Plasma Bupivacaine Research Articles

High-Performance Liquid Chromatography Using UV Detection for the Simultaneous Quantification of Ropivacaine and Bupivacaine in Human Plasma

A specific high-performance liquid chromatography assay coupled with UV detection has been developed and validated for the simultaneous determination of ropivacaine and bupivacaine in human plasma. A liquid-liquid back extraction procedure was used to increase specificity, and very good and consistent recoveries were obtained: 93%-95% for ropivacaine and 90%-96% for bupivacaine. The lowest limit of quantification was 4 and 8 ng/mL for ropivacaine and bupivacaine, respectively. The method was sensitive, reproducible (coefficient of variation <or= 6.9%), and accurate (96%-102%) and was applied successfully to study the pharmacokinetics of ropivacaine and bupivacaine in a perioperative context.

Midazolam dose for loss of response to verbal stimulation during the unilateral or bilateral spinal anesthesia

We have conducted this study to investigate whether unilateral or bilateral spinal anesthesia with bupivacaine induces different sensitivity to intravenous (i.v.) midazolam for sedation. Forty-two patients undergoing various elective unilateral lower extremity surgeries were allocated into two groups: (1) unilateral spinal anesthesia group (Group US, n=21; heavy bupivacaine 5 mg/ml, 9 mg) and (2) bilateral spinal anesthesia group (Group BS, n=21; heavy bupivacaine 5 mg/ml, 9 mg). One milligram of midazolam was injected i.v. at 30-s intervals until the patients did not respond to the hand grasp test beginning 15 min after spinal anesthesia. The concentration of plasma bupivacaine was evaluated every 15 min for the first 75 min after the start of the spinal anesthesia, and the bispectral index was monitored continuously. The mean venous plasma concentration of bupivacaine was not significantly different between Group US and BS. The dose of midazolam required to abolish responses to verbal commands was significantly lower in Group BS (mean 5.9+/-1.2 mg) vs. Group US (mean 9.0+/-1.4 mg). A higher dosage of midazolam is required for loss of response to verbal stimulation during unilateral spinal anesthesia than during bilateral spinal anesthesia.

A Comparison of the Combination of Epinephrine and Vasopressin with Lipid Emulsion in a Porcine Model of Asphyxial Cardiac Arrest After Intravenous Injection of Bupivacaine

In a porcine model, we compared the effect of the combination of vasopressin/epinephrine with that of a lipid emulsion on survival after bupivacaine-induced cardiac arrest. After administration of 5 mg/kg of a 0.5% bupivacaine solution i.v., ventilation was interrupted for 2 +/- 0.5 (mean +/- SD) min until asystole occurred. Cardiopulmonary resuscitation (CPR) was initiated after 1 min of untreated cardiac arrest. After 2 min of CPR, 10 animals received, every 5 min, either vasopressin combined with epinephrine or 4 mL/kg of a 20% lipid emulsion. Three minutes after each drug administration, up to three countershocks (4, 4, and 6 J/kg) were administered; all subsequent shocks with 6 J/kg. Blood for determination of the plasma bupivacaine concentration was drawn throughout the experiment. In the vasopressor group, all five pigs survived, whereas none of five pigs in the lipid group had restoration of spontaneous circulation (P < 0.01). There was no significant difference between groups in the plasma concentration of total bupivacaine. In this model of a bupivacaine-induced cardiac arrest, the vasopressor combination of vasopressin and epinephrine compared with lipid emulsion resulted in higher coronary perfusion pressure during CPR and survival rates.

Estimated Plasma Bupivacaine Concentration After Single Dose and Eight‐Hour Continuous Intra‐Articular Infusion of Bupivacaine in Normal Dogs

