Yukako Yoshikawa

Evaluation of intestinal pressure-controlled colon delivery capsule containing caffeine as a model drug in human volunteers.

The delivery ability of a pressure-controlled colon delivery capsule (PCDC) containing caffeine as a test drug was evaluated after oral administration to healthy male human volunteers. The driving force causing PCDC disintegration in the intestinal tract is the physiological luminal pressure which results from peristalsis. Three kinds of PCDCs having different thickness of a water-insoluble polymer membrane was prepared by coating the inner surface of the gelatin capsules with ethylcellulose (EC). The mean thickness were 40 +/- 1 (SE) for type 1, 44 +/- 1 for type 2 and 50 +/- 1 micron for type 3 PCDC, respectively. Caffeine was dissolved with a suppository base (PEGs 400 and 1000) and the capsules were filled. Doses were 15, 45 or 75 mg. After blank saliva samples were obtained, test preparations were orally administered to the volunteers and saliva samples were collected for 1 min intervals hourly from 1 to 10 h in the fasted state study, and from 1 to 20 h and at 25 h in the fed state study. Caffeine concentrations in the saliva samples were analyzed by HPLC. The maximum salivary caffeine excretion rate increased as the oral caffeine dose increased. The maximum salivary caffeine excretion rate increased predominantly compared to the pre-dose level in 75 mg dose study. Therefore, all following studies were performed with this dose. The first appearance time of caffeine into the saliva, TI, was used as a parameter to estimate the disintegration time of test preparations in the gastrointestinal tract. The mean TI of types 1, 2, and 3 PCDCs were 3.0 +/- 0.4, 4.0 +/- 0.4 and 4.5 +/- 0.3 h, respectively. After oral administration of 75 mg caffeine in pain gelatin capsule as a reference preparation, caffeine appeared in the saliva within 0.5 h. The mean hardness of the PCDCs were 1.05 +/- 0.10 (type 1), 1.55 +/- 0.06 (type 2) and 2.08 +/- 0.15 newton (type 3), respectively. There were good correlations between three parameters: EC coating membrane thickness, hardness and TI (determination coefficient r2 = 0.935 between TI and thickness, r2 = 0.998 between thickness and hardness, r2 = 0.958 between hardness and TI). The effect of food intake on the delivery ability was examined with type 3 PCDCs. Food intake prolonged the mean TI, from 4.5 +/- 0.3 to 7.8 +/- 1.3 h. This increase is thought to be ascribed to prolonged gastric emptying time. Comparison with reported colon arrival times indicates that the type 3 PCDC functions in colon delivery of caffeine and is thought to be applicable to other drugs.

Enhanced intestinal absorption of vancomycin with Labrasol and D-α-tocopheryl PEG 1000 succinate in rats

Vancomycin hydrochloride (VCM) is a glycopeptide antibiotic used for the treatment of infections caused by methicillin-resistant staphylococci. It is water soluble, having a high molecular weight, and poorly absorbed from the gastrointestinal tract. Mixtures of VCM with Labrasol and D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) were prepared to improve oral absorption of VCM. Administration of VCM solution to rat ileum at a dose of 20 mg/kg did not result in detectable plasma VCM concentration. Formulation containing 50% of Labrasol resulted in a Cmax value of 5.86+/-0.97 microg/ml and an AUC(0-6h) value of 16.06+/-1.78 microgh/ml. Addition of TPGS to VCM solution at 12.5% concentration also increased the plasma VCM concentration with a Cmax value of 4.98+/-0.45 microg/ml. But the AUC(0-6 h) (9.87+/-1.90 microgh/ml) was significantly lower than that obtained with Labrasol. The addition of 5.0 and 25.0% TPGS to solutions of VCM containing 50% of Labrasol did not result in any significant increase either in Cmax or AUC(0-6 h) of VCM. Whereas the addition of 12.5% of TPGS has resulted in an increase in Cmax and AUC(0-6 h) by 2.2 and 2.4 times, respectively, suggesting that this concentration of 50% Labrasol and 12.5% TPGS (1:0.25) was optimum for improving intestinal absorption of VCM. A dose dependent decrease in the Cmax and AUC(0-6 h) values was observed when the dose of absorption enhancers was decreased by 50% with formulation containing Labrasol and TPGS in 1:0.25 ratio. The results of the study indicate that formulations containing Labrasol and TPGS improve intestinal absorption of hydrophilic macromolecular drug, VCM.

