Absorption enhancement of polypeptide drugs by cyclodextrins. I. Enhanced rectal absorption of insulin from hollow-type suppositories containing insulin and cyclodextrins in rabbits.

The absorption of insulin (from porcine pancreas) from the rectum of rabbits after the administration of hollow-type suppositories containing insulin and five kinds of cyclodextrins (CyDs) was investigated. Three types of suppositories were employed: suppository I containing insulin (approximately 26 IU/mg) and various amounts of each CyD in citric buffer solution at pH 3.0 or powder in its cavity, suppository II containing CyD without insulin, and suppository III containing insulin without CyD. Without CyD, the insulin and glucose levels in plasma were unchanged, whereas a significant increase in the plasma insulin concentration and a marked decrease in the glucose levels were found following simultaneous administration of insulin and CyDs by suppository I. The enhancing effect of CyD on rectal insulin absorption (absorption-enhancing effect) by chemically modified CyDs (heptakis(2,6-di-O-methyl)-beta-CyD (DM-beta-CyD) and 2-hydroxypropyl-beta-CyD (HP-beta-CyD)) was higher than those by natural CyDs (alpha-, beta-, and gamma-CyD). The area under the plasma concentration-time curve (AUC) and Cmax of insulin significantly decreased with the preadministration (administration of CyD 6, 24 and 48 h before rectal insulin administration) of DM-beta-CyD. The absorption-enhancing effect disappeared 24 h after preadministration. These results suggest that CyDs enhance insulin absorption from the rectum, and that attenuation of the membrane transport barrier function in the rectum recovers at a maximum of 24 h after administration of CyDs.     

Enhancing effect of glyceryl-1-monooctanoate on the rectal absorption of gentamicin from hollow-type suppositories in rabbits

A hollow-type suppository containing gentamicin (GM) in its cavity was prepared using Witepsol H-15 (H-15) mixed with glyceryl-1-monooctanoate (MO) or MO alone in the body of the suppository (type I) and a suppository (type II) containing GM and MO in the cavity was constructed using H-15 in the body of the suppository. Without MO, GM (60 mg) was not absorbed (plasma GM levels less than 1 microgram/ml). However, the absorption of GM from the rectum of rabbits was enhanced by coadministered MO in types I and II. Even when the amount of GM was decreased to 6 mg (1/10), GM was observed in the plasma (Cmax, 3.5 +/- 0.3 micrograms/ml) after administration of the suppository made from MO mixed with H-15. The enhancing effect of MO on the rectal absorption of GM could not be further increased by incorporating an amount of MO larger than approximately 300 mg into the suppository. This study demonstrates that MO can be used in the two types of hollow suppositories as an effective enhancing agent of rectal absorption of poorly absorbed drugs such as GM.     

Studies on sustained-release suppositories. I. Effect of alginic acid addition on rectal absorption of bacampicillin in rabbits

Sustained-release suppositories of bacampicillin (BAPC) were prepared by the use of the adduct which was precipitated from an aqueous solution containing BAPC and alginic acid (Alg). As the suppository base, Witepsol H-15 and macrogol were used. Absorptions of BAPC from the suppositories were prolonged in rabbits, but the bioavailabilities were decreased compared to that after administration of BAPC alone. However, these absorptions were improved enormously by the addition of surface-active agents, that is, an excellent prolonged absorption and high bioavailability were obtained. Interestingly, similar prolonged absorption could be obtained only by mixing Alg with BAPC in a suppository base. Further, this absorption rate was found to be controlled by the amount of Alg addition. The absorption profiles from a suppository containing the mixture differed from that containing the adduct in exhibiting both high plasma level and prolonged absorption. This may be due to simultaneous fast absorption of BAPC itself and formation of the adducts. Thus, it seemed that BAPC preparations containing Alg may be practically useful as a rectal preparation with prolonged action and giving a high plasma level.     

