Nonformalin Fixative Agents: A Comparative Study of Fixative Efficacy and Histomorphology.

OBJECTIVES: Formalin is a gold standard fixative agent. However, formalin possesses health hazards and is not always available in rural areas. The objective of this study was to compare tissue fixative efficacy of nonformalin fixative agents with formalin. METHODS: Oral tissues obtained during impacted tooth removal were collected. The tissue fragments were sectioned and fixed in 4 different fixatives; 30% jaggery, 70% ethanol, 2% mepivacaine with 1:100 000 epinephrine, or formalin for 24 and 72 hours. All specimens were then immersed in formalin for another 24 hours and processed according to standard protocol. Hematoxylin and eosin-stained sections were evaluated by a pathologist. The cellular structure, cellular outline, and quality of staining were graded from 1 to 3 and average fixative efficacy scores were compared using 1-way analysis of variance. Fixative artifacts were described. RESULTS: Fixative efficacy scores of 70% ethanol and 30% jaggery at 24 and 72 hours were not statistically different from those of formalin. Conversely, 2% mepivacaine demonstrated significantly lower fixative efficacy scores than other agents. Although efficacy of each fixative was not statistically different between 24 and 72 hours, efficacy of 70% ethanol was markedly reduced at 72 hours when compared with others. Acantholysis of epithelial cells was the most notable artifact at 72 hours when fixed with nonformalin fixative agents. CONCLUSION: Both 70% ethanol and 30% jaggery provided acceptable fixative efficacy at 24 hours. However, only 30% jaggery maintained fixative efficacy at 72 hours. Nevertheless, negative effects on the epithelial cells were unavoidable and should be interpreted with caution.

The stability and microbial contamination of bupivacaine, lidocaine and mepivacaine used for lameness diagnostics in horses.

Local anaesthetics (LAs) are frequently used for diagnostic procedures in equine veterinary practice. The objective of this study was to investigate the physico-chemical stability and bacterial contamination of bupivacaine, lidocaine and mepivacaine used for lameness examinations in horses. The LAs were stored in 12 different groups at different temperatures (-18 °C to 70 °C), light intensities and in common veterinary field conditions for up to 16 months. The pH, presence of bacterial contamination and concentrations of LAs and methylparaben (a preservative present in lidocaine) were determined serially in both new and repeatedly punctured (RP) vials. Mepivacaine remained chemically stable. A 1.9% increase in bupivacaine concentration was evident in one group, whereas a 1.9-3.7% decrease was noted in six groups. Risk factors associated with a change in concentration were light and RP vials. Lidocaine concentration decreased 6.3% in one group and increased 5.3-7.2% in two groups. Risk factors for degradation were heat and RP vials whereas storage in practice vehicles was a risk factor for increased concentrations. Methylparaben decreased 8.3-75.0% in seven groups, and RP vials, heat and storage in practice vehicles were risk factors for degradation. No contamination was present in any of the LAs and pH remained stable. Commercially available solutions of lidocaine, mepivacaine and bupivacaine stored under common veterinary field conditions are extremely stable and sterile for extended periods. The minor changes in concentration documented in this study are unlikely to affect anaesthetic efficacy during equine lameness examinations. When using products containing methylparaben, degradation of the preservative over time is to be expected.

[Pharmakological characteristic of some amide local anesthetics, currently used in dental clinics].

Along with the brief history of amide local anesthetics development, their most important properties (from the viewpoint of use in clinical dental practice), are also reviewed. In particular, some properties of most commonly used local anesthetics, such as lidocaine, mepivacaine, prilocaine, bupivacaine and articaine are analysed. The most important data concerning pharmacological mechanisms of mentioned anesthetics' action, that cause certain features and peculiarities of their clinical application are given in condensed form. Besides, some precaution measures that must be taken into account in specific clinical cases together with the history and current status of each patient are mentioned as well.

Enhanced local anesthetic action of mepivacaine from the bioadhesive gels.