Gastrointestinal mucoadhesive patch system (GI-MAPS) for oral administration of G-CSF, a model protein

A new gastrointestinal mucoadhesive patch system (GI-MAPS) has been designed for the oral delivery of protein drugs. The system consists of four layered films, 3.0 x 3.0 mm2, contained in an enteric capsule. The 40 microm backing layer is made of a water-insoluble polymer, ethyl cellulose (EC). The surface layer is made of an enteric pH-sensitive polymer such as hydroxypropylmethylcellulose phthalate (HP-55), Eudragit L100 or S100 and was coated with an adhesive layer. The middle layer, drug-containing layer. made of cellulose membrane is attached to the EC backing layer by a heating press method. Both drug and pharmaceutical additives including an organic acid, citric acid, and a non-ionic surfactant, polyoxyethylated castor oil derivative (HCO-60), were formulated in the middle layer. The surface layer was attached to the middle layer by an adhesive layer made of carboxyvinyl polymer (Hiviswako 103). Fluorescein (FL), 30mg, was first used as a model drug for oral administration of GI-MAPS having different surface layers in beagle dogs. The plasma FL concentration vs. time profiles demonstrated that the targeting of the systems was obtained, because the Tmax, the time when plasma FL concentrations reaches to its maximum lelev, was 2.33+/-0.82 h for HP-55 system, 3.33+/-0.41 h for Eudragit L100 system and 5.00+/-0.00 h for Eudragit S100 system. The same three kinds of GI-MAPSs containing 125 microg of recombinant human granulocyte colony-stimulating factor (G-CSF) were prepared and orally administered to dogs and the increase in total white blood cell (WBC) counts were measured as the pharmacological index for G-CSF. Comparison with the total increase of WBCs after iv injection of the same amount of G-CSF (125 microg) indicated the pharmacological availabilities (PA) of G-CSF were 23%, 5.5% and 6.0% for Eudragit L100, HP-55 and Eudragit S100 systems. By decreasing the amount of HCO-60 and citric acid, the PA of G-CSF decreased. These results suggest the usefulness of GI-MAPS for the oral administration of proteins.

Absorption enhancing effect of Labrasol on the intestinal absorption of insulin in rats

The oral absorption enhancing effect of Labrasol™ has been studied in rats using insulin as a model peptide/protein drug. Insulin solution was prepared by dissolving insulin in pH 7.4 buffer followed by the addition of Labrasol. The insulin concentration was 50.0 IU/ml. The test insulin/Labrasol solution was administered to the jejunum, ileum and ascending colon of rats at 10.0 IU/kg. After administration, blood samples were collected for 5 h and serum glucose levels and insulin levels were measured. In another group of rats, insulin solution was injected intravenously at 1.0 IU/kg, and both serum glucose and insulin levels were measured. The pharmacological availability of insulin from Labrasol solution was found to be 3.9, 8.9 and 9.1% following jejunal, ileal and colonic administrations, respectively, by comparing the serum glucose level vs. time profiles obtained after intestinal and i.v. administrations. By comparing the serum insulin levels vs. time profiles, the bioavailability of insulin was found to be 0.25 and 0.20% for intra-ileum and colonic administrations, respectively. The hypoglycemic effect of insulin after intra-ileum administration showed a dose-dependency in the insulin dose range from 10.0 to 1.0 IU/kg. These results suggest the absorption enhancing effect of Labrasol on the intestinal absorption of insulin in rats.

Application of pressure-controlled colon delivery capsule to oral administration of glycyrrhizin in dogs.

A colon delivery system has been used to improve the bioavailability of glycyrrhizin, a glycoside of glycyrrhetic acid. The bioavailability of glycyrrhizin is low when administered in conventional oral galenic dosage forms because glycyrrhizin is enzymatically hydrolysed both in the stomach and in the intestine. It was reasoned that if large amounts of glycyrrhizin were directly delivered to the colon, enzymatic activity should be reduced due to saturation so that intact glycyrrhizin could be absorbed into the systemic circulation. Based on this assumption, pressure‐controlled colon delivery capsules (PCDCs) were used as a colon delivery system. Eight types of glycyrrhizin solution were prepared and were introduced into PCDCs. After oral administration of the test PCDCs to beagle dogs, blood samples were obtained over 24 h and plasma glycyrrhizin concentrations were measured by an HPLC method. With PCDCs containing aqueous glycyrrhizin and propylene glycol solutions, plasma glycyrrhizin levels were extremely low and the bioavailabilities of glycyrrhizin were 0.6% and 0.4%, respectively. When Labrasol was added to both types of glycyrrhizin solution, the bioavailability was improved to 4.6 % for aqueous solution and 3.8% for propylene glycol solution. When a surfactant, Polysorbate 80, was added in combination with Labrasol, synergistic effects were not obtained. Furthermore, dose‐dependent effects of Polysorbate 80 were not obtained. Labrasol, which is a component of self‐emulsifying drug delivery systems (SEDDS), has been shown to strongly improve the bioavailability of glycyrrhizin from the colon.