Studies on sustained-release suppositories. III. Rectal absorption of morphine in rabbits and prolongation of its absorption by alginic acid addition

Rectal absorption of morphine from various kinds of suppository bases was investigated. The extent of bioavailability of morphine by rectal administration varied with the bases used (30.5-97.5%), but every value was higher than that in the case of oral administration (13.5%). Witepsol bases were preferable to macrogol base for the rectal absorption of morphine. In particular, Witepsol S-55 or W-35 gave a higher plasma peak level than H-15 or E-75, whereas the difference in the mean residence times obtained from these bases could not be regarded as significant. Sustained-release suppositories of morphine could be prepared simply by mixing alginic acid (Alg) with morphine in a suppository base. Further, prolonged rectal absorption could be obtained by using these sustained-release suppositories, and the absorption rate was controlled by the amount of Alg added. It seems likely that the sustained release was due to the binding of morphine to Alg from the results of partition coefficient and binding ratio measurements in aqueous solution. The rapid initial absorption and the subsequent prolonged absorption of morphine simultaneously obtained from the morphine-Alg suppository may be useful in the clinical context.     

Studies on sustained-release suppositories. II. Evaluation of polymer electrolyte containing acidic groups for prolonged rectal absorption of bacampicillin in rabbits

Rectal absorption of bacampicillin hydrochloride (BAPC) was found to show the best bioavailability with Witepsol H-15 as suppository base among various Witepsol bases. However, an effective plasma concentration of drug (above 0.5 micrograms/ml) was only maintained for 2 h, so sustained-release suppositories of BAPC were studied. Bacampicillin reacts with acidic polymer electrolytes such as pectic acid (Pc), chondroitin sulfate (Cd) and precipitates as its adduct with the polymer in an aqueous solution. The dissolution rate of BAPC from the adducts in a solution was slower than that of BAPC itself. The absorptions of BAPC from the suppositories containing the adducts were prolonged, but the bioavailabilities were decreased compared to that from the suppository containing BAPC alone. Similar prolonged absorption could be obtained simply by mixing Pc or Cd with BAPC in a base. Further, the absorption rate was found to be controlled by the amount of the polymer addition, and both a high plasma level and excellent bioavailability were obtained. This desirable outcome may be due to the simultaneous occurrence of rapid absorption of BAPC itself and formation of the adducts.     

Indomethacin sustained-release suppositories containing sugar ester in polyethylene glycol base

Indomethacin (IM) sustained-release suppositories were prepared by the fusion method using sugar ester and polyethylene glycol 4000 (PEG). The suppositories were evaluated by in vitro release testing, X-ray analysis and in vivo absorption testing in rabbits. X-ray analysis showed that IM was amorphous in PEG-base suppositories. In a release test, slow-release was obtained when the sugar ester content of a suppository was 60%. The IM plasma level following the administration of the suppository was well sustained in the absorption test. The main slow-release mechanism is considered to be the release of IM from the matrix composed of sugar ester and PEG, which is represented by the Higuchi equation. A good correlation between the release test and the absorption test was obtained. It is considered that the amorphous state of IM in this type of sustained-release suppository would enhance the release and absorption of IM in the rectum of the rabbit, whose rectal fluid volume is small.     

Enhancing effect of cyclodextrins on nasal absorption of insulin and its duration in rabbits.

The absorption of insulin (from porcine pancreas) in rabbits after the nasal administration of aqueous preparations containing insulin and five kinds of cyclodextrins (CyDs) in phosphate buffer solution at pH 7.0 was investigated. Without CyD, the insulin and glucose levels in plasma were unchanged, whereas a marked increase in the plasma levels of insulin and a decrease in glucose concentrations were observed following the simultaneous administration of insulin and CyD such as alpha- and heptakis (2,6-di-O-methyl)-beta-CyD (DM-beta-CyD). The largest enhancing effect on the nasal absorption of insulin was obtained by DM-beta-CyD. To evaluate the duration of the absorption-enhancing effect of CyDs, preadministration (administration of CyD 0.5, 6, 12 and 24 h before insulin administration) was performed. The area under plasma concentration-time curve (AUC) and Cmax of insulin significantly decreased with the preadministration of DM-beta-CyD 6, 12 and 24 h before nasal administration. The absorption-enhancing effect disappeared 24h after the preadministration. These findings demonstrate that CyDs enhance the nasal absorption of insulin, and the recovery of the membrane transport barrier function in nasal mucosa is achieved, at the latest, 24 h after the administration of CyDs.     