Mepivacaine, an amide-type local anesthetic, has been used to relieve local pain. Among the many drug delivery systems, transdermal drug delivery has some advantages, as it provides controlled drug delivery for an extended period of time. To develop new gel formulations that have suitable bioadhesion, the bioadhesive force of hydroxypropyl methylcellulose (HPMC) was assessed using an auto-peeling tester. The effect of drug concentration on drug release from 2% HPMC gel was studied using synthetic cellulose membrane at 37±0.5°C. The drug concentrations tested were 0.5, 1, 1.5, 2, and 2.5%. The effect of temperature on drug release from the 2% drug gel was evaluated at 27, 32, 37 and 42°C. To increase the skin permeation of mepivacaine from HPMC gel, enhancers such as saturated and unsaturated fatty acids, pyrrolidones, propylene glycol derivatives, glycerides, and non-ionic surfactants were incorporated into the mepivacaine-HPMC gels. The enhancing effect of the enhancer on drug permeation was then examined in the modified Keshary-Chien cell. For the efficacy study, the anesthetic action of the formulated mepivacaine gel containing enhancer and vasoconstrictor was evaluated with the tail-flick analgesimeter. Among the various kinds of HPMC, HPMC-K100M gel showed the highest viscosity and bioadhesive force. As the viscosity of the HPMC gels increased, the bioadhesive forces increased. Increasing the drug concentration or temperature increased the drug release rate. Among the enhancers used, polyoxyethylene 2-oleyl ether showed the greatest enhancement of permeation. Based on the area under the efficacy curve of the rat tail flick test curve, mepivacaine gel containing polyoxyethylene 2-oleyl ether and tetrahydrozoline showed prolonged and increased local anesthetic action compared to the control. For bioadhesive mepivacaine gels with enhanced local anesthetic action, mepivacaine gels containing penetration enhancer and vasoconstrictor could be developed with the bioadhesive polymer, HPMC.

Brain penetration of local anaesthetics in the rat: Implications for experimental neuroscience.

Multiple experimental neuroscience techniques rely on the use of general anaesthesia to minimize the discomfort associated to animal restraint and to achieve a more effective control of relevant physiological parameters. In order to minimise potential interference on brain neuronal activity, such studies are typically conducted at low anaesthetic doses. This practice is often coupled to peripheral infiltration of local anaesthetics to provide supplementary analgesia and prevent sub-threshold activation of pain pathways that may confound central measurements of brain function. However, little is known of the effect of peripheral anaesthesia on central measurements of brain activity in small laboratory animal species. In order to begin to address this question, we measured total and free brain exposure of five different local anaesthetics following subcutaneous infiltration of analgesic doses in a surgical protocol widely used in rodent neuroimaging and electrophysiology studies. Notably, all the anaesthetics exhibited detectable total and free brain concentrations at all the time points examined. Lidocaine and mepivacaine showed the highest free brain exposures (>525 ng/g), followed by bupivacaine and ropivacaine (>70 ng/g). The ester-type local anaesthetic tetracaine produced the lowest free brain exposure (<8.6 ng/g). Our data suggest that peripheral administration of local anaesthetics in small laboratory animals could result in pharmacologically active brain exposures that might influence and confound central measurements of brain function. The use of the ester-type anaesthetic tetracaine produced considerably lower brain exposure, and may represent a viable experimental option when local anaesthesia is required.

Liposomal formulations of prilocaine, lidocaine and mepivacaine prolong analgesic duration.

