50 bpm). CONCLUSION: The transdermal formulation of bisoprolol may be useful for the early introduction of ß-blockers in patients admitted with cardiac symptoms associated with myocardial ischemia or heart failure. However, caution should be exercised because of the possible risk of hypotension.

A retrospective analysis of patient-specific factors on voriconazole clearance.

BACKGROUND: Voriconazole concentrations display a large variability, which cannot completely be explained by known factors. We investigated the relationships of voriconazole concentration with patient-specific variables and concomitant medication to identify clinical factors affecting voriconazole clearance. METHODS: A retrospective chart review of voriconazole trough concentration, laboratory data, and concomitant medication in patients was performed. The concentration/dose ratio (C/D-ratio) was assessed as a surrogate marker of total clearance by dividing voriconazole concentration by daily dose per kg of body weight. RESULTS: A total of 77 samples from 63 patients were obtained. In multiple linear regression analysis, increased C-reactive protein (CRP) level (p < 0.05) and decreased albumin (Alb) level (p < 0.05) were associated with significantly increased C/D-ratio of voriconazole, and coadministration with a glucocorticoid was associated with significantly (p < 0.05) decreased C/D-ratio of voriconazole (adjusted r (2) = 0.31). Regarding CRP and Alb, receiver operating characteristic curve analysis indicated that increased CRP level and decreased Alb level were significant predictors of toxic trough concentration of voriconazole. For CRP, area under the curve (AUC) and cutoff value were 0.71 (95 % confidence interval (CI), 0.57-0.86, p < 0.01) and 4.7 mg/dl, respectively. For Alb, AUC and cutoff value were 0.68 (95 % CI, 0.53-0.82, p < 0.05) and 2.7 g/dl, respectively. A significant difference was seen in voriconazole trough concentration between patients with hepatotoxicity and those without (5.69 µg/ml vs 3.0 µg/ml, p < 0.001). CONCLUSION: Coadministration of glucocorticoid and inflammation, reflected by elevated CRP level and hypoalbuminemia, are associated with voriconazole clearance. We propose that early measurement of voriconazole concentration before the plateau phase will lead to avoidance of a toxic voriconazole level in patients with elevated CRP level and hypoalbuminemia, although further studies are needed to confirm our findings.

Preparation and pharmaceutical evaluation of acetaminophen nano-fiber tablets: Application of a solvent-based electrospinning method for tableting.

In this study, we developed nano-fiber-based tablets with acetaminophen (AAP; LogPow=0.51) for controlled-release delivery systems and evaluated in vitro drug dissolution and in vivo pharmacokinetics in rats. Nano-fibers made from methacrylic acid copolymer S (MAC; EUDRAGIT S100) and containing AAP were prepared using a solvent-based electrospinning (ES) method. In vitro dissolution rate profiles of AAP showed tableting pressure-dependent decreases and pH-dependent increases. The results of tablet tracking by X-ray irradiation showed tablets based on MAC nano-fibers did not disintegrate in the upper intestinal lumen and had the properties of a long-term-acting tablet. In addition, the in vitro release profiles of AAP from nano-fiber tablets prepared by dissolving MAC with AAP (NFT), nano-fiber tablets prepared by adsorbing AAP to drug-free MAC nano-fibers (NFTadso), and tablets prepared by adsorbing half the amount of AAP to MAC nano-fibers containing the remaining amount of AAP (NFThalf) showed independent controlled-release aspects of AAP compared with physical mixture tablets (PMT). In vivo pharmacokinetic studies in rats after intraduodenal administration of 14 mg/rat AAP in NFT, NFTadso, and NFThalf demonstrated that all these tablets based on MAC nano-fibers showed sustained-release profiles compared with PMT, and showed ultra-sustained release properties for AAP. These new tablets based on MAC nano-fibers did not disintegrate in the intestine in the lower pH region, and the tablets could regulate the release of AAP in a pH-dependent manner. The ES method is a useful technique to prepare nano-fibers and showed promising results as an oral delivery system for sustained-release regulation.

Preparation of methacrylic acid copolymer S nano-fibers using a solvent-based electrospinning method and their application in pharmaceutical formulations.

