Brimonidine Concentrations Research Articles

Diadenosine tetraphosphate improves adrenergic anti-glaucomatous drug delivery and efficiency

The effect of the dinucleotide P1, P4-Di (adenosine-5') tetraphosphate (Ap4A) in improving adrenergic anti-glaucomatous delivery by modifying the tight junction proteins of the corneal epithelium was evaluated. Stratified human corneal epithelial cells (HCLE) were treated with Ap4A (100 μM) for 5 min and TJ protein levels and barrier function were analysed by western blotting and transepithelial electrical resistance (TEER), respectively. Western blot experiments showed a significant reduction at 2 h (45% reduction of ZO-1 and 65% reduction of occludin protein levels) as compared to non-treated (control) cells. Two hours after Ap4A treatment, TEER values were significantly reduced (65% as compared to control levels (p < 0.001)), indicating an increase in corneal barrier permeability. Topical application of Ap4A in New Zealand white rabbits two hours before the instillation of the hypotensor compounds (the α2-adrenergic receptor agonist, brimonidine and the β-adrenergic receptor antagonist, timolol), improved the delivery of these compounds to the anterior chamber as well as their hypotensive action on the intraocular pressure. The results obtained showed that, when Ap4A was topically applied two hours before the adrenergic compounds, the concentration of brimonidine in the aqueous humour increased from 64.3 ± 5.3 nM to 240.6 ± 8.6 nM and from 58.9 ± 9.2 nM to 183.7 ± 6.8 nM in the case of timolol, which also produces a more profound effect on IOP. Therefore, Ap4A treatment results in a better entrance of adrenergic anti-glaucomatous compounds within the eye and consequently improved therapeutic efficiency by increasing corneal epithelial barrier permeability.

Trans-meningeal drug delivery to optic nerve ganglion cell axons using a nanoparticle drug delivery system

The purpose of this study was to investigate if neuroprotective drugs can cross the optic nerve sheath in vitro. Four optic nerves were used for this study. Two porcine nerves were harvested at the time of euthanasia and two human nerves were obtained at the time of therapeutic globe enucleation. The optic nerve sheaths were dissected and placed as a membrane in a two chamber diffusion cell to test meningeal penetration by both brimonidine alone and brimonidine encapsulated in nanoparticle (NP-brimonidine). Brimonidine concentration was assayed by UV–vis spectrometer measurement of absorbance at 389 nm. Increasing concentration of brimonidine on the receiver side of the chamber was measured in both the brimonidine alone and the brimonidine encapsulated experiments. The human data were fitted with a two parameter exponential regression analysis (brimonidine alone donor r2 = 0.87 and receiver r2 = 0.80, NP-brimonidine donor r2 = 0.79 and receiver r2 = 0.84). Time constant (τ) was 10.2 h (donor) and 13.1 h (receiver) in the brimonidine study, and 24.0 h (donor) and 15.9 h (receiver) in the NP-brimonidine study. Encapsulated brimonidine had a longer time to reach equilibrium. Passage of brimonidine through the optic nerve sheath was demonstrated in the experiments. Increase in time constants when comparing the NP-brimonidine with the brimonidine curves in the human studiesindicates that diffusion is delayed by the initial parameter of drug being loaded in NP. Direct treatment of injured optic nerve axons may be possible by trans-meningeal drug diffusion.