Thermo-reversible flurbiprofen liquid suppository with HP-β-CD as a solubility enhancer: improvement of rectal bioavailability

The purpose of this work was to develop a thermo-reversible flurbiprofen liquid suppository base composed of poloxamer and sodium alginate for the improvement of rectal bioavailability of flurbiprofen. Cyclodextrin derivatives such as α-, β-, γ-cyclodextrin and hydroxypropyl-β-cyclodextrin (HP-β-CD) were used to enhance the aqueous solubility of flurbiprofen. The effects of HP-β-CD and flurbiprofen on the physicochemical properties of liquid suppository were then investigated. Pharmacokinetic studies were performed after rectal administration of flurbiprofen liquid suppositories with and without HP-β-CD or after intravenous administration of commercial Lipfen^® (flurbiprofen axetil-loaded emulsion) to rats, and their pharmcokinetic parameters were compared. HP-β-CD decreased the gelation temperature and reinforced the gel strength and bioadhesive force of liquid suppository, while flurbiprofen was opposed to HP-β-CD. Thermo-reversible flurbiprofen liquid suppository showed the physicochemical properties suitable for rectal administration. The flurbiprofen liquid suppository with HP-β-CD showed significantly higher plasma levels, AUC and C_max of flurbiprofen than those of the liquid suppository without HP-β-CD, indicating that flurbiprofen could be well absorbed due to the enhanced solubility by formation of inclusion complex. Moreover, the flurbiprofen liquid suppository with HP-β-CD showed an excellent bioavailability in that the AUC of flurbiprofen after its rectal administration was not significantly different from that after intravenous administration of commercial Lipfen^®. It is concluded that HP-β-CD could be a preferable solubility enhancer for the development of liquid suppository containing poorly water-soluble drugs.     

Enhancement effect of lauric acid on the rectal absorption of propranolol from suppository in rats.

In a previous paper, we have demonstrated that medium chain fatty acids significantly enhance the in vitro rectal absorption of propranolol (PL) and that the enhancement may be partly due to the formation of a complex with a fatty acid at a 1:1 molar ratio. To confirm in vivo the enhancement effect of lauric acid on PL absorption, PL suppositories with lauric acid at various molar ratios were administered to rat rectum. PL absorption from Witepsol and macrogol suppositories with lauric acid at a 1:1 molar ratio was much larger than that after PL alone and the 1:2 or 1:3 molar ratio ones. The bioavailability (BA) after administration of the 1:1 molar ratio suppository (PL, 4 mg/kg) was 1.6- and 2.1-fold for the Witepsol and macrogol formulations respectively, compared with that after PL alone. A similar result was obtained with the PL solid dispersion suppository with lauric acid at a 1:1 molar ratio, showing a 1.7-fold higher BA compared with PL alone. The release of PL from the macrogol suppository was significantly faster at a 1:1 molar ratio than that of other preparations, but not so in the solid dispersion suppository. There was not good agreement between the release rates of PL from the suppositories and the plasma levels after dosing. These results supported the concept that a portion of PL, by forming a 1:1 complex with lauric acid, would penetrate across the rectal mucosa more easily than PL alone.     

Indomethacin sustained-release suppositories containing sugar ester

We prepared indomethacin (IM) sustained-release suppositories using sugar ester (SE) as an additive. The suppositories were prepared by the fusion method with IM, SE, and Witepsol H-15 (H-15) and their availabilities in vitro and in vivo were evaluated mainly by the drug release test and the absorption test in rabbits, respectively. The softening point of the suppositories increased with increasing SE content. In the release test with the Muranishi method, slow-release profiles were obtained when the SE content was more than 52.5%. The absorption of IM from these suppositories, however, was very little. In the other release test, e.g. immersion method with gauze, all of the suppositories with SE showed slow-release profiles, and the drug release rates clearly depended on the SE content. The drug was released from the suppositories by the following leaching-type mechanism proposed by Higuchi. The suppository with a 30% SE content showed a sustained-plasma level of IM, but the other suppositories did not. It was concluded that an appropriate content of SE (i.e. 30%) in the suppository base was required to obtain sustained-release because it reasonably regulated the infiltration of rectal fluid into the suppository and the mechanical strength of the suppository against disintegration.     

Bioavailability of morphine in rabbits after rectal administration of suppository containing controlled release morphine tablet.