PURPOSE: A laboratory investigation was undertaken to compare the in vivo antinociceptive effects of 2% liposomal formulations of prilocaine (PLC), lidocaine (LDC) and mepivacaine (MVC) compared to plain solutions of each of these three local anesthetics. METHODS: Large unilamellar vesicles were prepared by extrusion (400 nm), at pH 7.4. The membrane/water partition coefficients were obtained from encapsulation efficiency values, after incorporation of each local anesthetic to the vesicles. The anesthetic effect of each liposomal formulation was compared to the respective local anesthetic solution in water, using the infraorbital nerve-blockade test, in rats. RESULTS: The partition coefficients were: 57 for PLC, 114 for LDC and 93 for MVC. In vivo results showed that local anesthetic-free liposomes, used as control, had no analgesic effect. In contrast, the encapsulated formulations induced increased intensities of total anesthetic effect (35.3%, 26.1% and 57.1%) and time for recovery (percentage increases of 30%, 23.1% and 56%), respectively, for PLC, LDC and MVC when compared to the plain solutions (P < 0.01). CONCLUSIONS: These results indicate that liposomes provide effective drug-delivery systems for intermediate-duration local anesthetics. Mepivacaine was affected to the greatest extent, while LDC benefited least from liposome encapsulation, possibly due to greater vasodilatory properties of LDC.

Solid-state study of mepivacaine hydrochloride.

Different crystalline forms of the local anaesthetic mepivacaine hydrochloride (MH) were revealed by Fourier transform infrared spectroscopy (FT-IR), not by conventional differential scanning calorimetry (DSC). The existence of two polymorphic anhydrous modifications was discovered and further characterized by X-ray powder diffraction and thermal analysis: Form II, the commercial one, and the more stable Form I, obtained by re-crystallization from Form II. Two pseudopolymorphs were also obtained: Form III, a solvate crystallized from ethanol and Form IV, a solvate crystallized from methanol. Single crystal X-ray diffraction data for both solvates were collected and their structures were determined. Form II, metastable and monotropically related to Form I, generates through desolvation of Form III, very often present in industrial processing, where crystallization from ethanol solution is a common practice. For the sake of clarity, the presence of polymorphic forms should be reported in the drug master files of MH. However, since MH is readily water soluble, the observed polymorphism has no relevance to its typical clinical use as aqueous solutions.

Delayed-type hypersensitivity to amide local anesthetics.

We present a 54-year-old woman who suffered eczematous eruptions on her face after the administration of lidocaine and mepivacaine for dental surgeries. Patch tests showed delayed-type-hypersensitivity to the amide local anesthetics lidocaine and mepivacaine with cross reaction to other amynoacylderivatives (prilocaine,bupivacaine) but not articaine.

Fast screening of low molecular weight compounds by thin-layer chromatography and "on-spot" MALDI-TOF mass spectrometry.

Fast screening of low-MW compounds is performed by thin-layer chromatography (TLC) followed by direct on-spot matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification with nearly "matrix-free" mass spectra using an UV-absorbing ionic liquid matrix. Owing to minimal background ions from the proton donor triethylamine/alpha-cyano-4-hydroxycinnamic acid ionic liquid matrix, three arborescidine alkaloids, the anesthesics levobupivacaine and mepivacaine, and the antibiotic tetracycline were readily characterized most frequently by the MS detection of their protonated molecules. The technique is fast and sensitive, requires little sample preparation and manipulation, and is therefore suitable for fast screening with TLC separation and MS identification of low-MW compounds, with potential applications in areas such as phytochemistry, synthetic chemistry, and product manufacturing quality monitoring.

Membrane effects of ropivacaine compared with those of bupivacaine and mepivacaine.

We compared the effects of ropivacaine, bupivacaine and mepivacaine on membrane lipids in an attempt to determine the anaesthetic mechanism of ropivacaine with structure-dependent potency. The membrane effects were determined by measuring anaesthetic-induced changes in the phase transition temperature and the fluorescence polarization of liposomal membranes prepared with cholesterol and phosphatidylcholine. Bupivacaine, ropivacaine and mepivacaine depressed the membrane lipid phase transition and decreased the polarization of liposomal membranes at 0.0625-1.0 mg/mL, indicating that these anaesthetics fluidize membranes at concentrations lower than those in clinical use. Ropivacaine and bupivacaine were effective in fluidizing the membrane core rather than the membrane surface, whereas mepivacaine was a membrane fluidizer acting equally on both regions. In the comparison of membrane fluidization at an equimolar concentration (3.0 mmol/L), ropivacaine was found to be less potent than bupivacaine and more potent than mepivacaine. This membrane-fluidizing potency was also consistent with the hydrophobic properties of these substances evaluated by reversed-phase chromatography. Structure-dependent membrane fluidization associating with hydrophobicity appears to underlie the local anaesthetic effect of ropivacaine as well as those of bupivacaine and mepivacaine.