To estimate maximum plasma concentration (C(max)) and time to maximum plasma (t(max)) bupivacaine concentration after intra-articular administration of bupivacaine for single injection (SI) and injection followed by continuous infusion (CI) in normal dogs. Cross-over design with a 2-week washout period. Healthy Coon Hound dogs (n=8). Using gas chromatography/mass spectrometry, canine plasma bupivacaine concentration was measured before and after SI (1.5 mg/kg) and CI (1.5 mg/kg and 0.3 mg/kg/h). Software was used to establish plasma concentration-time curves and estimate C(max), T(max) and other pharmacokinetic variables for comparison of SI and CI. Bupivacaine plasma concentration after SI and CI best fit a 3 exponential model. For SI, mean maximum concentration (C(max), 1.33+/-0.954 microg/mL) occurred at 11.37+/-4.546 minutes. For CI, mean C(max) (1.13+/-0.509 microg/mL) occurred at 10.37+/-4.109 minutes. The area under the concentration-time curve was smaller for SI (143.59+/-118.390 microg/mL x min) than for CI (626.502+/-423.653 microg/mL x min, P=.02) and half-life was shorter for SI (61.33+/-77.706 minutes) than for CI (245.363+/-104.415 minutes, P=.01). The highest plasma bupivacaine concentration for any dog was 3.2 microg/mL for SI and 2.3 microg/mL for CI. Intra-articular bupivacaine administration results in delayed absorption from the stifle into the systemic circulation with mean C(max) below that considered toxic and no systemic drug accumulation. Intra-articular bupivacaine can be administered with small risk of reaching toxic plasma concentrations in dogs, though toxic concentrations may be approached. Caution should be exercised with multimodal bupivacaine administration because plasma drug concentration may rise higher than with single intra-articular injection.

Plasma bupivacaine levels during continuous iliac crest wound infusion

Plasma bupivacaine levels during continuous iliac crest wound infusion

Determination of Local Anesthetics in Human Plasma by Liquid-Liquid-Liquid Microextraction Coupled with High Performance Liquid Chromatography

Liquid-liquid-liquid microextraction (LLLME) coupled with high-performance liquid chromatography was developed for the simultaneous determination of three local anesthetics, lidocaine, bupivacaine, and tetracaine in human plasma. The effects of the stirring rate, the extraction time, and the pH in the donor phase on the enrichment factor of analytes were investigated. Optimal experimental conditions, 200 μL benzene as the organic phase, 1.0 μL 0.2 mol/L HCl as the acceptor phase, 250 rpm of the stirring rate, and 45 min of the extraction time, were obtained. Under these conditions, high enrichment factors (more than 305-fold for all analytes) were achieved. The linear ranges of analytes studied were from 0.025 to 5 mg/L except tetracaine, for which the range was from 0.05 to 5 mg/L with correlation coefficients higher than 0.996. The limits of detection were 0.005 mg/L for lidocaine, 0.015 mg/L for bupivacaine, and 0.025 mg/L for tetracaine at signal-to-noise ratio of 3 ( S/ N=3). The relative standard deviations were lower than 5%. With LLLME, some substances in the human plasma, which interfered in the determination of analytes, were eliminated efficiently. The method with little solvent consumption was sensitive and suitable for determination of lidocaine, bupivacaine, and tetracaine in human plasma. This method was applied to the specimen of the patient who was treated with local anesthesia of lidocaine and bupivacaine revealed that simultaneous determination of above two local anesthetics in human plasma was possible.

Plasma Bupivacaine Concentrations Are Too Low to Explain Reduced Vecuronium Requirement with an Epidural

Plasma Bupivacaine Concentrations Are Too Low to Explain Reduced Vecuronium Requirement with an Epidural

Accelerated rehabilitation after total knee replacement

This study records the length of hospital stay of 50 total knee arthroplasty patients involved in an accelerated postoperative rehabilitation protocol, and a control group of patients undergoing routine rehabilitation. This protocol involved modifications to normal knee replacement procedure, including infiltration of bupivacaine and adrenaline to the divided tissue layers at the time of surgery, spinal anaesthesia, and mobilisation on the day of surgery. These modifications were combined with an organised multidisciplinary approach anticipating issues that may delay discharge. In addition, patients and hospital staff were encouraged to expect an earlier discharge from the hospital. The mean length of hospital stay after surgery was reduced to 3.6 (S.D. 1.0) days, from a previous departmental average of 10.5 days. The control group inpatient stay was 6.6 (S.D. 2.6) days. Plasma bupivacaine levels were found to be well within safe levels, and pain records indicated that the protocol did not cause increased levels of discomfort. American Knee Society and Oxford knee scores demonstrated good levels of knee function at 6 weeks post surgery. In addition, it was noted that no postoperative blood transfusions were required. This is regarded as a significant further benefit.