In this study, we applied an electrospinning (ES) method, which is mainly employed in the textile industry, to the field of pharmaceuticals. We developed and modified an ES instrument and then utilized it to produce methacrylic acid copolymer S (MAC) nano-fibers to prepare tablets. By attaching a conductor rod made from stainless steel to the central part of the nano-fiber-collection plate of the ES apparatus, a MAC nano-fiber sheet could be produced effectively. In addition, we studied various operating conditions for this new ES method, including needle gauge, voltage between the electrodes, distance between the needle and nano-fiber-collection plate and the flow rate of MAC polymer solution, but these had no significant effect on the diameter of MAC nano-fibers. On the other hand, the viscosity (concentration) of MAC polymer solution and permittivity of solvent used to dilute MAC were closely related to the mean diameter of the nano-fibers. Tableting of MAC nano-fibers was performed using a tableting machine without lubricants, and addition of Tween 20 to the tablets enabled regulation of the release profile of a water-soluble drug. The modified ES method reported here is a useful technique for the controlled-release of drugs and has wide-ranging potential for pharmaceutical applications.

Comparison of the expression and function of ATP binding cassette transporters in Caco-2 and T84 cells on stimulation by selected endogenous compounds and xenobiotics.

Caco-2 and T84 cells are intestinal epithelial model cells. Caco-2 cells are more commonly used in drug transport studies, whereas only a few studies have used T84 cells, and the two cell lines have not been compared. We cultured Caco-2 and T84 cells on plastic dishes or polycarbonate Transwell filters and compared the expression and function of ATP binding cassette (ABC) transporters, including multidrug resistance protein (MDR) 1 and multidrug resistance-associated protein (MRP) 2 and MRP3, in response to various compounds. Overall, the pattern of change in transporter mRNA expression in response to compounds was very similar regardless of culture conditions (plastic dish or polycarbonate filter) and cell line (Caco-2 or T84), and changes in MDR1 function was accompanied by expression changes. The cells cultured on Transwell filters were more sensitive to the tested compounds, regardless of the cell line. On comparing the two cell lines, the intrinsic function of MDR1 was stronger in Caco-2 cells, while sensitivity to the tested compounds was more prominent in T84 cells. These results suggest that Caco-2 cells are more suitable for identifying whether MDR1 mediates drug transport, while T84 cells are more useful for assessing the induction capacity of compounds.

Preparation and pharmaceutical evaluation of new sustained-release capsule including starch-sponge matrix (SSM).

The focus of current study was to demonstrate a new sustained-release capsule including starch-sponge matrix (SSM) and to investigate how the pharmaceutical properties of SSM affect the drug release or its pharmacokinetic properties. Three representative drugs (uranine [UN], indomethacin [IMC] and nifedipine [NFP]) with different physicochemical properties (LogP(ow): 0.10, 1.18 and 3.23, respectively) were selected as model drugs. Model drug was dispersioned in pastelike cornstarch (starch glue) after heating 2.0-3.0% cornstarch suspension with electromagnetic wave at 2450 MHz (700 W) for l min. Then the drug mixture was encapsulated into a gratin capsule by a syringe, and the SSM including drug was prepared by means of a freeze-dried method. Essentially, drug-free SSM has a porous and netlike structure, and the distribution aspect of model drugs in the SSM depends on physicochemical properties between cornstarch glue and drugs. UN with much lower lipophilicity exists in continues phase of SSM, and IMC or NFP with a moderate or a higher lipophilicity exist in continues phase or porous space of the SSM. In the in vitro dissolution study, the release rate of drug from the SSM was mainly dependent on the lipophilicities of drugs, showing a rank order of the release rate of UN>IMC>NFP. In addition, the in vitro release rate for each drug was well regulated by changing the initial concentration of cornstarch suspension. In vivo absorption studies after intraduodenal administration of SSM capsule including model drug revealed that the sustained-release effects also could be regulated by the initial concentration of starch suspension. Moreover, the sustained-release effect of SSM capsule was enhanced with an increase in the lipophilicity of drug, and local-residential and mucoadhesive properties of SSM in the intestine provided stable supply of drugs from the SSM. The SSM capsule we developed here shows promising results as an oral drug delivery system for sustained-release regulation or target specificity.

Effect of cyclosporine on drug transport and pharmacokinetics of nifedipine.