Identification of the precipitate in alkalinized solutions of mepivacaine and bupivacaine at 37 degrees C.

Commercial and control solutions of bupivacaine (0(.)75%) and mepivacaine (1(.)5%) were alkalinized with bicarbonate until cloudy at room temperature. The solutions were heated to 37 degrees C for 2(.)5 h. The precipitates were filtered, lyophylized and analysed by fast atom bombardment mass spectrometry. Analysis showed the precipitates to be predominantly the free base of the local anaesthetic. The precipitate of the commercial bupivacaine solution also contained a small amount of the hydrochloride salt. The mepivacaine control crystals contained an unknown at molecular weight 528, which may represent a dimer of the free base and hydrochloride salt.

Transient radicular irritation after spinal anesthesia induced with hyperbaric solutions of cerebrospinal fluid-diluted lidocaine 50 mg/ml or mepivacaine 40 mg/ml or bupivacaine 5 mg/ml.

BACKGROUND: Transient radicular irritation (TRI) is common after spinal anesthesia induced with hyperbaric lidocaine 50 mg/ml. The purpose of this study was to determine the incidence of TRI after spinal anesthesia with hyperbaric lidocaine 50 mg/ml diluted with cerebrospinal fluid (CSF) 1:1 and hyperbaric mepivacaine 40 mg/ml and hyperbaric bupivacaine 5 mg/ml. METHODS: Ninety ASA class I-IV patients undergoing mostly brief urological procedures under spinal anesthesia were randomly allocated to receive either hyperbaric lidocaine 50 mg/ml diluted with CSF 1:1 (Group L), hyperbaric mepivacaine 40 mg/ml (Group M) or hyperbaric bupivacaine 5 mg/ml (Group B). Characteristics of the patients and details of the surgical procedures and spinal anesthesias were similar in all groups except for the intensity of motor block. The patients were evaluated on the first postoperative day by an anesthesiologist who did not know which spinal anesthetic agent had been used. RESULTS: Six patients (20%) in group L, 11 patients (37%) in Group M and none (0%) in Group B experienced pain in the legs and/or back (TRI) after spinal anesthesia. CONCLUSION: TRI is frequent after spinal anesthesia induced with hyperbaric lidocaine 50 mg/ml diluted with CSF 1:1. The incidence of TRI after hyperbaric mepivacaine 40 mg/ml is of the same magnitude. TRI could not be observed after bupivacaine spinal anesthesia.

Stability of local anesthetics in heparinized blood, plasma and sulfuric acid.

BACKGROUND: This study was undertaken to investigate the stability of local anesthetics in different media. METHODS: 60 patients with axillary plexus block, using prilocaine, mepivacaine or lidocaine 1%, or with lumbar epidural catheter-anesthesia, using bupivacaine 0.5%, were investigated. Stability was evaluated in heparinized whole blood for 24 h at +4 degrees C, in plasma for 8 week at -25 degrees C and in sulfuric acid for 6 months at -25 degrees C. RESULTS: Mean recovery rates were between 96.4% and 102.6% in all local anesthetics and media. No statistical significance became obvious for differences between recovery rates and initial values. CONCLUSION: Stability of prilocaine, mepivacaine, lidocaine and bupivacaine can be expected when samples are stored as heparinized whole blood for 24 h at +4 degrees C, plasma for 8 weeks at -25 degrees C, and after preparation into sulfuric acid for 6 months at -25 degrees C.