Separation and determination of bupivacaine in plasma by capillary electrophoresis with tris(2,2′‐bipyridyl)ruthenium(II) electrochemiluminescence detection

A new, rapid, selective and sensitive method is described for determination of bupivacaine by capillary electrophoresis coupled with tris(2,2'-bipyridyl)ruthenium(II) [Ru(bpy)(3)2+] electrochemiluminescence detection. The influence of parameters such as detection potential, Ru(bpy)(3)2+ concentration, buffer concentration and pH, injection time and separation voltage on separation efficiency and ECL peak intensity was systematically investigated. Under optimized conditions, the calibration curve was linear in the range 0.02-10 microg/mL. The RSD was 4.0% (n = 6). The detection limit was 3 ng/mL. The recoveries obtained were about 90%. This method was tested in the analysis of plasma samples taken from a rat after it had received bupivacaine injections.

Sensitive bioassay of bupivacaine in human plasma by liquid-chromatography-ion trap mass spectrometry

A sensitive high-performance liquid chromatography–tandem mass spectrometric (HPLC–MS–MS) method, using an ion trap spectrometer, was developed for quantitation of bupivacaine in human plasma. Bupivacaine and an internal standard (ropivacaine) were extracted in a single step from 100 μL of alkalinized plasma with diethyl-ether. The mobile phase consisted of acetonitrile with 0.1% formic acid (50:50, v/v), and was delivered at a flow rate of 0.3 mL/min. The effluent was detected by MS–MS in positive ion mode. Ionisation was performed, using an electrospray ion source, operating at 200 °C. The selected reaction monitoring transitions m/ z 289 → m/ z 140 and m/ z 275 → m/ z 126 were chosen for bupivacaine and ropivacaine, respectively. Calibration curves were linear over the concentration range of 3.90–500 μg/L with determination coefficients >0.996. The method is accurate (bias <10%) and reproducible (intra-assay and inter-assay precision <15%), with a quantitation limit of 3.90 μg/L, using only 100 μL of plasma. The high specificity and sensitivity, achieved by this fast method (total run-time <3 min), allowed the determination of bupivacaine plasma levels in pediatric patients, following epidural administration of bupivacaine.

Dexmedetomidine Decreases the Convulsive Potency of Bupivacaine and Levobupivacaine in Rats: Involvement of α2-Adrenoceptor for Controlling Convulsions

Dexmedetomidine, a highly selective alpha(2)-adrenoceptor agonist, is used in combination with local anesthetics for sedation and analgesia. We tested the hypothesis that dexmedetomidine used for sedation alters the convulsive potency of racemic bupivacaine and levobupivacaine in awake, spontaneously breathing rats. In the first experiments, male Sprague-Dawley rats were randomly divided into six groups: bupivacaine with no dexmedetomidine (bupivacaine control; BC), bupivacaine with small-dose dexmedetomidine (BS), bupivacaine with large-dose dexmedetomidine (BL), levobupivacaine with no dexmedetomidine (levobupivacaine control; LC), levobupivacaine with small-dose dexmedetomidine (LS), and levobupivacaine with large-dose dexmedetomidine (LL) (n = 10 for each group). Continuous infusion of dexmedetomidine (Groups BC and LC, 0 microg x kg(-1) x h(-1); Groups BS and LS, 3.6 microg x kg(-1) x h(-1); and Groups BL and LL, 10.8 microg x kg(-1) x h(-1)) was started after bolus injection (Groups BC and LC, 0 microg/kg; Groups BS and LS, 0.5 microg/kg; and Groups BL and LL, 1.5 microg/kg). Fifteen minutes after the start of the dexmedetomidine infusion, continuous infusion of bupivacaine (Groups BC, BS, and BL) or levobupivacaine (Groups LC, LS, and LL) at 1 mg x kg(-1) x min(-1) was started and continued until tonic/clonic convulsions occurred. Dexmedetomidine achieved significantly different sedation levels both in Groups BC, BS, and BL and in Groups LC, LS, and LL (P < 0.05). Convulsive doses of bupivacaine and levobupivacaine were significantly larger in Groups BL and LL than in Groups BC and LC, respectively (P < 0.01 for both). Concentrations of bupivacaine and levobupivacaine in plasma and in brain at the onset of convulsions were also larger in Groups BL and LL than in Groups BC and LC (P < 0.01 for both). In the second experiment, yohimbine (1 mg/kg) administered 10 min before and 5 min after the start of dexmedetomidine infusion completely reversed the sedative effect of dexmedetomidine (bolus 1.5 microg/kg, followed by 10.8 microg x kg(-1) x h(-1)). Convulsive doses and plasma and brain concentrations of bupivacaine and levobupivacaine at the onset of convulsions in rats receiving yohimbine and dexmedetomidine were significantly smaller than in those receiving only dexmedetomidine (P < 0.05 for all) and were similar to those without dexmedetomidine or yohimbine. We conclude that dexmedetomidine used for sedation decreases the convulsive potency of both bupivacaine and levobupivacaine in rats. Alpha(2)-adrenoceptor agonism may be involved in this anticonvulsant potency.

