Glaucoma is a heterogeneous group of multifactorial optic neuropathies and the leading cause of irreversible blindness and visual impairment. Epidemiological data has estimated that in 2020 there will be more than 80 million individuals affected by the disease worldwide. Nowadays, intraocular pressure (IOP) lowering is carried out mainly by pharmacotherapy, with different drugs. The study of ocular pharmacokinetics of antiglaucoma drugs, crucial for better understanding of ocular distribution, bioavailability, and pharmacodynamic parameters, can benefit the development of antiglaucoma drugs or formulations. Bioanalysis of drugs in ocular matrices is still underestimated, since it is challenging and rarely performed. Therefore, this review summarized the chromatographic methods employed for the quantification of several antiglaucoma drugs in different ocular matrices, discussing bioanalytical steps, such as sample preparation, separation, and detection. Animals and matrices as well as the challenges faced in ocular bioanalysis were also discussed. Ocular bioanalysis has been performed mainly in rabbits, the most adequate animal model for ocular studies. The matrix most used is aqueous humor, because it is cleaner and easier to sample. Sample preparation was carried out primarily employing classic techniques, such as liquid-liquid extraction, protein precipitation, and solid-phase extraction, with conventional solvents and sorbents. Chromatographic separation was achieved predominantly by reversed-phase liquid chromatography. Ultraviolet spectrophotometry and tandem mass spectrometry prevailed for detection, although other techniques, such as fluorimetry, have also been used. It was evidenced that more efforts must be directed towards miniaturized, eco-friendly, and non-terminal sampling for sample preparation. In its turn, ultra high-performance liquid chromatography and mass spectrometry should gain prominence in ocular bioanalysis for separation and detection, respectively, since it combines high separation capacity with selectivity and sensitivity, in addition to being an environmental friendly approach.

青光眼是多因素视神经病变的异质性群体，是不可逆性失明和视力障碍的主要原因。流行病学数据估计，到2020年，全世界将有超过8000万人受到该疾病的影响。如今，降低眼压（IOP）的方法主要是通过药物治疗，并使用不同的药物。抗青光眼药物的眼药代动力学研究对于更好地了解眼的分布，生物利用度和药效学参数至关重要，可有助于抗青光眼药物或制剂的开发。眼基质中药物的生物分析仍然被低估了，因为它具有挑战性并且很少进行。因此，这篇综述总结了用于定量不同眼基质中几种抗青光眼药物的色谱方法，并讨论了生物分析步骤，例如样品制备，分离和检测。还讨论了动物和基质以及眼部生物分析面临的挑战。眼部生物分析主要在兔子中进行，兔子是进行眼部研究的最合适的动物模型。最常用的基质是房水，因为它更清洁且更易于采样。样品制备主要采用经典技术进行，例如液-液萃取，蛋白质沉淀和固相萃取，以及常规溶剂和吸附剂。色谱分离主要通过反相液相色谱实现。紫外分光光度法和串联质谱法主要用于检测，尽管也使用了其他技术，例如荧光法。有证据表明，必须更加努力地进行小型化，生态友好和非终端采样以进行样品制备。反过来，超高效液相色谱法和质谱法在眼用生物分析的分离和检测中也应得到重视，因为它不仅是一种环境友好的方法，而且兼具高分离能力，选择性和灵敏度。