Nifedipine (NFP) is an anti-hypersensitive drug and a well-known substrate of cytochrome P450 3A4 (CYP3A4), while cyclosporine (CSP) is a potent p-glycoprotein (P-gp) inhibitor. P-gp is a drug transporter, which determines the absorption and bioavailability of many drugs that are substrates for P-gp. Drugs that induce or inhibit P-gp may have a profound effect on the absorption and pharmacokinetics (PK) of drugs transported by P-gp within the body, possibly compromising their bioavailability. But the role of P-gp in the NFP efflux and its impact on PK profile is not known. Hence in our present study we attempted to investigate the effect of CSP on oral absorption and PK of NFP. Rhodamine 123 (Rho 123), a known P-gp substrate was used as a positive control. Male Wistar rats (350-400 g) were used for the study. Rats were divided into 4 groups (n=6 each); one group was treated with vehicle (cremophor) followed by NFP (0.2 mg/kg; i.v. bolus) and the other group with CSP (10 mg/kg; i.v.) followed by NFP. Group 3 and 4 were treated with vehicle (cremophor) followed by Rho 123 (0.2 mg/kg, i.v.) and CSP (10 mg/kg; i.v.) followed by Rho 123 (0.2 mg/kg, i.v.) respectively. The blood samples were collected at 0, 5, 10, 15, 30, 60, 90, 120, 180 and 240 min after NFP administration. NFP concentrations in plasma were analyzed by LC-MS-MS and Rho 123 was analyzed by fluorimetric detector. NFP efflux was significantly decreased in CSP treated rats (49.1% decrease, P<0.05), while NFP concentration in plasma were not changed. However the decrease in NFP efflux did not show any significant changes in NFP PK parameters (T(max); 2.0 vs. 2.5 min, C(max); 0.084 vs. 0.076 microg/ml, T(1/2); 84.0 vs. 91.4 min, AUC(0-t); 4.183 vs. 3.467 microg h/ml, AUC(infinity); 5.915 vs. 4.769 microg h/ml, AUMC(0-t); 224.073 vs. 173.063 microg h/ml, AUMC(infinity); 776.871 vs. 575.038 microg h/ml, MRT(0-t); 53.608 vs. 49.538 microg h/ml, MRT(infinity); 118.194 vs. 115.246 microg h/ml, CL(tot); 0.0375 vs. 0.0433 l/h, Vd(ss); 3.999 vs. 4.641 l in NFP alone vs. CSP+NFP groups respectively). Thus the results indicate that NFP would belong to a group of P-gp substrate. The decrease in efflux of NFP by CSP, through inhibition of P-gp, into the intestinal lumen did not show any impact on PK. This could be due to the activity of other transporters and/or CYP3A4 may have more limiting role than P-gp on NFP metabolism and disposition that is why inhibiting P-gp did not lead to increase the bioavailability and PK alterations.

Gender-related differences in expression and function of hepatic P-glycoprotein and multidrug resistance-associated protein (Mrp2) in rats.

To clarify whether gender-related differences exist in the expression and function of hepatic P-glycoprotein- and/or multidrug resistance-associated protein (Mrp2), we measured the hepatobiliary excretion of doxorubicin and their protein levels in male and female Sprague-Dawley rats. When rats received a single intravenous injection of doxorubicin (5 mg/kg), a delay in the disappearance of doxorubicin from plasma was observed in male rats. When rats received a constant-rate infusion of doxorubicin, no significant gender-related differences in the apparent biliary clearance of doxorubicin based on the steady state plasma concentrations were observed between male and female rats. However, the net biliary clearance of doxorubicin based on the liver concentration, which represents the actual function of P-glycoprotein and/or Mrp2, was higher in female rats than in male rats. These results suggest that the actual function of the hepatobiliary transport of doxorubicin is greater in female than in male rats. Western blot analysis revealed that the expression of P-glycoprotein and Mrp2 in the liver of female rats was significantly higher than in male rats, similar to results of hepatobiliary excretion experiments. The expression of hepatic cytochrome P450 (CYP) 2B1, which is involved in the metabolism of doxorubicin, was significantly higher in male than in female rats. By pretreatment with testosterone (10 mg/day for 7 days), the actual biliary clearance of doxorubicin in female rats was nearly that of male rats. The protein levels of P-glycoprotein and Mrp2 in female rats were also lowered by treatment with testosterone so as to be nearer those in male rats. These results suggest that gender-related differences exist in P-glycoprotein- and Mrp2-mediated hepatobiliary transport and that these two transporters may be regulated by sex hormones.

[Objective evaluation of clinical pharmacy lectures and experience in a master program using visual analog scale method].

We conducted an investigation to determine whether the visual analogue scale (VAS) method could be utilized in evaluating the lectures and pharmacy experience of the Division of Pharmacy and Health Sciences, Graduate School of Natural Science and Technology, Kanazawa University. Graduate students who had finished the 1-year pharmacy experience at the Kanazawa University Hospital were asked to make a self-evaluation of the understanding/attainment of lectures and experience in the course. Since the experience was carried out as a one-student-to-one-pharmacist system, the preceptors (pharmacists) were also asked to evaluate their corresponding students. When evaluating the necessity of the lectures, students tended to feel that the medical science- or pharmacotherapy-related subjects were important and those of social sciences were less important. These results suggest the need to review the contents of the lectures to enhance the interests of the students in the latter. By comparing the extent of understanding of each lecture before and after pharmacy experience, it was found that students had a better understanding of the lectures through their experience. In most results from the answers in pharmacy experience, students also scored themselves higher than their preceptors. Therefore comparisons of evaluation may provide more objective results in pharmacy experience. It was demonstrated that utilization of the VAS method and comparing the data are very useful in evaluating not only students' understanding/attainment but also the importance and usefulness of lectures and pharmacy experience in an objective way.