The P-glycoprotein inhibitor quinidine decreases the threshold for bupivacaine-induced, but not lidocaine-induced, convulsions in rats.

To examine whether inhibition of P-glycoprotein (P-gp) activity by quinidine affects the central nervous system toxicity of lidocaine and racemic bupivacaine (bupivacaine). Forty male Sprague-Dawley rats were randomly divided into four groups (n = 10). Fifteen minutes following administration of 15 mg x kg(-1) of quinidine (QL and QB groups) or saline (L and B groups), lidocaine (L and QL groups, 4 mg x kg(-1) x min(-1)) or bupivacaine (B and QB groups, 1 mg x kg(-1) x min(-1)) was infused until convulsions occurred. Concentrations of lidocaine and its primary metabolite, monoethylglycinexylidide (MEGX) and bupivacaine in plasma and in the brain at the onset of convulsions were measured by high-performance liquid chromatography. There were no differences in the dose of lidocaine required to induce convulsions between the L and QL groups. There were no differences in the concentrations of total (L = 17.2 +/- 3.5, QL = 16.6 +/- 2.6 micro g x mL(-1)) or unbound lidocaine (L = 7.8 +/- 2.5, QL = 7.3 +/- 2.3 micro g x mL(-1)), total (L = 1.2 +/- 0.5, QL = 1.3 +/- 0.7 micro g x mL(-1)) or unbound MEGX (L = 0.9 +/- 0.5, QL = 0.8 +/- 0.4 micro g x mL(-1)) in plasma, total lidocaine or MEGX in the brain at the onset of convulsions between the L and QL groups. The dose of bupivacaine required to induce convulsions was comparable in the B and QB groups. At the onset of convulsions, plasma concentrations of both total (B = 4.9 +/- 1.1, QB = 4.0 +/- 0.6 micro g x mL(-1), P = 0.03) and unbound bupivacaine (B = 1.4 +/- 0.6, QB = 0.9 +/- 0.2 micro g x mL(-1), P = 0.02) were significantly lower in the QB group than in the B group. There were no differences in concentration of total bupivacaine in the brain between the B and QB groups. These results suggest that quinidine inhibited P-gp activity, resulting in increased brain/plasma concentration ratio of bupivacaine, but not of lidocaine, and decreased the threshold of plasma concentration for bupivacaine-induced convulsions.

Determination of ropivacaine and bupivacaine in human plasma by programmed temperature vaporiser-fast gas chromatography-mass spectrometry (PTV/fast GC/MS) utilising in-vial liquid-liquid extraction

In-Vial liquid-liquid extraction has been investigated as a sample-preparation method in combination with fast gas chromatography and mass spectrometric detection for analysis of ropivacaine and bupivacaine in human plasma. The method is simple, quick, and readily automated. It offers a new possibility for the GC analysis of drugs in biological fluids. Validation showed that the method is accurate and selective. The acceptance criteria for the study validation were well in line with international criteria.

Liquid chromatographic analysis of local anesthetics in human plasma after sample preparation by on-line dialysis. Optimization by use of experimental design

A fully automated method involving dialysis, clean-up and enrichment of the dialysate on a pre-column packed with a strong cation-exchange phase, and subsequent liquid chromatographic (LC) analysis with UV detection at 220 nm has been developed for the determination of local anesthetics (prilocaine, mepivacaine, and bupivacaine) in human plasma. All sample-handling operations were executed automatically by means of an Asted XI system. To identify the most important conditions affecting analyte recovery from the dialysis and trace-enrichment processes, a seven-factor D-optimal design with 16 experimental points was elaborated as a screening test A four-factor D-optimal design with 24 experimental points was then used to predict and optimize analyte recovery. Derringer's desirability function was also used to deduce optimum conditions for analyte recovery and dialysis time within the experimental domain. Finally, the method developed was validated. Mean recoveries were approximately 72% for bupivacaine and approximately 67% for mepivacaine and prilocaine. The limits of quantification were 28 ng mL(-1) for bupivacaine and 25 ng mL(-1) for mepivacaine and prilocaine.