Enantioseparation of anaesthetic drugs by capillary zone electrophoresis using cyclodextrin-containing background electrolytes.

The enantiomers of five racemic anaesthetic drugs were resolved with cyclodextrins using capillary zone electrophoresis. Parameters which affected the chiral resolution, such as type and concentration of cyclodextrin, temperature, and addition of organic modifier were investigated. The results show that the enantiomeric discrimination of the solutes is influenced by the structural shape of the solute molecules, separation temperature, and type of cyclodextrin. It was found that alpha-cyclodextrin was the best enantioselector for resolution of prilocaine and ketamine, while the enantiomers of mepivacaine, ropivacaine, and bupivacaine were resolved with beta-cyclodextrin and/or modified beta-cyclodextrins, i.e., methyl- and 2-hydroxypropyl-beta-cyclodextrin, as chiral selectors. The length of the alkyl chain on the amino group of the drug molecule had a strong effect on the enantioresolution of mepivacaine, ropivacaine, and bupivacaine. Baseline separation of racemic ketamine was achieved with alpha- and methyl-beta-cyclodextrin at 15 degrees C. Addition of 5 M urea to the running buffer containing beta-cyclodextrin at high concentrations resulted in the enantioseparation of prilocaine, mepivacaine, and ketamine. Enantioresolution was improved upon the addition of 10% methanol to the buffer containing urea and beta-cyclodextrin. Generally, the complex formed between the S-enantiomers and modified beta-cyclodextrins was stronger than the corresponding R-forms. An exception was prilocaine where the R-form gave a more stable complex both with alpha- and beta-cyclodextrin.

Pharmacokinetics of the enantiomers of bupivacaine and mepivacaine after epidural administration of the racemates.

UNLABELLED: We investigated the pharmacokinetics of the enantiomers of bupivacaine and mepivacaine after epidural injection of the racemate of each drug into six surgical patients. After epidural administration of either bupivacaine/HCl (115 mg) or mepivacaine/HCl (460 mg), blood samples were collected for 24 h. Unbound fractions were determined by using ultrafiltration for bupivacaine and equilibrium dialysis for mepivacaine. Concentrations in plasma, ultrafiltrate, and dialysate were determined by using stereoselective high-performance liquid chromatography. Peak plasma concentrations of R(+)-bupivacaine (389 +/- 93 ng/mL) and R(-)-mepivacaine (1350 +/- 430 ng/mL) were smaller than those of S(-)-bupivacaine (449 +/- 109 ng/mL, P < 0.0001) and S(+)-mepivacaine (1740 +/- 490 ng/mL, P < 0.002), respectively. However, the unbound peak concentrations of R(+)-bupivacaine (20 +/- 11 ng/mL) were larger than those of S(-)-bupivacaine (15 +/- 9 ng/mL, P < 0.005); unbound peak concentrations of R(-)-mepivacaine (485 +/- 158 ng/mL) and S(+)-mepivacaine (460 +/- 139 ng/mL) did not differ. These observations reflect differences in the systemic disposition (distribution and elimination) of the enantiomers, because the systemic absorption was not enantioselective with either drug. This study supports the opinion that the use of single enantiomers, rather than racemates, is preferable, particularly for bupivacaine. IMPLICATIONS: Measurements of the plasma concentrations of the enantiomers of bupivacaine and mepivacaine after epidural administration of the racemates demonstrated that the systemic disposition, but not the systemic absorption, of these drugs is enantioselective and supports the opinion that the use of single enantiomers, rather than racemates, is preferable.

Structure-affinity relationships and stereospecificity of several homologous series of local anesthetics for the beta2-adrenergic receptor.