An extensive study of electrochemical behavior of brimonidine and its determination at glassy carbon electrode

The electrochemical behavior of brimonidine (BRIM), an antiglaucoma agent applied in therapy for lowering high intraocular pressure, was investigated by cyclic voltammetry, differential pulse voltammetry and square wave voltammetry using a glassy carbon electrode (GCE).The reduction of BRIM occurs as one-step quasi-reversible reaction in acid and neutral medium, reaching the full reversibility in alkaline solutions. Reduction process involves the transfer of two electrons and two protons at the pyrazine ring of quinoxaline moiety, forming a dihydro-derivative. In acid and neutral solutions, brimonidine reduction product is partly oxidized to its hydroxy-derivative. BRIM is also oxidized irreversibly with the transfer of one electron and one proton at secondary amine moiety. The effects of pH of the electrolyte solution, scan rate and BRIM concentration were monitored. The nature of the electrode process was found to be controlled by the adsorption at pH>6 and the total surface concentration of brimonidine adsorbed onto the GCE surface at pH 7, ΓBRIM=1.35×10−10molcm−2 was obtained. Based on this study, differential pulse voltammetric method was developed, validated and suggested for rapid electroanalytical determination of the low concentration of brimonidine. The linearity was achieved within the concentration range from 5×10−7 to 5×10−6M with LOD=1.6×10−7M and LOQ=5.3×10−7M. The method was applied for brimonidine determination in pharmaceutical dosage form, eye drops.

An Electroanalytical Approach to Brimonidine at Boron Doped Diamond Electrode Based on Its Extensive Voltammetric Study

AbstractAn extensive electrochemical study of brimonidine at boron doped diamond electrode (BDDE) was done by applying cyclic voltammetry (CV) and square‐wave voltammetry (SWV) in sulfuric acid of different concentrations (pH ranged from 0.6 to 1.6), and in BR buffer (pH ranged from 2.0 to 9.0). It was found that the reduction of brimonidine occurred in a one‐step quasi‐reversible mechanism, involving the transfer of two electrons and two protons. The reduction process took place at the quinoxaline ring and the corresponding mechanism of reduction was confirmed to be the same as for the other quinoxaline derivatives. The nature of the electrode process was found to be diffusion controlled in acid medium, while in a more alkaline medium a certain degree of adsorption was noticed. Based on this study, two sensitive voltammetric methods, differential pulse (DPV) and square wave (SWV) were developed, fully validated and suggested for rapid electroanalytical determination of low concentrations of brimonidine. The linearity was achieved within the concentration range from 2×10−6 M to 3×10−5 M for DPV (LOD=6.31×10−7 M, LOQ=2.1×10−6 M) and from 5×10−7 M to 1.5×10−5 M for SWV (LOD=1.28×10−7 M, LOQ=4.28×10−7 M). The methods were applied for brimonidine determination in pharmaceutical dosage form, eye drops. The obtained good recoveries suggested these simple and accurate methods for quality control of brimonidine in dosage form. Due to its high sensitivity, the SWV method could be a good alternative for determination of low concentrations of brimonidine, even in biological samples.

Hybrid Dendrimer Hydrogel/PLGA Nanoparticle Platform Sustains Drug Delivery for One Week and Antiglaucoma Effects for Four Days Following One-Time Topical Administration

We report a novel hybrid polyamidoamine (PAMAM) dendrimer hydrogel/poly(lactic-co-glycolic acid) (PLGA) nanoparticle platform (HDNP) for codelivery of two antiglaucoma drugs, brimonidine and timolol maleate. This platform was not cytotoxic to human corneal epithelial cells. Cellular uptake of Nile red-encapsulating PLGA nanoparticles was significantly increased by dendrimer hydrogel. A prolonged residence time of nanoparticles was demonstrated through investigation of FluoSpheres loaded into dendrimer hydrogel. Both brimonidine and timolol maleate were slowly released in vitro over a period of 28-35 days. Following topical administration of one eye drop (30 μL of 0.7% w/v brimonidine and 3.5% w/v timolol maleate) in normotensive adult Dutch-belted male rabbits, the HDNP formulation resulted in a sustained and effective IOP reduction (18% or higher) for 4 days. Furthermore, the HDNP maintained significantly higher concentrations of brimonidine in aqueous humor and cornea as well as timolol maleate in the aqueous humor, cornea, and conjunctiva up to 7 days as compared to saline, DH, and PLGA nanoparticle dosage forms, without inducing ocular inflammation or discomfort. Histological analysis of the cornea and conjunctiva did not reveal any morphological or structural changes. Our work demonstrated that this new platform is capable of enhancing drug bioavailability and sustaining effective IOP reduction over an extended period of time. This newly developed platform can greatly reduce dosing frequency of topical formulations, thus, improving long-term patient compliance and reducing enormous societal and economic costs. Given its high structural adaptability, many other chronic ocular diseases would benefit from long-lasting drug delivery of this new platform.