Two kinds of sustained release morphine suppositories have been prepared; one is an oleaginous base suppository (MSC) containing a controlled release morphine tablet (MST: MS Contin), and the other is a hollow-type suppository (MSCH) containing MST and morphine powder packed in its hollow space. In vitro release tests and in vivo rectal absorption experiments in rabbits were performed. The profiles of morphine release from MST and MSC in vitro were similar, and revealed that suppository bases had no effect on the release profile of morphine from the preparation. Morphine release from MSCH was rapid in the early phase, and then enclosed morphine was slowly and continuously released from MST. Phamacokinetics of morphine from the suppository were compared with the orally administered MST, and it was found that there was no difference in the maximum plasma concentration (Cmax) and the peak time (Tmax) between MSC and MST, but the mean residence time (MRT) of MSC was approximately three times longer than that of MST, and the extent of bioavailability (BA) of MSC was significantly larger than that of MST (71.6 +/- 14.2% and 11.9 +/- 4.0%, respectively). Cmax can be altered arbitrarily by changing the morphine content in the hollow space of MSCH. As in the case of MSC, the plasma concentration of morphine from MSCH was maintained. It is concluded from the above results that MSC is a satisfactory sustained release morphine suppository for the treatment of cancer pain, administering it twice a day, and that MSCH is effective due to its fast analgesic effect and sustained release nature not only for cancer pain but also for surgical operations.     

Studies on the crystal structure of (D-Ala)-B0 porcine insulin at 1.9 A resolution

The crystal structure of (D-Ala)-B0 porcine insulin has been determined, using data to 1.9 A and atomic parameters of 2 Zn porcine insulin as a starting model, and through the use of the difference method and the restrained least square method, to a final R-factor of 0.211 and r.m.s. deviation of 0.057 A for the bond lengths. The electron densities of B0 residues were very clear. Introduction of B0 residues into the molecules had reduced the thermal vibration of the N-terminus of B-chain for both molecules I and II and made the molecules pack closer in the crystal. The obvious differences between the crystal structures of 2 Zn and (D-Ala)-B0 porcine insulin were the conformations of partial polar groups around the possible receptor binding surface and the assembly mode of two helixes of A-chain in molecule I. In the local environment of the N-terminus of B-chain there were great differences between the crystal structures of (D-Ala)-B0 porcine insulin, (Trp)-B1 porcine insulin and Des B1(Phe) bovine insulin. In this paper the structure-immunoactivity relationships of insulin molecule have also been discussed briefly.     

Cisplatin suppository: preparation, release characteristics and clinical evaluation

Cisplatin (CDDP) has attracted attention as a chemotherapeutic agent for the treatment of uterine endometrial carcinoma but causes serious side effects, including renal toxicity. CDDP suppositories containing NaCl at different concentrations were prepared to enhance the efficacy and to reduce the side effects of CDDP. The release characteristics, melting point and viscosity of the suppositories were first studied. The rate of CDDP release increased as the NaCl concentration increased: it was 12% 12 h after administration of suppositories containing no NaCl, but 32% with 0.2% NaCl. The melting point was raised by addition of NaCl: 35.5 degrees C without NaCl and 36.5 degrees C with 0.2% NaCl. Addition of 0.2% NaCl doubled the viscosity. Clinically, the suppository containing 0.06% NaCl was given to 3 patients with endometrial carcinoma twice a week for 3 weeks to examine serum CDDP levels and endometrial absorption. Patients with endometrial carcinoma showed different peak plasma platinum (Pt) levels which were as low as 0.12, 0.06 and 0.22 micrograms Pt/ml with similar patterns of change in the level. Radiographic analysis revealed many Pt particles in sections of necrosed endometria after 21d of the treatment. No side effects of CDDP were found in biochemical testing or subjective symptoms.     

