Strontium hydroxyapatite in situ gel-forming system - a new approach for minimally invasive bone augmentation.

OBJECTIVES: To achieve ideal functional and aesthetic requirements, ridge augmentation is often required before dental implant placement. Bone augmentation (especially vertical), which normally consists of complex and invasive surgeries, still remains challenge. This study seeks to investigate the feasibility of an injectable in situ gel-forming system containing strontium hydroxyapatite (SrHA) and alginate for minimally invasive bone augmentation in a rat calvarial model. MATERIAL AND METHODS: SrHA-alginate solution was prepared by mixing SrHA powder with alginate solution (20 mg/mL) to the final concentration of 0.5% (w/v). Each animal received a 200-µL single subperiosteal injection of either SrHA-alginate solution or alginate solution. The new bone formation was assessed at 0, 4, and 8 weeks histologically and radiologically. RESULTS: The SrHA-alginate solution materials could form solid gel once injected. As such, no sutures were required to close the injection site. Significantly greater amount of new bone formation was observed in the SrHA-alginate group compared with the alginate group both by micro-CT and by histological section. The newly formed bone in the SrHA-alginate group originated both from the underlying original bone and from the elevated periosteum. A 2.3-fold increase of the vertical bone height was observed in the SrHA-alginate group compared with 1.3-fold increase in the alginate group. CONCLUSIONS: Rat calvarial bone augmentation was achieved by a single subperiosteal injection of SrHA-alginate solution without any administration of stem cells or growth factors. The in situ gel-forming material may hold potential therapeutic benefits for local bone augmentation in a minimally invasive manner.

Effects of bead size and polymerization in PMMA bone cement on vancomycin release.

Polymethylmethacrylate (PMMA) bone cement loaded with antimicrobial agents is used for the treatment and prevention of infections in orthopaedic surgery. The use of antimicrobial-loaded bone cement allows for high local doses while avoiding systemic toxicity. The release of vancomycin (VCM) from bone cement has been reported. However, the exact mechanism behind the release is unknown. We studied the influence of bead size and polymerization time on elution, and considered the release mechanism for VCM. We used CMW Endurance Bone Cement. Cements were prepared by mixing 6 g of VCM with 40 g of polymer, and then 10 g of liquid monomer was added. We kneaded and shaped the preparation into spheres containing 10.7 w/w% VCM. We measured the release of VCM from PMMA beads of three different sizes. Average weights of the beads were 0.96 g (SB) (n = 6), 2.86 g (MB) (n = 2) and 5.65 g (LB) (n = 1). Additionally, we studied beads made with different polymerization times. The polymerization time was taken as the period from the making of the beads until the start of the study, and was 15 min (B15), 20 min (B20), 60 min (B60) or 180 min (B180). The release of VCM showed a bimodal curve with a high initial release followed by a sustained release. Regarding the size of the beads, SB released 7.2%, MB released 4.3% and LB released 3.1%. Regarding polymerization time, B15 released 10.0%, B20 released 6.5%, B60 released 6.3% and B180 released 4.3%, respectively. The release of VCM from PMMA beads was influenced by bead size and polymerization time. Those beads which were smaller and had a shorter polymerization time released more VCM. Total pore volume of beads that polymerization time was 30 min after drug-release test was 1.33 times grater than that of control beads that polymerization time was one week before drug-release test. This suggested that the short polymerization time caused the beads to leak more VCM. We proposed a model with four kinds of the dissolution from bone cement. (A) Dissolution from drug particles on the cement surface. This type shows the burst effect of release curve. (B) Dissolution from micropores near the cement surface. It is responsible the grater part of the curve based on Higuchi's equation. (C) Dissolution from ink-bottle-neck-type micropores. It causes a release based on a non-Higuchi's equation. (D) No dissolution from Encapsulation micropores.It can be concluded that the release of VCM from bone cements is controlled by a combination of surface area and polymerization time. PMMA beads loaded with VCM should be used carefully in orthopaedic surgery, taking into consideration the influence of bead size and polymerization time.

