Injectable porous hydroxyapatite microparticles as a new carrier for protein and lipophilic drugs.

Hydroxyapatite (Ca10 (PO4)6(OH)2) is a biodegradable material that forms a major component of bones and teeth. We prepared injectable spherical porous hydroxyapatite microparticles (SP-HAp) as a drug carrier by the spray-drying method. We then examined the usefulness of SP-HAp as a carrier for drugs such as interferon alpha (IFNalpha), testosterone enanthate (TE), and cyclosporin A (CyA). SP-HAp had an average diameter of 5 mum and a porosity of approximately 58%. It could be injected subcutaneously through a 27-gauge needle. SP-HAp was observed to be biodegradable. The speed of degradation of SP-HAp could be regulated by altering the calcination temperature. IFNalpha was adsorbed well to SP-HAp particles, but INFalpha was released faster from the particles, than the particles could degrade in both in vitro and in vivo experiments. Addition of human serum albumin and zinc (reinforcement) to IFNalpha-adsorbed SP-HAp caused marked prolongation of release in vivo. The in vivo release of testosterone enanthate and CyA from SP-HAp preparation, which was easily injectable, was similarly prolonged to that from the oil preparation. In conclusion, the SP-HAp seems to be useful as a biodegradable and subcutaneously injectable drug carrier. It is suggested that the reinforcement of the SP-HAp is very effective on the sustained release of drugs.

Role of zinc in formulation of PLGA/PLA nanoparticles encapsulating betamethasone phosphate and its release profile.

The purpose of this study was to develop poly(D,L-lactic/glycolic acid) (PLGA) or poly(D,L-lactic acid) (PLA) nanoparticles of less than 200 nm in diameter that encapsulated water-soluble corticosteroid derivatives for sustained release and targeting to inflammatory sites. Nanoparticles were prepared with PLGA (or PLA), zinc, betamethasone phosphate and surfactant by an oil-in-water solvent diffusion method. With this method, the efficiency of encapsulating betamethasone phosphate in the nanoparticles and the particle size were significantly affected by various factors, such as the concentration of PLGA (or PLA) and the amount of zinc added. Nanoparticles ranging from 80 to 250 nm in diameter could be prepared, with a maximum betamethasone phosphate content of 8% (w/w). Betamethasone phosphate was gradually released from the nanoparticles in diluted serum, and the release rate depended on the glycolic/lactic acid ratio and on the molecular weight of PLGA or PLA. Betamethasone was gradually released over at least 8 days from murine macrophages that had internalized betamethasone phosphate-encapsulated nanoparticles in vitro, and the rate of release was slower than from nanoparticles prepared without zinc. These results suggest that zinc increases the efficiency of encapsulating betamethasone phosphate in nanoparticles and also promotes sustained release of betamethasone phosphate from the nanoparticles.

Optimum formulation for sustained-release insulin.

Our aim was to prepare an optimum formulation for a sustained-release preparation of insulin using biodegradable polymer composed of co-poly(D,L-lactic/glycolic) acids (PLGA)(L/G ratio: 50/50). Various kinds of PLGA microcapsules containing 3% insulin were administered subcutaneously (250 U/kg) as a single dose to rats with streptozotocin-induced diabetes, and plasma insulin levels were monitored. The following results were obtained. (1) Glycerin and water were suitable additives to prepare a reproducible injectable formulation. (2) Addition of zinc compounds was essential to diminish rapid insulin release and six-fold molar excess of ZnO to insulin was desirable. (3) The size of insulin particles showed the following order: human insulin > lyophilized human insulin > Zn-free human insulin. Zn-free insulin was similar to lyophilized insulin with respect to control of rapid release, so a smaller particle size was essential. (4) The size of the microcapsules also affected the release of insulin. With larger microcapsules (approximately 30 microm), there was gradual release and a significant second phase of insulin release, while smaller microcapsules did not allow sustained release. Some variation in microcapsule size contributed to more constant and sustained release. (5) Based on the insulin release profile in vivo, a suitable molecular weight for PLGA was around 6000. The biological activity of insulin extracted from the formulation was similar to that of normal insulin. These experiments allowed us to prepare a desirable sustained-release insulin formulation.

A novel insulin formulation can keep providing steady levels of insulin for much longer periods in-vivo.