Effect of brimonidine tartrate 0.10% ophthalmic solution on pupil diameter

To evaluate the effect of brimonidine tartrate 0.10% ophthalmic solution on pupil diameter under light and dark luminance conditions. Ophthalmology Department, Tel Aviv Medical Center, Tel Aviv, Israel. Case series. The pupil diameter was measured with a Colvard pupillometer in eyes of healthy volunteers under light (5.0 candelas [cd]/m(2)) and dark (0.0 cd/m(2)) luminance conditions before brimonidine 0.10% instillation and after 30 minutes, 3 hours, and 6 hours. The mean age of the 26 volunteers (52 eyes) was 34.73 years (range 19 to 60 years). Under light conditions, the mean pupil diameter was 4.98 mm ± 0.83 (SD) before instillation of brimonidine 0.10% and 4.64 ± 0.82 mm after instillation. The difference was not statistically significant, with 13.4% of eyes having a clinically significant reduction (>1.0 mm) in pupil diameter after 6 hours. Under dark conditions, the mean pupil diameter was 6.76 ± 1.08 mm before instillation of brimonidine 0.10% and 5.30 ± 0.85 mm after instillation; the difference was statistically significant (P<.001). There was a clinically significant reduction in pupil diameter after 6 hours in 90.4% of eyes. The antimydriatic effect under dark luminance conditions was more pronounced in eyes with light irides. Brimonidine 0.10% ophthalmic solution had an antimydriatic effect under dark luminance conditions and had a negligible effect on pupil diameter under light luminance conditions. The effect was similar to that reported in studies using higher concentrations of brimonidine.

Brimonidine in the treatment of glaucoma and ocular hypertension

Treatment in glaucoma aims to lower intraocular pressure (IOP) to reduce the risk of progression and vision loss. The alpha2-adrenergic receptor agonist brimonidine effectively lowers IOP and is useful as monotherapy, adjunctive therapy, and replacement therapy in open-angle glaucoma and ocular hypertension. A fixed combination of brimonidine and timolol, available in some countries, reduces IOP as effectively as concomitant therapy with brimonidine and timolol and offers the convenience of 2 drugs in a single eyedrop. Brimonidine is safe and well tolerated. Its most common side-effects are conjunctival hyperemia, allergic conjunctivitis, and ocular pruritus. The newest formulation of brimonidine, brimonidine-Purite 0.1%, has a higher pH to improve the ocular bioavailability of brimonidine. This formulation contains the lowest effective concentration of brimonidine and is preserved with Purite(R) to enhance ocular tolerability. Brimonidine-Purite 0.1% is as effective in reducing IOP as the original brimonidine 0.2% solution preserved with benzalkonium chloride. Recent results from preclinical and clinical studies suggest that brimonidine may protect retinal ganglion cells and their projections from damage and death independently of its effects on IOP. The potential for neuroprotection with brimonidine is an added benefit of its use in glaucoma and ocular hypertension.