A comparative study of the metastable equilibrium solubility behavior of high-crystallinity and low-crystallinity carbonated apatites using pH and solution strontium as independent variables.

UNLABELLED: Using solution strontium and pH as independent variables, the metastable equilibrium solubility (MES) behavior of two carbonated apatite (CAP) samples has been examined, a high-crystallinity CAP (properties expected to be similar to dental enamel) and a low-crystallinity CAP (properties expected to be similar to bone mineral). CAP samples were prepared by precipitation/digestion: (CAP A: high-crystallinity, 1.3 wt% CO3, synthesized at 85 degrees C; CAP B: low-crystallinity, 6.4 wt% CO3, synthesized at 50 degrees C). Baseline MES distributions were determined in a series of 0.1 M acetate buffers containing only calcium and phosphate (no strontium) over a broad range of solution conditions. To assess the influence of strontium, MES profiles were determined in a similar fashion with 20, 40, 60, and 80% of the solution calcium being replaced on an equal molar basis by solution strontium. To determine the correct function governing CAP dissolution, ion activity products (IAPs) were calculated from the compositions of buffer solutions based on the hydroxyapatite template (Ca(10-n)Sr(n)(PO4)6(OH)2 (n = 0-10)) and the calcium/hydroxide deficient hydroxyapatite template (Ca(9-n)Sr(n)(HPO4)(PO4)5OH (n = 0-9)). FINDINGS: (a) for CAP A, at high solution strontium/calcium ratios, the MES profiles were essentially superimposable when the solution IAPs were calculated using the stoichiometry of Ca6Sr4(PO4)6(OH)2 and for CAP B by a stoichiometry of Ca7Sr2(HPO4)(PO4)5OH; (b) for CAP A, at low strontium/calcium ratios, the stoichiometry yielding MES data superpositioning was found to be that of hydroxyapatite and for CAP B, that of calcium/hydroxide deficient hydroxyapatite. When other stoichiometries were assumed, good superpositioning of the data was not possible.

Antibiotic delivery system using bioactive bone cement consisting of Bis-GMA/TEGDMA resin and bioactive glass ceramics.

A novel drug delivery system containing cephalexin (CEX) as a model drug using a new bioactive bone cement consisting of 15% bisphenol-alpha-glycidyl methacrylate (Bis-GMA), 15% triethylene-glycol dimethacrylate (TEGDMA) resin and 70% apatite- and wollastonite-containing glass-ceramic (A-W GC) powder was investigated. A-W GC powder containing CEX powder hardened within 5 min after mixing with Bis-GMA/TEGDMA resin, and furthermore its compressive strength was expected to be higher than that of polymethylmethacrylate cement. In vitro CEX release from bioactive bone cement pellets in a simulated body fluid at pH 7.25 and 37 degrees C continued for more than 2 weeks. The drug release rate increased with increasing amount of CEX in the mixture. All of the drug release profiles followed the Higuchi equation at the initial stage, but not at later stages. As hydroxyapatite was precipitated out on the cement surface, the drug release rate decreased. These results suggest that the CEX release rate from bioactive bone cement could be controlled by varying the amount of drug in the cement system.

Solid dosage form preparations from oily medicines and their drug release. Effect Of degree of surface-modification of silica gel on the drug release from phytonadione-loaded silica gels.