Evaluation of an intestinal pressure-controlled colon delivery capsules prepared by a dipping method

A new method for preparation of large amounts of empty pressure-controlled colon delivery capsules (PCDCs) by a dipping method has been developed. Empty PCDCs are composed of two polymer membranes. The inner one was a water-insoluble polymer membrane, ethylcellulose (EC). The outer one was an enteric polymer membrane, hydroxypropylmethylcellulose phthalate (HPMCP) or hydroxypropylmethylcellulose acetate succinate (HPMCAS). By consequently dipping into an ethanolic EC solution and an alkalized enteric polymer solution, empty PCDCs were obtained after both the capsule body and cap were adjusted to the size of #2 capsules. With each enteric polymer, two types of empty PCDCs of different thickness were prepared. Fluorescein (FL) was formulated with suppository base, PEG1000, and used as a model drug. FL/PEG1000 suspension was introduced into empty PCDCs which were then sealed with enteric polymer solution. The PCDCs were evaluated by an in vivo experiment using beagle dogs. After oral administration of the test PCDC preparations containing 30 mg of FL, blood samples were obtained from the jugular vein and serum FL levels were measured. The thickness of the EC membrane layer varied in both the capsule body and cap. HPMCAS PCDCs had 62.1+/-5.0 (S.E.) microm (body) and 49.7+/-3.3 microm (cap) with thicker ones and 55.7+/-6.6 microm (body) and 46.8+/-6.2 microm (cap) with thinner ones. HPMCP PCDCs had 28.1+/-3.3 microm (body), 30.9+/-1.0 microm (cap) with thinner ones and 43.1+/-9.8 microm (body), 42.4+/-8.2 microm (cap) with thicker ones. The mean T(i) values, the first appearance time, of FL in the serum of HPMCAS PCDCs were 2.0+/-0.7 h for thicker ones and 3.8+/-0.5 h for thinner ones, while the mean T(i) values of HPMCP PCDCs were 2.0+/-0.0 h for thinner ones and 3.5+/-0.7 h for thicker ones. Since the colon arrival time in beagle dogs was 3.5+/-0.3 h as determined by a sulfasalazine test, thinner HPMCAS PCDCs and thicker HPMCP PCDCs were thought to deliver FL to the colon.

Retention and transit of intestinal mucoadhesive films in rat small intestine

The retention and transit characteristics of intestinal mucoadhesive film systems have been studied after intraduodenal administration in rats. Small size four layered film preparations, 0.5x0.5 mm, were prepared, where the backing layer (45.1+/-2.9 microm thick) was made of a water-insoluble polymer, ethylcellulose (EC), the surface layer was made of enteric pH-sensitive polymers, Eudragit L100, S100 or HP-55 and the middle layer was made of cellulose membrane. The surface layer was attached to the middle layer with an adhesive layer composed of carboxyvinyl polymer (Hiviswako(R) 103). After administration of ten films to the duodenum, the rats were sacrificed hourly and the distribution of the films in the whole small intestine was directly observed after abdominal incision. The HP-55, Eudragit L100 and S100 film systems were found to adhere to the upper, middle and lower part of the small intestine after 1, 2 and 4 h, respectively, for 2-3 h. Direct inspection study suggests that intestinal mucoadhesive film system has functions of: (1) pH-dependent intestinal adhesion site specificity; (2) adhesion to the intestinal wall; and (3) retention in the small intestinal adhesion site for at least 2 h. Intestinal mucoadhesive film system has been suggested to be a targeting system for drugs to the gastrointestinal tract.