Vitreous Concentration of Topically Applied Brimonidine-Purite 0.15%

The aim of this study was to determine the vitreous brimonidine concentration of topically applied brimonidine-Purite 0.15%. Patients scheduled for a pars plana vitrectomy were invited to participate in this study after institutional review board (IRB) approval was obtained. Each patient was asked to apply brimonidine-Purite (0.15%) drops in the designated eye either every 12 h (b.i.d.; 9 patients) or every 8 h (t.i.d.; 10 patients) for the 2 weeks proceeding scheduled surgery. The importance of the last topical dose being 12 h (b.i.d. group) or 8 h (t.i.d. group) before the scheduled surgery, was emphasized. Four (4) patients served as controls and did not receive any drops. Vitreous (approximately 0.5-1.0 mL) was aspirated prior to opening the infusion line. Specimens were frozen at -68 degrees C until analyzed. In the b.i.d. group, the mean concentration of brimonidine was 16.74 nM+/-10.33 standard deviation (range, 0.42-34.68 nM; median, 16.38); in the t.i.d. group, the mean concentration of brimonidine was 19.16 nM+/-15.40 standard deviation (range, 0.22-39.48 nM; median, 16.98). A significant difference was observed between the (no drug) control vitreous brimonidine levels and b.i.d. or t.i.d. vitreous levels (P<0.01, <0.01, respectively; n=4, 9, and 10, respectively); and in brimonidine levels between t.i.d. phakic patients and t.i.d. patients with posterior chamber lens (P=0.04; n=4 and 6, respectively). Brimonidine-Purite 0.15%, topically applied b.i.d. or t.i.d. for 2 weeks prior to collection, acquired vitreous levels of brimonidine at or above the 2-nM concentration known to activate the neuroprotective alpha-2 receptor in animals.

Brimonidine evokes heterogeneous vasomotor response of retinal arterioles: diminished nitric oxide-mediated vasodilation when size goes small

Brimonidine, an alpha2-adrenergic receptor (AR) agonist, has been employed in the treatment of glaucoma due to its beneficial effects on intraocular pressure reduction and neuroprotection. In addition, some studies have implicated that brimonidine might influence ocular blood flow; however, its effect on the retinal microcirculation has not been documented. Herein, we examined the vasomotor action of brimonidine on different branching orders of retinal arterioles in vitro and determined the contribution of the alpha2-AR subtype and the role of endothelium-derived nitric oxide (NO) in this vasomotor response. First- and second-order retinal arterioles of pigs were isolated, cannulated, and pressurized for functional studies. Videomicroscopic techniques were employed to record diameter changes in response to brimonidine. RT-PCR was performed for detection of alpha-AR and endothelial NO synthase (eNOS) mRNA in retinal arterioles. All first-order arterioles (82 +/- 2 microm ID) dilated dose dependently to brimonidine (0.1 nM to 10 microM) with 10% dilation at the highest concentration. Second-order arterioles (50 +/- 1 microm ID) responded heterogeneously with either dilation or constriction. The incidence and magnitude of vasoconstriction were increased with increasing brimonidine concentration. Administration of the NO synthase inhibitor NG-nitro-L-arginine methyl ester abolished the brimonidine-induced vasodilation in first- and second-order arterioles. Regardless of vessel size, vasomotor responses (i.e., vasodilation and vasoconstriction) of retinal arterioles were sensitive to the alpha2-AR antagonist rauwolscine. Consistent with the functional data, alpha2A-AR and eNOS mRNAs were detected in retinal arterioles. Collectively, our data demonstrate that brimonidine at clinical doses evokes a consistent NO-dependent vasodilation in first-order retinal arterioles but a heterogeneous response in second-order arterioles. These vasomotor responses are mediated by the activation of alpha2-AR. It appears that brimonidine, depending on the concentration and vessel size, may alter local retinal blood flow.