A biologically active insulin analogue with modification in the A2 position. [2-Valine-A] sheep insulin

The synthesis and biological evaluation of [2-Valine-A] insulin ([Val2-A]insulin) is reported. In this insulin, the isoleucine residue in position A2, invariant in the majority of mammalian insulins, is substituted by valine. The same substitution, along with four others, occurs naturally in the insulin produced by the owl monkey. Owl monkey insulin exhibits ca. 20% of the activity of porcine insulin in in vitro insulin assays using human cells in culture. [Val2-A]insulin displays 20-22% of the activity of bovine insulin in in vitro insulin assays using rat liver plasma membranes or isolated rat adipocytes. We suggest that the substitution of valine for isoleucine at position A2 is responsible for all or most of the diminution in potency of owl monkey insulin relative to porcine insulin. The data are discussed with regard to previous findings with insulin analogues in which isoleucine A2 was replaced with norleucine, glycine and alanine.     

Functional Monomers and Polymers. LX. Effects of Counterions on the Structure of Cu(II)-Polyvinylimidazole and Cu(II)lmidazole Complexes in Aqueous Solution

The effect of counterion types on the structure of Cu(II) complexes of imidazole and poly-4(5)-vinylimidazole in aqueous solution was studied by spectroscopic and electron spin resonance (ESR) techniques. Visible and ESR spectra of both complexes in aqueous solution containing NaClO₄, KNO₃, KC1, and KBr showed only a slight change from each other in the presence of different counterions. On the other hand, the frequency of the center of absorption band in the visible region (λ_max) and ESR parameters of the complex in aqueous solution did not correspond to those reported for its crystalline state, and in addition, those of the polymeric Cu(II) complex were different from those of the low molecular weight Cu(II) complex. It was proposed that in aqueous solution, water molecules are coordinated to the axial position of the complex where counterion molecules are present in the crystalline state, and that the structure of the polymeric Cu(II) complex is different from that of the low molecular weight Cu(II) complex due to steric hindrance of the polymeric ligand.     

Preparation and characterization of insulin-loaded acrylic hydrogels containing absorption enhancers

The objectives of this study were to prepare insulin-loaded acrylic hydrogel formulations containing various absorption enhancers, to perform in vitro and in vivo characterization of these formulations, and to evaluate the factors which affecting insulin availability on rectal delivery of insulin using this hydrogel system. The acrylic block copolymer of methacrylic acid and methacrylate, Eudispert, was used to make the hydrogel formulations. As absorption enhancers, 2,6-di-O-methyl-beta-cyclodextrin (DM-beta-CyD), lauric acid (C12), or the sodium salt of C12 (C12Na), were incorporated into the hydrogels. In an in vitro release test, the release rate of insulin from the hydrogels decreased as the polymer concentration of the hydrogel increased. The addition of C12Na to the hydrogel further increased the insulin release rate, which was greater at higher concentrations of the enhancer. A portion of the C12Na was found to remain bound to the acrylic polymer in dissolution medium. Serum insulin levels were determined at various time points after the administration of insulin solution or insulin-loaded (50 units/kg body weight) Eudispert hydrogels containing 5% (w/w) of C12, C12Na, or DM-beta-CyD to in situ loops in various regions of the rat intestine. The most effective enhancement of insulin release was observed with formulations containing C12Na. The bioavailability of insulin from the hydrogels was lower than that from the insulin solutions. Hydrogel formulations containing 7% or 10% Eudispert remained in the rectum for 5 h after rectal administration. However, the 5% (w/w) C12Na solution stained with Evan's-blue had diffused out and the dye had reached the upper intestinal tract within 2 h. Finally, the rectal administration of insulin-loaded hydrogels, containing 4%, 7%, or 10% (w/w) Eudispert and 5% (w/w) of enhancer (C12, C12Na, or DM-beta-CyD) to normal rats was shown to decrease serum glucose concentrations. The greatest effect was found with insulin-loaded 7% (Eudispert) hydrogel containing C12Na which having cosiderable large insulin release rate and bioadhesive characteristics.     

Preparation and evaluation of eudragit gels. V. Rectal gel preparations for sustained release and avoidance of first-pass metabolism of lidocaine.