A surface-modified silica gel was produced to improve the surface affinity to an oily medicine, phytonadione (VK1). The effect of the degree of surface modification of the silica gel on the drug release behavior from the silica porous matrix was investigated. The silica gels were surface-modified using the silan coupling agent, 3-methacryloxypropyltrimethoxysilane (C7), octadecyltriethoxysilane (C18), or 3,3,3-trifluoropropyltrimethoxysilane (F3). A mixture of VK1 solution and surface-modified silica gel was evaporated under reduced pressure at room temperature, then the resulting powder was dried in vacuo. The degree of surface modification was evaluated based upon elementary analysis. The dissolution profiles of the samples were investigated in Japanese Pharmacopoeia XII, 1st fluid buffer (pH 1.2, 37+/-0.5 degrees C) containing 1.5% sodium lauryl sulfate. The FT-IR spectra of VK1-loaded surface-modified silica gels suggested that the amount of hydrogen-bonded VK1 with the silanol group on the gel surface decreased with increasing hydrophobicity of the silica gel. Since the modified group was rotating on the silica gel surface, and inhibited the adsorption of VK1 to the surface, the attractive molecular interaction between VK1 and the silica gel surface might decrease with increasing length of the modified functional group. However, the characteristics of the affinity of VK1 to the functional groups significantly differed among the groups. The VK1 release from the modified silica gels was initially rapid, slowed markedly after 1 h, and continued for more than 24 h. The amount of VK1 released from the modified surface silica gels by C7, C18 or F3 increased with increasing density of the surface modification group. The mean drug release moment (MDT) decreased with an increase in surface-modified group density.

Effect of geometrical cement size on in vitro and in vivo indomethacin release from self-setting apatite cement.

The relationship between in vitro and in vivo indomethacin (IMC) release from a self-setting bioactive apatite cement and cement size were investigated. Differently sized apatite cements (total weight, 500 mg); either 64 of the small size (2 mm diameter x 2 mm thickness), sixteen of the medium size (4 mm x 2 mm) or one of the large size (15 mm x 2 mm) were obtained from cement bulk powder containing tetracalcium phosphate, dicalcium phosphate dihydrate and hydroxyapatite. In vitro IMC release from the 1, 2 and 5% drug-loaded apatite cement systems in simulated body fluid (SBF) (pH 7.25) at 37 degrees C increased with increasing concentrations of IMC and with decreasing geometrical size of the cement. The plots of in vitro IMC release per unit area against the square root of time increased with increasing IMC concentrations, but not with decreasing geometrical size of the cement. After subcutaneous (s.c.) implantation of differently sized 1% IMC-loaded cements in male Wistar rats, the plasma IMC concentration and the area under the curve increased with decreasing cement diameter. The in vivo IMC release profiles of the cement were deconvoluted from the plasma IMC profiles after s.c. administration of IMC solution. The plots of in vivo IMC release per unit area against the square root of time suggested that the initial release from all 1% drug-loaded cements was very rapid, slowed after one day, but continued for over two weeks. The relationship between the in vitro release in SBF and the in vivo release in rats of IMC-loaded cements was linear.

Initial dissolution rate studies on dental enamel after CO2 laser irradiation.

The influence of CO2 laser irradiation on the dissolution behavior of human dental enamel has been investigated. Human enamel was irradiated by a continuous-wave CO2 laser at 10.6 microns and initial dissolution rates (IDRs) were measured in 0.1 mol/L acetate buffer, pH = 4.5, both with and without calcium and/or phosphate common ion, by means of a rotating disk assembly. The effects of (1-hydroxyethylidene) bisphosphonic acid (EHDP), fluoride (F), and dodecylamine HCl (DAC) at various levels upon the IDR were also determined. All of the findings were consistent with the hypothesis that CO2 laser irradiation converts dental enamel to hydroxyapatite (HAP) possessing site #2 character (Yamamoto et al., 1986). The dissolution driving force function, KHAP = aCa10aPO4(6)aOH2, was found to have a value of 10(-129.9) after being lased, as compared with 10(-121.4) before being lased. The IDR values for EHDP (3 mmol/L) and DAC (3 mmol/L) were essentially zero as expected for site #2 HAP. For solution F, the deduced dissolution driving force function, KFAP = aCa10aPO4(6)aF2 was 10(-128.6) after being lased as compared with 10(-116.3) before being lased. These results all support the hypotheses (1) that laser irradiation may convert the surface of human dental enamel to an apatite of significantly lower effective solubility (i.e., site #2 HAP) than that of unlased enamel; and (2) that there is significant synergism between laser treatment and these chemical dissolution rate inhibitors (again consistent with site #2 HAP). Simple model calculations indicate that, in both the presence and absence of fluoride, these laser-induced changes in the driving force for dissolution should dramatically lessen the susceptibility of enamel to the types of acid challenge that might be encountered in the mouth.