Role of Alpha-2 Agonists in Neuroprotection

Four criteria are used to evaluate the potential usefulness of an agent for neuroprotection in glaucoma: 1) the agent must have a target in the retina; 2) it must be neuroprotective in animal models; 3) it must reach neuroprotective concentrations in the posterior segment after clinical dosing; and finally, 4) it must be shown to be neuroprotective in clinical trials. The alpha-2 adrenergic agonist brimonidine has met the first three criteria and clinical trials to establish the fulfillment of the fourth criterion are ongoing. The effects of brimonidine are mediated by its interaction with alpha-2 adrenergic receptors that are present in the retina. Activation of alpha-2 receptors by brimonidine has been shown to effectively promote the survival and function of retinal ganglion cells in a variety of animal models of optic injury relevant to glaucoma such as the chronic ocular hypertensive rat and rat optic nerve crush. Brimonidine has also been shown to be neuroprotective in the rat ischemia reperfusion model that evaluates general hypoxic damage to the whole retina. Clinical dosing of the topical formulation of brimonidine results in brimonidine concentrations in the posterior segment that are sufficient for both pharmacological activity at alpha-2 adrenergic receptors and neuroprotection. Finally, clinical trials are in progress to investigate the ability of brimonidine to protect human retinal ganglion cells and the visual field in glaucoma-related disease.

Up-regulation of brain-derived neurotrophic factor expression by brimonidine in rat retinal ganglion cells.

Brimonidine tartrate ophth, an alpha(2)-adrenergic agonist, is widely used as an antiglaucoma agent for lowering intraocular pressure. Recent studies suggest that brimonidine may be neuroprotective for retinal ganglion cells (RGCs) following optic nerve crush injury. Brain-derived neurotrophic factor (BDNF), a potent neuroprotective factor present in the RGCs, promotes RGC survival in culture and following optic nerve injury. We tested the hypothesis that a possible mechanism of brimonidine neuroprotection is through up-regulation of endogenous BDNF expression in the RGCs. A single dosage of brimonidine tartrate ophth solution (0.85-34 microM) was injected intravitreally into Sprague-Dawley rat eyes. The fellow eyes of each animal were injected with balanced salt solution (BSS) and used as control eyes. To determine BDNF messenger RNA expression, animal eyes were enucleated and processed for in situ hybridization, or retinas were isolated and processed for Northern blot analysis using rat BDNF radiolabeled riboprobes. In the control eyes injected with saline, BDNF was present in a minority of the RGCs. Two days after brimonidine injection, the number of BDNF-positive RGCs was increased from 55% to 166%, depending on brimonidine concentrations, when compared with those in the controls. In addition, the BDNF signal intensities in individual RGCs were elevated 50% in brimonidine-injected eyes compared with control eyes. Northern blot revealed a 28% increase of BDNF expression in the brimonidine group compared with the controls (P <.003). No significant difference was observed in BDNF receptor, trk B, expression between brimonidine, or BSS control groups. A single dose of a low concentration of intravitreal brimonidine is sufficient to significantly increase endogenous BDNF expression in RGCs. These results suggest that brimonidine neuroprotection may be mediated through up-regulation of BDNF in the RGCs. The BDNF should be further investigated regarding its role in the neuroprotective effects reported with brimonidine. Brimonidine may be (potentially) used clinically as a neuroprotective agent in optic neuropathy, including glaucoma, and ischemic and traumatic optic neuropathy.

Distribution of brimonidine into anterior and posterior tissues of monkey, rabbit, and rat eyes.

The objectives of the study were to evaluate the distribution of brimonidine (alpha2-adrenergic agonist) into anterior and posterior ocular tissues. Single or multiple doses of a 0.2 or 0.5% brimonidine tartrate solution were administered to one or both eyes of monkeys or to one eye of rabbits. Brimonidine was administered intraperitoneally to rats. After topical administration, [14C]brimonidine was rapidly absorbed into the cornea and conjunctiva and distributed throughout the eye. [14C]Radioactivity was higher and cleared more slowly in pigmented tissues (iris/ciliary body, choroid/retina, and optic nerve) than in nonpigmented tissues. Single and multiple dosing led to a similar drug distribution, with higher levels of brimonidine measured in pigmented tissues after multiple dosing. Most of the radioactivity extracted from ocular tissues represented unchanged brimonidine. In the rabbits and the monkey treated in only one eye, levels of radioactivity in the untreated eye were low, consistent with the low systemic levels and rapid drug clearance. Posterior ocular tissue concentrations of radioactivity exceeded systemic blood concentrations. The vitreous humor brimonidine concentrations in monkeys treated topically with 0.2% brimonidine tartrate was 82 +/- 45 nM. Vitreous levels in rabbits confirmed the penetration of brimonidine to the posterior segment. Similar concentrations of brimonidine (22 to 390 nM) were measured in the vitreous and retina of rats injected intraperitoneally with brimonidine. Both topically applied and systemically administered brimonidine reach the back of the eye at nanomolar concentrations sufficient to activate alpha2-adrenergic receptors. The brimonidine levels achieved at the retina are relevant for neuroprotection models.