We have recently succeeded in preparing insulin-loaded microcapsules that release the insulin in a strictly controlled manner with little initial rapid release in-vitro or in-vivo. We show here the superiority of the best formulation prepared with co-poly(D,L-lactic/glycolic) acids (PLGA) (mean MW 5800, L/G ratio 50:50) with a main diameter of 15 approximately 30 microm in-vivo. When 3.2 % insulin-loaded PLGA microcapsules were subcutaneously given as a single dose to streptozotocin-induced hyperglycaemic rats (250 U kg(-1)), plasma insulin levels gradually increased and constant levels (30.3-94.1 microU mL(-1)) were sustained. Rats receiving the formulation once a week showed not only steady plasma insulin levels, but also gained weight at a similar speed to normal rats. Meanwhile, daily treatment with Humulin U (25 U kg(-1)) caused a transient high insulin level (2723.9 microU mL(-1) at 1 h) in plasma, but the body weight of the rats was little changed. A pharmacological study in female Cynomolgus monkeys also revealed that the microcapsular formulation provided a flat release of insulin for longer periods and showed no immunogenic activity. In the near future, therefore, this insulin formulation could become very beneficial as a provider of basal insulin levels for insulin-dependent diabetic patients.

A novel sustained-release formulation of insulin with dramatic reduction in initial rapid release.

To ensure a strictly controlled release of insulin, a preparation method for insulin-loaded microcapsules was designed. Microcapsules were prepared with an injectable, biodegradable polymer composed of co-poly(D,L-lactic/glycolic) acids (PLGA) (mean molecular weight 6600, LA/GA ratio 50:50). Morphological examination using scanning electron microphotography demonstrated spherical particles with a main diameter of 15-30 microm. When 3% insulin-loaded PLGA microcapsules were administered subcutaneously as a single dose (250 U/kg) to streptozotocin-induced hyperglycemic rats, plasma insulin levels increased and were sustained at levels showing hypoglycemic effects. When glycerin, ethanol, or distilled water was used throughout the preparation procedure, the resultant microcapsules dramatically reduced the initial burst. The formulation in which glycerin was added to an oil phase containing PLGA, insulin, and ZnO increased plasma insulin levels to 86.7, 108.4, and 84.9 microU/ml at 1, 2, and 6 h, respectively. The levels remained at 36.2-140.7 microU/ml from day 1 to day 9. The AUC(0-24 h)/AUC(0-336 h) ratio was calculated to be 9.7%. The formulation prepared without additives gave such a rapid insulin release that animals receiving it became transiently hypoglycemic.

Prevention of bone loss in ovariectomized rats by pulsatile transdermal iontophoretic administration of human PTH(1-34).

Serum human parathyroid hormone (1-34)[hPTH(1-34)] levels and the anabolic effect of hPTH(1-34) were compared after administration using multiple pulses of iontophoresis or subcutaneous (sc) intermittent injections to ovariectomized (OVX) Sprague Dawley rats. Triple-pulse iontophoretic administration of hPTH(1-34) (doses: 40-400 microg/patch), achieved by repeated 30-min applications of a 0.1 mA/cm(2) current separated by 45-min rest intervals, produced three sharp peaks in the serum hPTH(1-34) level in response to application of the current. Each peak appeared at the end of the 30-min current application period and was proportional to the hPTH(1-34) dose. Compared with once-daily sc injections (7 pulses/week), triple-pulses iontophoretic administered 3 times/week (9 pulses/week) for 4 weeks produced dose-related increases in the bone mineral density (BMD) of the distal 1/3 femur. For the sc administration, the relative BMD values using the vehicle injection as a reference standard for 1, 5, and 25 microg/kg/day were 104, 114, and 121%, respectively. For iontophoretic administration, the relative BMD values using the placebo patch as a reference standard for 40, 120, and 400 microg/patch were 104, 110, and 116%, respectively. The increase in the BMD plotted against the area under the hPTH(1-34) serum level-time curve (AUC) over 1 week resulted in similar straight lines in the 9 pulses/week iontophoretic administration and the 7 pulses/week sc administration groups. The estimated iontophoretic dose giving an equivalent BMD to once-daily sc administration at 5 microg/kg/day was 120 microg/patch. These findings strongly suggest that three iontophoretic pulses administered on alternate days will exert an anabolic effect equivalent to that of daily sc administration at doses giving the same weekly AUC. Furthermore, this method of administering hPTH(1-34) might enable self-medication, a useful advance in the treatment of osteoporosis.