Plasma concentrations of bupivacaine after combined spinal epidural anaesthesia in infants and neonates.

The unbound and bound plasma concentration of bupivacaine in 50 infants less than 55 weeks postconceptual age was determined following combined spinal and epidural anaesthesia (csea). Plasma concentrations were determined at 15-min intervals up to 60 min postspinal anaesthesia. Maximum plasma bupivacaine levels were recorded between 45 and 60 min post CseA. Total plasma concentrations above a toxic threshold level of 4 microg.ml(-1) were recorded in 4% of patients and above 2.5 microg.ml(-1) in 10% of patients. Unbound bupivacaine levels were greater than a presumed toxic level of 0.25 microg.ml(-1) in 16% of cases and above 0.3 microg. ml(-1) in 14% of cases. A wide range of protein binding was measured (varying from 53.8-98.2%) and could not be correlated with standard indicators of local anaesthetic binding. Two neonates had brief apnoeas in the immediate perioperative phase but no adverse cardiac or central nervous system events attributable to the performance of Csea were demonstrated.

Determination of free concentrations of ropivacaine and bupivacaine in plasma from neonates using small-scale equilibrium–dialysis followed by liquid chromatography-mass spectrometry

Attempts to determine a safe plasma concentration of ropivacaine and bupivacaine in neonates have not been consistent. This might be due to an underestimation of free drug in small plasma samples by currently used techniques, e.g., ultrafiltration. We describe a simple microscale equilibrium–dialysis technique for the separation of free and bound ropivacaine and bupivacaine. The free drug in the dialysate was determined using solid-phase extraction and liquid chromatography with mass spectrometry. Pentycaine was used as an internal standard and added to the dialysates prior to extraction. The method is very selective and sensitive, as no compounds other than the analyte and internal standard were observed in the resulting chromatograms at low ng/ml levels. The limit of quantitation was 2.5 ng/ml. The calibration curve was linear in the range of 2 to 1000 ng/ml. The precision of the whole procedure was 8.1% ( n=10) and 6.5% ( n=7) for ropivacaine and bupivacaine, respectively. The method was tested in the analysis of plasma samples taken from neonates who had received epidural injections.

Disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments in rats.

This study was designed to determine the disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments, using multiple sampling time points in chronically prepared awake pregnant rats. All animals received an intravenous infusion of bupivacaine at a rate of 0.33 mg. kg-1. min-1 over a period of 15 min. The fetuses were delivered either at the end of infusion or at 2 or 4 h after dosing. Maternal and fetal blood and tissue samples were obtained for the assays of bupivacaine and its metabolites using capillary gas chromatography-mass spectrometry. The elimination half-life of bupivacaine was 37.7 min. The major metabolite was 3'-hydroxybupivacaine. Bupivacaine and 3'-hydroxybupivacaine were present in all samples at the end of administration. The fetal to maternal concentration ratio of bupivacaine in plasma was 0.29, and in the placenta was 0.63. The amnion contained the highest bupivacaine concentration: threefold higher in the maternal and 11-fold higher than in the fetal plasma. At 4 h after dosing, bupivacaine was no longer detectable in any maternal and fetal samples, whereas 3'-hydroxybupivacaine was still present in all tissues except the fetal plasma and heart. These data indicate that a considerable amount of bupivacaine is taken up by both sides of the placenta, as well as the amnion and myometrium. 3'-Hydroxybupivacaine was present in all tissues except the fetal plasma and heart samples, even after the parent compound became no longer detectable. Whether this slow elimination of 3'-hydroxybupivacaine causes any adverse effects on the fetus-newborn needs to be explored.