Heat-treatment-induced reduction in the apparent solubility of human dental enamel.

Holcomb and Young (1980) have shown a significant increase in human dental enamel (HE) structural order resulting from heat treatment in the temperature range of from 275 to 400 degrees C. Also, previous work in our laboratory had shown dramatic decreases in the initial dissolution rates (IDRs) of both carbonated apatite (CAP) heated at similar temperatures (from 300 to 500 degrees C) and HE exposed to CO2 laser irradiation for which calculated surface temperatures were in this same range. We hypothesize that thermal treatment shifts the apparent solubility distribution profile of HE toward lower apparent solubilities, paralleling the observed increased in crystal structural order and the decrease in IDRs. Powdered HE was heated in a furnace at temperatures ranging from 150 to 500 degrees C for 24 hours. The apparent solubility distributions of both heated and unheated HE powders were measured by equilibration for 24 hours in a series of partially saturated solutions simulating various amounts of HE dissolved in a pH 4.5 dissolution medium. The apparent solubility distribution for the unheated HE showed a peak at KHAP [the ion activity product based on the Ca10(PO4)6(OH)2 stoichiometry] of 10(121.0). Heat treatment shifted the apparent solubility distribution to lower solubilities. The peak KHAP values were approximately 10(124.8) at 200 degrees C; approximately 10(127.8) at 300 degrees C; and approximately 10(-129.1) from 400 to 500 degrees C. This approximately 8 orders of magnitude decrease in KHAP for HE heated at from 400 to 500 degrees C correlates with the previously observed reduction in the IDR driving force for laser-treated HE experiencing a similar surface temperature.(ABSTRACT TRUNCATED AT 250 WORDS)

Purification and some properties of neuraminidase isolated from the culture medium of oral bacterium Streptococcus mitis ATCC 9811.

Neuraminidase acting on the salivary bacterial agglutinating factor was isolated and purified from the culture medium of Streptococcus mitis ATCC 9811. The molecular weight and the isoelectric point of the enzyme were determined to be 42,000 and a pH of 4.6, respectively. The enzyme showed high activity against human glycoprotein substrates, especially the salivary bacterial agglutinating factor.

In vivo liver-directed gene transfer in rats and pigs with large anionic multilamellar liposomes: routes of administration and effects of surgical manipulations on transfection efficiency.

Conventional large (500-800 nm) multilamellar liposomes encapsulating DNA have been used in vivo as gene vectors into rat and pig liver. By using the intraportal vein route, high dose DNA (10 mg/kg) provided low efficiency and transient luciferase gene expression in the liver. This gene expression was, however, increased by liver resection (> 50%), ischemia (20 min) or orthotopic transplantation. As evidenced by histochemical analysis of beta-galactosidase expression, the gene transfection mainly ensued in Kupffer cells, but spleen and lung were contaminated. In comparison, injection into the bile duct of even 25-fold lower dose of liposome-encapsulated DNA (0.4 mg/kg) produced higher (100-fold) and long-lasting (during 6 days, at least) luciferase expression in rat liver. The gene expression was restricted to the liver and enhanced by liver resection. By this route, transgene-expressing cells were mainly hepatocytes. A treatment with colchicine prior to the administration of the vector allowed the persistence of relative high gene expression for at least 7 days. In pigs, qualitatively similar, but quantitatively less efficient gene expression was obtained by either the portal vein or the bile duct route. These results indicate that intrabile duct route might render large non-viral vectors applicable to gene transfer into the hepatocytes. The efficiency of liposome-mediated gene transfer into the liver can be increased by liver resection, ischemia or transplantation performed before DNA injection.