Importance of dissolution process on systemic availability of drugs delivered by colon delivery system

The relationship between in vitro drug release characteristics from colon delivery systems and in vivo drug absorption was investigated using three kinds of delayed-release systems. 5-aminosalicylic acid (5-ASA), tegafur (FT) and carbamazepine (CBZ) were selected as model drugs. Pressure-controlled colon delivery capsules (PCC) for liquid preparations, time-controlled colon delivery capsules (TCC) for liquid and solid preparations and Eudragit S coated tablets for solid preparations were used in this study. At first, in vitro dissolution tests for all preparations were performed. Drug release from solid preparations was delayed compared to that from liquid preparations with all three drugs. Next, these preparations were administered to fasted beagle dogs. For 5-ASA, the mean Cmaxs (peak level) of Eudragit S coated tablets and PCC were 5.52 and 16.89 micrograms ml-1, respectively. The mean Tmaxs (time when drug reached peak level) were 3.0 and 5.3 h. AUCs were 22.57 and 48.09 micrograms.h ml-1, respectively. For FT, Cmaxs of Eudragit S coated tablet and PCC were 0.87 and 1.46 micrograms ml-1, and Tmaxs were 7.0 and 6.7 h, respectively. AUCs were 9.73 and 15.55 micrograms.h ml-1 and bioavailabilities were 43.79 and 70.84%. For CBZ, the mean Cmaxs of liquid preparations and solid preparations were 0.37 and 0.22 micrograms ml-1, respectively. The mean Tmaxs were 4.7 and 4.3 h. AUCs were 0.673 and 0.392 micrograms.h ml-1. With liquid preparations, drug was thought to contact to the colonic membrane easily because of lack of interference by stools, and to be absorbed well as compared with solid preparations. From these findings, drug release from colon delivery systems and drug dissolution in the colonic lumen are very important factors for the systemic availability of drugs from the colon delivery systems.

Highly sensitive quantification of vancomycin in plasma samples using liquid chromatography–tandem mass spectrometry and oral bioavailability in rats

We developed a highly sensitive liquid chromatography-tandem mass spectrometry assay (LC-MS-MS) for a glycopeptide antibacterial drug, vancomycin (VCM), in rat plasma. After precipitating 100 micro l of plasma with 300 micro l of 10% trifluoroacetic acid-methanol (2:1, v/v), the supernatant was diluted with 300 micro l of distilled water and was passed through a filter. LC-MS-MS equipped with electrospray ionization in the positive ion mode used a pair of ions at 725/144 m/z for VCM in the multiple reaction-monitoring mode with a sample injection volume of 20 micro l. The calibration curve had a linear range from 0.01 to 20 micro g/ml when linear least square regression was applied to the concentration versus peak area plot. The drug in the sample was detected within 5 min. Precision, accuracy and limit of quantitation indicated that this method was suitable for the quantitative determination of VCM in rat plasma. Using this method, we defined for the first time that the oral bioavailability of VCM in rats was 0.069%. This method can be applied to basic pharmacokinetic and pharmaceutical studies in rats.

New preparation method of intestinal pressure-controlled colon delivery capsules by coating machine and evaluation in beagle dogs

A new method for preparing large amounts of pressure-controlled colon delivery capsules (PCDCs) which employs a pharmaceutical coating machine, Hicoater-mini, has been developed. In contrast to our original method for preparing PCDCs where the inner surfaces of gelatin capsule were coated with the water-insoluble polymer ethylcellulose (EC), PCDC were directly prepared by coating the capsular shaped suppositories with EC. As a model drug, fluorescein (FL) was used in this study. FL powder was suspended with the suppository base, polyethylene glycol (PEG) 1000, at 50 degreesC, and was hardened in the capsular shape the sizes of which were #0 and #2. The capsular shaped suppositories were coated with 5% w/v ethanolic EC (7G grade) solution by a coating machine. By increasing the coating time from 55 to 75 min, the mean coating thickness of #0 PCDCs increased from 141+/-7 to 211+/-4 micrometer. In the case of #2 PDDCs, the mean coating thickness increased from 102+/-3 to 110+/-5 micrometer by increasing the coating time from 35 min to 40 min. Several kinds of #0 PCDCs having the mean EC coating membrane thickness of 141+/-7 micrometer (type 1), 166+/-4 micrometer (type 2), 188+/-4 micrometer (type 3), 211+/-4 micrometer (type 4) as well as #2 PCDCs having thickness of 102+/-3 micrometer (type 5) and 110+/-5 micrometer (type 6) were used for in vivo evaluation using beagle dogs. After oral administration of the test preparations containing 30 mg of FL, blood samples were obtained from the jugular vein and plasma FL levels were measured. The first appearance time, Ti, of FL in the plasma was used as a parameter for the estimation of the release time of FL from PCDCs in the gastrointestinal tract. The mean Ti of #0 PCDCs were 2.3+/-0.5 for type 1, 3.3+/-0.5 for type 2, 4.8+/-1.0 for type 3 and 7.8+/-1.7 h for type 4 preparations while the mean Ti of #2 PCDCs were 3.2+/-0.4 for type 5 and 3.8+/-0.4 h for type 6, respectively. There were good correlations between EC coatings.