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Laboratory Science

Alkalinization increases penetration of lidocaine across the human cornea

Fuchsjäger-Mayrl, Gabriele MDa; Zehetmayer, Martin MD*,a; Plass, Herbert PhDb; Turnheim, Klaus MDb

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Journal of Cataract & Refractive Surgery: April 2002 - Volume 28 - Issue 4 - p 692-696
doi: 10.1016/S0886-3350(01)01233-0
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Abstract

In the past decade, topical anesthesia has been widely used for small-incision phacoemulsification, especially clear corneal cataract surgery. Topical anesthesia is considered safe and effective, greatly reducing the risk of complications and systemic toxicity and eliminating the use of needles.1–4

Lidocaine 4% and bupivacaine 0.5% or 0.75% are among the agents most frequently used for topical anesthesia in small-incision cataract surgery. Both have a rapid onset time.5 Lidocaine is known to be less toxic to the corneal epithelium than bupivacaine.6 Short-term exposure of corneal endothelium to intracameral lidocaine appears to be safe.7,8

Lidocaine is a weak base with a pKa value of 7.8. Hence, at pH 5, only 0.16% of the molecules are nonionized in aqueous solution, calculated from the Henderson-Hasselbalch equation. According to the principle of “nonionic diffusion” or “partition hypothesis,” only the nonionized moiety of a weak electrolyte is expected to penetrate lipid membranes.9,10 Increasing the pH of the anesthetic solution by adding sodium bicarbonate (pH adjustment) should augment the rate of penetration of the local anesthetic agent into the tissue.5,11 In a previous in vivo study, aqueous humor levels of lidocaine after topical application of lidocaine 4% in a pH 7.2 solution were 210% higher than in a pH 5.2 solution.11

In the present study, we examined the pH dependence of the penetration of lidocaine across the human cornea in vitro.

Materials and Methods

Ten pairs of human corneas were obtained from the Eye Bank, Department of Ophthalmology, University of Vienna. The corneas were not considered for corneal transplantation because of 1 or more of the following reasons: enucleation more than 6 hours after death of donor, potential human immunodeficiency virus or hepatitis C virus infection (positive or missing results due to hemolytic sera), advanced donor age, and systemic donor infection or leukemia.12 The use of human corneas in the study was approved by the Ethics Committee of the Medical School, University of Vienna.

The mean donor age of the corneas was 64 years ± 7 (SD). Eight donors were men and 2, women. The mean postmortem enucleation time was 7.9 ± 0.6 hours. Sterile conditions were maintained throughout the enucleation procedure. The mean corneal endothelial cell density, examined by specular microscopy (Labovert® large-angle microscope, Leitz), was 2635 ± 150 cells/mm2. The corneas were stored in a culture medium (Optisol®, Chiron Vision) at 2°C to 4°C. Only transparent corneas with clinically intact epithelium were used. The mean time between enucleation and the experimental procedure was 5.5 ± 2 days (range 2 to 8 days).

A pair of corneas were mounted simultaneously in an in vitro perfusion system (modified Ussing chambers, Figure 1) and short circuited to eliminate electrical driving forces for the charged species of lidocaine using an automatic voltage clamp (VCC 600, Physiologic Instruments).13 Electrode asymmetries and fluid resistance were automatically compensated for by the voltage clamp.

Figure 1.
Figure 1.:
(Fuchsjäger-Mayrl) Schematic diagram of the modified Ussing chamber for corneal permeability studies. I = electric current, V = voltmeter.

Special care was taken to minimize corneal distortion or edge damage. On the endothelial side of the chamber, a rubber O-ring was inserted. The inner diameter of the Ussing chamber was 8 mm, resulting in an exposed corneal area of 50 mm2. By appropriate addition of Na+-acetate buffer, pH 5, or NaHCO3 solution, the osmolality of the epithelial bathing solutions was set to 290 mOsm/L, measured with a vapor pressure osmometer 5520 (Wescor), resulting in isotonic solutions with pH 5 and pH 7, respectively. The pH values remained unchanged during the entire experiment. The endothelial bathing solution consisted of (mM) 132 Na2+, 132 Cl, 5.4 K+, 1.2 Mg2+, 1.2 Ca2+, 2.4 HPO42−, 0.6 H2PO4, pH-adjusted to 7.4 by adding Hepes-Tris buffer. The osmolality of this solution was also 290 mOsm/L. All solutions contained 10 mM glucose. Both bathing solutions were recirculated by a gas-lift system with pure O2 at 37°C.

Transcorneal fluxes of lidocaine were determined by adding 14C-labeled lidocaine to the epithelial bathing solution together with 3H-polyethylene glycol (PEG), MM 4000, as a marker for the extracellular pathway. Fifty microliter aliquotes were sampled from the endothelial bathing solution every 15 minutes for 180 minutes to measure the radioactivity in a liquid scintillation counter (LS 6500, Beckman Instruments Inc.). Transcorneal fluxes of lidocaine were calculated from the differences in the appearance rate of 14C-labeled lidocaine and 3H-PEG in the endothelial solution; hence, lidocaine fluxes were corrected for extracellular movements.