Hygroscopic stability and dissolution properties of spray-dried solid dispersions of furosemide with Eudragit.

Spray-dried solid dispersions of furosemide-Eudragit RS100 and RL100 (ethyl methacrylate chlorotrimethylammoniumethyl methacrylate copolymer) were studied to determine their stability at 45 degrees C and 0, 44, and 75% relative humidity and their dissolution characteristics. The crystallization rates of the solid dispersions were measured by X-ray powder diffractometry and calculated on the basis of the Jander equation to estimate physicochemical stability in the presence of water vapor. The stability of the solid dispersion depended on the kind of acrylic resins and the drug content. The solid dispersion with a drug: Eudragit RL100 ratio of 1:3 was the most stable at 75% relative humidity. The dissolution profiles of the solid dispersions were measured in a pH 6.8 buffer at 37 degrees C. The equilibrium drug concentration of the solid dispersions were estimated from the dissolution profiles after 24 h. The equilibrium drug concentration decreased with an increasing Eudragit quaternary ammonium group: furosemide molar ratio. The equilibrium drug concentrations of the solid dispersions with > 0.3 mol of the quaternary ammonium group were approximately 0. Thus, the drug was completely adsorbed on Eudragit.

A novel skeletal drug delivery system using self-setting calcium phosphate cement. 2. Physicochemical properties and drug release rate of the cement-containing indomethacin.

A novel drug delivery device based on a self-setting bioactive cement formed from tetracalcium phosphate and dicalcium phosphate has been developed and tested in vitro with indomethacin as a model drug. Equimolar mixtures of the calcium phosphate powders containing 2 and 5% of indomethacin were transformed into a hydroxyapatite after being mixed with a dilute phosphoric acid solution. X-ray diffraction and differential scanning calorimetry results suggested that indomethacin transformed into an amorphous form in the pores of the cement matrix as it hardened. In vitro drug release from cement pellets into a 0.1 mol/L phosphate buffer at pH 7.40 and 37 degrees C continued for > 3 weeks. Release from 2 and 5% drug-loaded cements followed the Higuchi model equation. The drug release profiles of 5% drug-loaded cements with different thicknesses (0.5, 1.0, and 1.5 g) overlapped up to 90% drug release, indicating that the drug concentration gradient in the pore was independent of the thickness of the cement as expected from the model equation.

A novel skeletal drug-delivery system using self-setting calcium phosphate cement. 3. Physicochemical properties and drug-release rate of bovine insulin and bovine albumin.

A novel drug-deliver device based on a self-setting bioactive cement formed from tetracalcium phosphate and dicalcium phosphate has been developed and tested in vitro using bovine insulin and bovine albumin as model polypeptide drugs. Equimolar mixtures of the calcium phosphate powders containing bovine insulin and bovine albumin were transformed into a hydroxyapatite cement after being mixed with a dilute phosphoric acid solution. X-ray diffraction and FT-IR spectra results suggested that the raw materials transformed into lower crystallinity of hydroxyapatite as it hardened. In vitro drug release from cement pellets into a 0.1 mol/L phosphate buffer at pH 7.40 and 37 degrees C continued for more than 3 weeks. Release from the drug-loaded cements followed the Higuchi model equation.

A novel skeletal drug-delivery system using self-setting calcium phosphate cement. 4. Effects of the mixing solution volume on the drug-release rate of heterogeneous aspirin-loaded cement.