The release of lidocaine from hydrogel and xerogel preparations was remarkably suppressed compared with polyethylene glycol (PEG) 2000 suppository. The release rate of lidocaine from hydrogel and xerogel increased with the increase in the amount of sodium hydroxide incorporated within the range of 3 to 7 milliequivalent (meq). After an oral administration of lidocaine HCl solution, the plasma concentration of lidocaine was considerably lower than that after intravenous administration for all time periods. The absolute bioavailability (F(oral)) was 5.63%. For the Witepsol S-55 and PEG 2000 suppositories, the plasma levels of lidocaine were higher than those for the oral preparation, and Cmax and area under the concentration-time curve (AUC) values significantly improved (p < 0.01). The absolute bioavailabilities were 21.3 and 29.6%, respectively. On the other hand, Eudispert hv-hydrogel and xerogel preparations showed the characteristics of a sustained-release preparation, especially the xerogel preparation with 5 meq NaOH. Absolute bioavailability for hydrogel and xerogel preparations increased significantly (p < 0.05) by approximately 1.7-3.4 folds compared with those of Witepsol S-55 and PEG 2000 suppositories.     

Antihyperglycemic effect of insulin from self-dissolving micropiles in dogs

As a percutaneous delivery device, self-dissolving micropiles (SDMPs) composed of chondroitin sulfate and insulin were prepared under room temperature from highly concentrated solution, glue. The mean weight of SDMP was 1.03+/-0.04 mg. One insulin SDMP was percutaneously administered to the shaved abdominal skin of four beagle dogs at insulin dose level of 1.0 and 2.0 IU/dog. After administration, blood samples were collected for 6 h and plasma glucose levels were measured. The time when minimum plasma glucose level appeared, T(min), was 1.38+/-0.2 h for 1.0 IU study and 1.38+/-0.1 h for 2.0 IU study and clear dose-dependent hypoglycemic effect of insulin was observed in the dose range. By comparing the area above the plasma glucose level vs. time curve (AAC) between insulin SDMP and subcutaneous (s.c.) injection solution, the relative pharmacological availabilities were 99% (1.0 IU) and 90% (2.0 IU), respectively. To ascertain the usefulness of insulin SDMP, oral glucose tolerance test (OGTT) was performed. When dogs were treated with insulin SDMPs, 2.0 IU, followed by an OGTT 30 min, glycemia did not appear for 5 h. On the other hand, when OGTT was performed at 1 h after insulin SDMP administration, hypoglycemia appeared as in the case of s.c. injection of insulin solution, 2.0 IU. Insulin SDMP improved the oral glucose challenge for 3 h, with a maximum effect at 30 min before the administration of glucose. Those results suggest the usefulness of a SDMP for the percutaneous delivery of peptide/protein drugs like insulin.     

PDMS-chemistry of angiotensin II and insulin in glucose glass thin films

A plasma desorption mass spectrometry study was made on the properties of glucose and glucose/glucuronic acid thin films as matrices for amino acids, small and large peptides and insulin. Amino acids and small peptides are distributed throughout the film as it is formed from aqueous solutions and the mass spectra are similar to what is observed for nitrocellulose matrices. AngiotensinII (angII), insulin, and reduced insulin containing the separated A- and B-chains concentrate at the surface of the film due to the hydrophobic interaction. Extensive positive and negative fragmentation patterns are observed for angII using the glucose glass film. The fragment ions appear to be formed from layers just below the surface of the film. The co-matrix of glucuronic acid/glucose produces a higher molecular ion yield. The spectrum of insulin in glucuronic acid/glucose consists mainly of positive ions with a fragmentation pattern from the B-chain. The spectrum of reduced insulin using a nitrocellulose matrix gives B-chain ions but glucose/glucuronic acid gives A-chain ions in both the positive and negative ion spectra. The fragmentation patterns of the A-chain and B-chain ions are sensitive to the nature of the matrix. An extensive negative ion A-chain fragmentation pattern was observed with glutamate ions serving as the charge centers. The reasons for the behavior of the A- and B-chain fragmentation patterns in these matrices is not clear.     

Analysis of insulin preparations by reversed-phase high-performance liquid chromatography

A reversed-phase high-performance liquid chromatographic system is described for the rapid and complete separation of bovine and porcine insulin from their readily formed monodesamido derivatives under isocratic conditions in the presence of the ion-pairing agent cetrimide. The system is suitable for the direct analysis of formulations of insulins of mixed bovine and porcine origin, and gives satisfactory results with a number of readily available commercial packings. Human insulin is not resolved from porcine in this system, but an alternative system allows the complete separation of all three insulins and their monodesamido derivatives, although acceptable peak shapes were obtained only on a limited number of packings.