The 14C-lidocaine and 3H-PEG were purchased from New England Nuclear. All other chemicals and reagents were supplied by local dealers.

The tissue content of lidocaine in the cornea was obtained from the time span until the unidirectional transcorneal 14C-lidocaine fluxes reached their steady state.14

The results are given as means ± standard deviations. The statistical significance of the mean differences was analyzed using paired t tests. Probabilities less than 0.05 were considered significant.

Results

Unidirectional transcorneal lidocaine fluxes reached a steady state after about 90 minutes. At all sampling times, transcorneal fluxes of lidocaine were significantly higher at pH 7 than at pH 5 (P < .05). The mean steady-state flux at pH 5 was 59 ± 34 nmol/min·cornea and at pH 7, 101 ± 37 nmol/min·cornea. This difference was significant (P < .002). Permeation of lidocaine via PEG-accessible shunts was always below 15% of the total flux.

The time-dependent accumulation of lidocaine on the endothelial side of isolated human corneas at pH 5 and pH 7 is shown in Figure 2. The total amount of lidocaine transferred through the cornea to the endothelial side within 180 minutes was significantly higher at pH 7 than at pH 5 (16.1 ± 5.5 μmol/cornea versus 9.1 ± 4.7 μmol/cornea; P < .001).

Figure 2.
Figure 2.:
(Fuchsjäger-Mayrl) Time-dependent accumulation of lidocaine on the endothelial side of the cornea following addition of 14C-labeled lidocaine to the epithelial side. The negative y-axis intersect of the linear regression of the steady-state fluxes with time (interrupted line) represents the corneal content of lidocaine. ∗Differences in lidocaine accumulation at pH 5 and pH 7 were significant (P < .005).

The steady-state tissue content of lidocaine in the cornea at pH 7 was 2.8 ± 0.9 μmol/cornea and at pH 5, 1.7 ± 1.2 μmol/cornea (not significant). The increase in corneal lidocaine content at pH 7 may be attributed to the increased solubility of the mainly undissociated lidocaine in membranes.

Discussion

Corneal penetration enhancers such as the disinfectants chlorhexidine and benzalkonium chloride increase corneal drug penetration.15,16 However, their use is often limited by problems of corneal toxicity.17 Another way to enhance ocular drug penetration is to modify the physicochemical properties of drugs.18 Alkalinization of local anesthetic agents is a well-known method to improve the anesthetic effect and its duration.19–21 Neutral solutions of lidocaine induce significantly less pain during infiltration than acidic solutions22–25 and have a more rapid onset.26 In a study in rabbit eyes,5 various topical anesthetic agents at different concentrations and acidity were investigated. Compared with unbuffered solutions, buffered preparations of lidocaine and bupivacaine had a significantly longer anesthetic effect.

In the present study, penetration of lidocaine across human corneas in vitro was about 70% faster at pH 7 than at pH 5. This agrees with the results in a previous clinical study in which the accumulation of lidocaine in the aqueous humor of the anterior eye chamber was 3-fold higher when topical lidocaine was administered in a solution of pH 7.2 than in a solution of pH 5.2.11 The fact that in the present in vitro study transcorneal lidocaine penetration was stimulated less by alkalinization may be related to differences in the penetration areas (central part of cornea in the present in vitro study versus cornea and conjunctiva), to the effect of solution pH on lacrimation and blinking in vivo,27 to the buffering action of tears,28 or to structural changes in the isolated corneas related to the postmortem time and storage. It is not reasonable to study lidocaine solutions with pH values higher than 7.2 to 7.4, since in alkaline solutions, lidocaine tends to precipitate during autoclavation.11

In conclusion, the present in vitro study showed that lidocaine penetration across the human cornea and the corneal content of lidocaine was significantly higher at pH 7 than at pH 5. According to the Henderson-Hasselbalch equation, at pH 7, 16.00% of lidocaine is in its undissociated state compared with 0.16% at pH 5. These findings agree with the notion of nonionic diffusion or the pH partition hypothesis of diffusion, according to which a weak electrolyte penetrates lipid biomembranes primarily in its undissociated state.9,10 In the cornea, lipid barriers are primarily formed by the cell membranes of the epithelium and endothelium, whereas the stroma of the cornea is predominantly hydrophilic.29 Alkalinization of the lidocaine solution by adding NaHCO3 is easy to perform and inexpensive.

The main clinical advantages of adjusting the pH of anesthetic solutions to more physiological values include less local irritation and lacrimation as well as augmentation and prolongation of the anesthetic action.11

References

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