Brimonidine Tartrate: A One-month Dose Response Study

Brimonidine tartrate is a relatively selective alpha2-agonist that effectively reduces mean intraocular pressure (IOP) and the incidence of IOP spikes after laser trabeculoplasty. The authors were interested in evaluating the dose response of brimonidine when applied topically for a longer duration in patients with elevated IOPs. The authors conducted a 1-month, multicentered, double-masked, randomized, placebo-controlled, parallel clinical study in 194 patients with ocular hypertension or glaucoma (mean IOP, 25.6 +/- 3.2 mmHg). The authors administered three concentrations of brimonidine (0.08%, 0.2%, and 0.5%) or placebo bilaterally every 12 hours for 1 month. The authors evaluated the following parameters: IOP, heart rate, blood pressure, visual acuity, pupil size, basal tear secretion as well as patient comfort at baseline, day 1, week 1, week 3, and week 4. All concentrations of brimonidine significantly reduced IOP, compared to baseline and placebo, at all follow-up visits. Maximum mean IOP decreases from baseline of 20.8%, 27.2%, and 30.1% were observed for the 0.08%, 0.20%, and 0.5% treatment groups, respectively. On days 1 and 21, the 0.2% and 0.5% treatment groups exhibited significantly greater IOP decreases than did the 0.08% group. After the initial steep decline in IOP, the effect decreased slightly and stabilized at day 14 at the level that was maintained throughout the study. The most frequent side effects reported were fatigue and dry mouth. No significant changes in heart rate were reported. Statistically significant decreases in mean blood pressure without clinical symptoms were observed within the 0.2% and 0.5% treatment groups. Brimonidine 0.2% is well tolerated, efficacious, and shows potential as an agent in the long-term treatment of elevated IOP.

Measurement of brimonidine concentrations in human plasma by a highly sensitive gas chromatography/mass spectrometric assay

Brimonidine is an α 2-adrenergic agonist that is efficacious in lowering intraocular pressure in humans. A highly sensitive and selective gas chromatography/mass spectrometry (GC/MS) assay is described for quantitation of brimonidine in human plasma following ocular installation. Brimonidine in 1 ml of plasma was extracted together with tetradeuterated brimonidine (internal standard) and clonidine (carrier) by solvent extraction. After solvent evaporation, 3,5-bis(trifluoromethyl)benzoyl derivatives were formed and injected onto a GC/MS appartus under negative chemical ionization conditions. The ions monitored for derivatized brimonidine and tetradeuterated brimonidine were m/ z 691 [M - HBr] and m/ z 694 [M - DBr], respectively. Calibration curves were linear from 2 to 1000 pg ml −1 ( r 2 = 0.981-0.996). The method was specific for brimonidine relative to endogenous compounds in plasma. The inter-day relative standard deviation for analysis of quality controls was 12% or less, and the inter-day assay accuracy ranged from 97 to 104% of nominals. The GC/MS assay showed adequate sensitivity for analysis of human samples from volunteers ocularly dosed with 0.5% brimonidine tartrate solution. Overall, the GC/MS assay showed excellent precision and accuracy, and a minimum quantifiable concentration of 2 pg ml −1.