Nitric Oxide Modulation Affects the Tissue Distribution and Toxicity of Bupivacaine

We have previously demonstrated that inhibition of nitric oxide synthase (NOS) alters the toxicity of local anesthetics including bupivacaine. Because significant changes in blood distribution are associated with the use of nonselective NOS inhibitors, the purpose of this study was to determine whether modification of bupivacaine toxicity by nonselective NOS inhibition is due to alteration in tissue disposition of bupivacaine. Rats were anesthetized with halothane and pretreated with either: 1) a nonselective NOS inhibitor, N ω-nitro- l-arginine methyl ester ( l-NAME, 2 mg/kg/min, IV for 30 min); 2) a neuronal NOS inhibitor, 7-nitroindazole (7-NI, 30 mg/kg, IP); or 3) vehicle (control). Thirty minutes later, bupivacaine 2 mg/kg/min IV was infused until onset of seizures, arrhythmias, or asystole. l-NAME caused a rapid increase in plasma bupivacaine concentrations (3–4 times faster than in the other groups), which was associated with markedly lower bupivacaine doses (mg/kg) required to produce arrhythmias in l-NAME (4.2 ± 0.5) vs. control (26 ± 3, p < 0.01) and 7-NI groups (17 ± 3, p < 0.01). Myocardial bupivacaine concentrations at arrhythmia onset were slightly lower in the l-NAME group. Bupivacaine seizure doses in 7-NI and l-NAME pretreated animals were similar to control but significantly different from each other. Brain bupivacaine concentrations at seizure onset were similar among the groups. There were no significant differences between 7-NI and control groups in any parameter observed. We conclude that enhanced cardiotoxicity of bupivacaine by nonselective NOS inhibition is primarily due to rapid increases in plasma and myocardial distribution of bupivacaine.

Plasma bupivacaine concentrations and effects of epinephrine after superficial cervical plexus blockade in patients undergoing carotid endarterectomy

Objective: To determine if epinephrine (EPI) added to a solution of bupivacaine (BUP) injected for use in superficial cervical plexus blockade (SCPB) lowers plasma BUP concentrations after injection and whether this addition of EPI resulted in tachycardia, cardiac arrhythmias, or both. Design: Randomized, unblinded prospective clinical interventional study. Participants: Patients scheduled to undergo carotid endarterectomy using SCPB consenting to study. Setting: University-affiliated tertiary care hospital operating room. Interventions: Twenty patients were given SCPB with BUP 0.5% and were randomized to receive either no EPI or 1: 300,000 EPI. This study block was followed by a second period in which 20 patients were given SCPB with BUP 0.25% randomized to receive either no EPI or 1: 300,000 EPI. Continuous electrocardiogram monitoring was performed during and after the block and analyzed for heart rate and rhythm changes. Measurements and Main Results: Arterial plasma BUP concentrations were measured 2.5 to 120 minutes after initiation of SCPB. Plasma BUP concentrations were highest in the 0.5% no EPI group, followed by the 0.5% EPI, 0.25% no EPI, and 0.25% EPI groups. The use of EPI did not significantly affect heart rate or change the incidence of cardiac arrhythmias. Conclusions: BUP 0.25% consistently produced the lowest plasma BUP concentrations, particularly when EPI was added to the solution. BUP 0.5% without EPI can produce plasma BUP concentrations previously reported to be associated with central nervous system effects. The use of EPI in this setting does not produce untoward cardiac side effects.

Bupivacaine-loaded comatrix formed by albumin microspheres included in a poly(lactide-co-glycolide) film: in vivo biocompatibility and drug release studies.

Bupivacaine-loaded comatrix, formed by bupivacaine-loaded microspheres included in a poly(lactide-co-glycolide) film, was assayed for the controlled release of the drug 'in vivo'. The comatrix, with 66.37 microg of bupivacaine, signifying a dose of 265.5 microg/kg, was subcutaneously implanted in the back of rats. Maximum plasma bupivacaine concentration was 147.6 +/- 5.0 ng/ml 95 h after the device implantation, and the drug was detected in plasma for 17 days. The half-life time of bupivacaine improves by more than 50 times with regard to that of the drug administered in a solution by intraperitoneal injection. After 15 days of implantation the comatrix was included in a thin fibrous capsule and degradation of the microspheres was observed. The histological studies show good biocompatibility of this comatrix. After 50 days the comatrix was degraded and its remains were almost indistinguishable from the surrounding tissue. Small number of microspheres was observed and they were surrounded by conjunctive tissue. Nerve packets and small blood vessels were also observed in the periphery of the implant.