Experimental demonstration of the unstirred water layer effect on drug transport in Caco-2 cells.

We previously demonstrated that P-glycoprotein and MRP2 contribute to the secretory transport of grepafloxacin in the small intestine. Although inhibitors of these secretory transporters increased absorptive transport of grepafloxacin, secretory transport was not altered in Caco-2 cells, as determined by a conventional Transwell method. Because the value of the permeability coefficient of grepafloxacin is high, permeation through the unstirred water layer (UL) might be the rate-limiting step. To examine the possibility that the UL effect may mask the involvement of membrane transporters in the transport of drug with high permeability in Caco-2 cells, transport experiments were performed by agitating the experimental solution to decrease the thickness of the UL, and by lowering the temperature to decrease permeation via active transporters. Under these conditions, the UL effect was not rate limiting, and the inhibitory effects of transporter modulators were reflected in the apparent permeability as a decrease in secretory transport as well as an increase in absorptive transport. In conclusion, it was demonstrated that the UL can be the rate-limiting factor for transport of drugs with high membrane permeability in Caco-2 cells. When the UL affects the apparent permeability in an experimental apparatus in vitro, careful analysis is required to evaluate the contributions of transporters from the apparent permeability of drugs.

PepT1 mRNA expression is induced by starvation and its level correlates with absorptive transport of cefadroxil longitudinally in the rat intestine.

PURPOSE: To establish how closely intestinal transport activity for beta-lactam antibiotics is correlated with PepT1 expression, absolute expression level of PepT1 mRNA and transport activity were determined longitudinally in the small intestine of fed and starved rats. METHODS: For evaluation of absolute expression levels of PepTl mRNA, quantitative RT-PCR by LightCycler was used. The transport function was determined by quantifying the absorptive transport of cefadroxil across intestinal tissue sheets in a Ussing chamber. RESULTS: PepT1 mRNA expression was highest at the lower region and lowest at the upper region in the fed rats. The value of PepT1 was about 1/5-1/6 of that of GAPDH. The expression level in the starved rats was increased in all segments, but more profoundly in the upper region. Cefadroxil transport across intestinal tissue was higher in the lower region and lower in the upper region in fed rats, and increased in the upper region in starved rats. An excellent correlation was observed between expression levels and the permeability coefficients (r2 = 0.859, p < 0.05). CONCLUSIONS: The intestinal transport of cefadroxil is directly proportional to PepT1 expression, suggesting that the PepT1 expression level in the rat small intestine is the major determinant of the absorption of peptide-like compounds.

Involvement of multidrug resistance-associated protein 2 in intestinal secretion of grepafloxacin in rats.

We investigated the contribution of multidrug resistance-associated protein 2 (MRP2) to the secretory transport of grepafloxacin and compared its functional role with that of P-glycoprotein (P-gp) by using Sprague-Dawley rats (SDRs) and Eisai hyperbilirubinemic rats (EHBRs), in which MRP2 is hereditarily defective. In intestinal tissue from SDRs mounted in Ussing chambers, the level of transport in the direction from the serosal layer to the mucosal layer was twofold greater than that in the direction from the mucosal layer to the serosal layer. This secretory transport of grepafloxacin was diminished by both probenecid, an MRP2 inhibitor, and cyclosporine, a P-gp inhibitor. In intestinal tissue from EHBRs, the secretory transport of grepafloxacin was lower than that in intestinal tissue from SDRs and was inhibited by cyclosporine but not by probenecid. The absorption of grepafloxacin from intestinal loops in SDRs was in the order of duodenum > jejunum > ileum and was increased by cyclosporine but not by probenecid. The absorption in EHBRs was not higher than that in SDRs. The intestinal secretory clearance in SDRs after intravenous administration of grepafloxacin was shown to be greater for the ileum than for the duodenum, which is in good agreement with the previously reported regional expression profile of MRP2 mRNA. The intestinal secretory clearance was lower in EHBRs than in SDRs. Accordingly, in addition to P-gp, MRP2 might play a role in the secretory transport of grepafloxacin. The function of MRP2 in facilitating grepafloxacin transport in the secretory direction is more pronounced both in vitro and in vivo, while the restriction of entry from the lumen into the cell by MRP2 seems to be negligible, compared with that by P-gp, in the case of grepafloxacin.