The effect of the mixing solution volume was investigated on the in vitro drug-release rate of a novel drug-delivery device based on a self-setting bioactive calcium phosphate cement containing aspirin as a model drug. Equimolar mixtures of metastable calcium phosphate powders containing various proportions (3-40 w/w %) of seed hydroxyapatite crystals transformed into hydroxyapatite after being mixed with dilute phosphoric acid. The drug release from cement pellets in vitro into a 0.1 mol/L phosphate buffer at pH 7.40 and 37 degrees C by the rotating disk method continued for more than 1 week. The drug-release rate from the cement increased with increasing volumes of mixing solution. The relationship between the liquid/powder ratio and the porosity of the cement was a straight line, indicating that the cement porosity depended on the amount of the mixing solution, but was independent of the amount of seed crystals. Drug release from the cement followed the modified Fick's law, with the rate increasing with the amount of mixing solution, since the porosity depended on the amount. The tortuosity of the cements was estimated from the modified Fick's equation, and the relationships between the drug release rate and the tortuosity of the pore in the drug-loaded cement in the plots were nonlinear. The results suggested that the drug-release rates from the cement were controlled by the drug diffusion in the pores.

Calculation of intercrystalline solution composition during in vitro subsurface lesion formation in dental minerals.

Applications of a novel technique to calculate intercrystalline solution composition during enamel demineralization are presented. Bovine tooth enamel blocks and carbonated apatite (CAP) compressed disks were demineralized in an in vitro subsurface lesion system. The demineralization medium was a 0.1 M acetate buffer at pH 4.5, containing calcium, phosphate, and fluoride (0.5 ppm). Mineral samples were demineralized for various times, and fluoride profiles and mineral density profiles of these samples were determined by electron microprobe and X-ray microradiography, respectively. A model independent data analysis (MIDA) technique uses these data along with the differential equations for mass transfer and permits calculation of the local intercrystalline solution composition profiles inside the porous mineral matrix as functions of time and position. The invariance in diffusivity with time as calculated in the analysis was taken as an indicator of the physical reasonableness of the method. Current outcomes suggest that it is the sharp gradient of fluoride concentration in the intercrystalline solution which causes the formation of subsurface lesions. Since the driving force for mineral dissolution is a function of solution composition, a gradient of this driving force is consequently formed. Using a compressed disk of carbonated apatite powder as a model for block enamel excluded the possibility of the existence of a gradient of mineral composition which could also cause a gradient of the driving force for mineral dissolution. An FAP surface complex hypothesis is consistent with the current view that fluoride in the intercrystalline solution has a stronger inhibition effect on the dissolution of mineral than does fluoride in the mineral phase. With the help of the MIDA technique, calculated results indicate that the mechanism of the formation of subsurface lesions is dynamically controlled by the intercrystalline solution composition.

Self-setting hydroxyapatite cement: a novel skeletal drug-delivery system for antibiotics.

A novel approach using a self-setting hydroxyapatite (HAP) cement as a skeletal drug-delivery system has been proposed to solve the problem of delivering drugs to skeletal tissue at sufficiently high local concentrations for desirable therapeutic effects. HAP cements loaded with antibiotics can be formed in situ and can be used as bonding materials between bone and prostheses, as well as drug-release devices. The cement also possesses sufficient mechanical strength to be a potential bone grafting material. Using cephalexin and norfloxacin as model drugs, we observed continuous in vitro release profiles of these compounds from cement pellets loaded 0.9-4.8% by weight with one of the drugs. This drug-release pattern correlated well with the Higuchi model. This hydroxyapatite cement drug-delivery system can be applied in the treatment of osteomyelitis and infected compound fractures.

A novel skeletal drug delivery system using a self-setting calcium phosphate cement. 5. Drug release behavior from a heterogeneous drug-loaded cement containing an anticancer drug.

A novel drug delivery device based on a self-setting bioactive calcium phosphate cement formed from tetracalcium phosphate and dicalcium phosphate has been developed and tested in vitro using the anticancer agent 6-mercaptopurine (6-MP) as a model compound. X-ray diffraction results suggest that equimolar mixtures of the calcium phosphate salts were transformed into hydroxyapatite after being mixed with a dilute phosphoric acid solution, even in the presence of various amounts of 6-MP powder. The inclusion of 6-MP in the reaction mixture resulted in the formation of a homogeneous drug-containing cement. Alternatively, the drug was loaded after cement formation to produce a heterogeneous drug-containing pellet. In vitro drug release from both the homogeneous and heterogeneous drug-loaded cement pellets into simulated body fluid (pH 7.25, 37.0 degrees C) was measured using the rotating-disk method. Release from the homogeneous 5% drug-loaded cements did not obey the Higuchi equation. The release rate from the heterogeneous drug-loaded cements of different thicknesses (1, 2, and 3 mm) was a function of thickness, indicating that release kinetics could be controlled by the design of the cement formulation.