UNLABELLED: Local anesthetics inhibit binding of ligands to beta2-adrenergic receptors (beta2ARs), and, as a consequence, inhibit intracellular cAMP production. We hypothesized that among homologous local anesthetics, their avidity at inhibiting binding of tritiated dihydroalprenolol (3H-DHA) to beta2ARs would increase with increasing length of alkyl substituents and would demonstrate stereospecificity. Specific binding of 3H-DHA to human beta2ARs was assayed in the presence of six different members of the 1-alkyl-2,6-pipecoloxylidide class of local anesthetics (including mepivacaine, ropivacaine, and bupivacaine), the R(+) and S(-) bupivacaine enantiomers, lidocaine, prilocaine, etidocaine, procaine, and tetracaine. Avidity of binding to beta2ARs increased with increasing length of the alkyl chain (pKi values = 2.4, 3.6, 4.3, 4.1, 4.1, 5.9 for the methyl [mepivacaine], ethyl, S(-)propyl [ropivacaine], butyl [bupivacaine], pentyl, and octyl derivatives, respectively). We found no evidence for bupivacaine stereospecificity (pKi values = 4.3 and 4.9 for the S(-) and R(+) isomers, respectively). Other amide and ester local anesthetics also showed increasing potency with increasing length of alkyl substituents (pKi values = 3.6, 3.8, and 4.3 for lidocaine, prilocaine, and etidocaine; 4.2 and 5.6 for procaine and tetracaine, respectively). The correlation between increased inhibition of beta2AR binding and alkyl chain length resembles the correlation between local anesthetic potency at nerve block and increased alkyl chain length. The lack of clear stereospecificity is consistent with the relatively low potency these agents demonstrate at inhibition of beta2AR binding. Finally, the relatively potent inhibition of beta2ARs by etidocaine, tetracaine, and bupivacaine suggests that their propensity for cardiovascular depression after accidental intravenous overdose could result from beta2AR or beta1AR blockade and inhibition of cAMP production. IMPLICATIONS: Local anesthetics demonstrate a rank order of avidity for displacing ligands from beta2-adrenergic receptors such that larger molecules displace ligands at lower concentrations than smaller local anesthetic molecules. This relationship between molecular size and receptor avidity could explain the greater propensity for cardiovascular toxicity of relatively large local anesthetics such as bupivacaine.

The relationship between pH and concentrations of antioxidants and vasoconstrictors in local anesthetic solutions.

pH affects the efficacy of local anesthetics by determining the percentage of the lipid-soluble base form of the anesthetic available for diffusion and penetration of the nerve sheath. The purpose of this study was to determine the relationship between pH and the concentrations of antioxidant and vasoconstrictor in dental local anesthetic solutions over real-time and after accelerated aging. Several batches of lidocaine and mepivacaine with vasoconstrictors were tested. Results showed that, immediately upon receipt from the manufacturers, three batches were below the USP pH limit (pH 3.3), and two batches contained less than the minimum limit of vasoconstrictors (90%). Real-time tests on batches that were within normal limits revealed that solutions were stable past 4 yr. Accelerated aging tests revealed a strong correlation between a decrease in pH and loss of antioxidants and vasoconstrictors. In conclusion, a quality batch of local anesthetic should remain efficacious long past the manufacturer's stated shelf life; a batch that is less than optimal, or one that is exposed to environmental stresses, will degrade rapidly, and efficacy may be affected by decreases in pH and loss of vasoconstrictor. pH may be an inexpensive, readily available screening test for efficacy of local anesthetics.

[Changing compatibility by temperature of local anesthetics with sodium bicarbonate].

We investigated the amount of 7% sodium bicarbonate which could be added to local anesthetics (bupivacaine, mepivacaine and lidocaine) without any precipitation when temperature was altered (5 degrees C, 25 degrees C, 40 degrees C). Precipitations occurred at any temperatures and with any dose of bupivacaine. Interestingly, the higher the temperature, the more frequently precipitation with mepivacaine and lidocaine occurred. This was probably because the solubility of the precipitate produced by the interaction of the base of local anesthetics with bicarbonate was reduced when the temperature increased.