A novel skeletal drug delivery system using self-setting calcium phosphate cement. 7. Effect of biological factors on indomethacin release from the cement loaded on bovine bone.

The use of self-setting bioactive calcium phosphate cement containing indomethacin as a model drug in bovine bone was investigated by means of an in vitro drug release test, mercury porosimetry, and scanning electron microscopy (SEM). Calcium phosphate cements containing 2 and 5% indomethacin after being mixed with dilute phosphoric acid were applied to defect sites and the medullary cavity of bovine bone and transformed into hydroxyapatite. The in vitro drug release from the cement loaded on the defect site into a simulated body fluid (SBF) containing 2.5 mM Ca2+ and 1.0 mM HPO4(2+) or 0.1 M phosphate buffer at pH 7.25 and 37 degrees C continued for more than 3 weeks. The release profiles of the drug-loaded cements in phosphate buffer were linear using the Higuchi plot; however, that was not the case for SBF. The drug release in SBF was much lower than that in phosphate buffer. The total pore volume of the cement after the drug release test in SBF was lower than its initial value. However, the pore size of 0.1-0.01 microns after drug release in phosphate buffer was higher than that seen in SBF. The micropore distribution results suggested that hydroxyapatite crystallized from SBF and the pore volume in the cement decreased after drug release. However, in phosphate buffer it appeared to dissolve. The SEM observations for cements loaded on the bone after drug release in phosphate buffer suggested that there was a boundary layer between the cement and natural bone, but this was not the case in SBF, where the cement bonded with the natural bone. The drug release rates from the cement-loaded bone were significantly higher than those from cement loaded on the dissolution holder. The results suggested that cement formation and drug release were affected by the presence of protein from natural bone. The drug release rates from the cement loaded on the defective bone were slower than those from the medullary cavity.

Cleavage action of a trypsin-like protease from Bacteroides gingivalis 381 on reduced egg-white lysozyme.

Soluble reduced lysozyme was extensively digested by a trypsin-like protease purified from the culture supernatant of the bacterium. The digestion peptides were separated and purified by reversed-phase high-performance liquid chromatography, and were subjected to amino acid analysis. The fragments were identified by their amino acid composition, and the cleavage sites in the lysozyme chain were determined. Like mammalian trypsin, the enzyme from B. gingivalis split peptide bonds non-specifically at carboxyl sides of internal arginine and lysine residues, but the lysine present at the amino terminus of the lysozyme chain was not released. In addition, the enzyme cleaved the peptide linkage at the amino side of lysine and bonds between leucine-glycine, alanine-leucine and leucine-serine. Thus the trypsin-like protease from B. gingivalis has some cleavage actions on lysozyme different from those of mammalian trypsin.

Degradation of human secretory immunoglobulin A by protease isolated from the anaerobic periodontopathogenic bacterium, Bacteroides gingivalis.

This bacterium is implicated in periodontal diseases of human adult type. Secretory immunoglobulin A (IgA) purified from human colostrum (HC-IgA) was incubated with Bacteroides gingivalis cells or protease isolated from the culture supernatant of B. gingivalis; the digestion of IgA was determined by immunoelectrophoresis and sodium dodecyl sulphate-polyacrylamide gel electrophoresis. B. gingivalis cells almost completely degraded HC-IgA; protease isolated from the culture supernatant cleaved both HC-IgA and secretory IgA in human parotid saliva. Thus by degradation of IgA, the protease may mediate in part the periodontopathogenic role of B. gingivalis by decreasing the oral defence mechanism.