ORIGINAL ARTICLE Detection of Raman Spectra in Ocular Drugs for Potential In Vivo Application of Raman Spectroscopy Mari Elshout, Roel J. Erckens, Carroll A. Webers, Henny J. Beckers, Tos T. Berendschot, John de Brabander, Fred Hendrikse, and Jan S. Schouten Abstract Purpose: Raman spectroscopy holds potential for the assessment of intraocular pharmacokinetics. Raman spectra of ocular drugs were acquired, to determine the drug-specific Raman signature. The ability of the Raman technique to quantify drug concentrations was also investigated. Methods: The experimental setup was based on a High Performance Raman Module 2500 Raman module, designed for 180° ‘‘backscatter’’ signal detection in the wavenumber range of 400–1,800 cm - 1 . Excitation source was a diode laser emitting a beam with a wavelength of 785 nm and a power of 10 mW. Laser light was focused in the sample with a long-working-distance microscope objective (25 · /0.50). Samples were measured in quartz cuvettes in 10 sequential measurements, with exposure time 30 s. The total number of measured drugs was 49. To determine whether signal intensity and drug concentration correlate, 2 drugs were diluted in water and measured with 120 s exposure time at different concentrations. Results: An active ingredient–specific Raman signature was detected in 4 glaucoma drugs, 6 mydriatics, 5 anti- biotics, 4 anesthetics, 3 anti-inflammatory drugs, 2 types of artificial tears, and 5 other drugs. In 20 drugs, no specific Raman signature was detected. Linear correlation of drug concentration with signal intensity was high (R 2 ‡ 0.94). Conclusions: Using low laser powers, Raman signatures for 29 commonly used ocular drugs were detected. Correlation of drug concentration with signal intensity is high, which is essential for monitoring drug concen- tration in ocular media. The presented results encourage the use of Raman spectroscopy to acquire detailed information on the pharmacokinetics of these ocular drugs. Introduction F or many years, a wide range of drugs has been in use for diagnostic purposes or treatment of ocular diseases such as glaucoma, corneal ulcers, uveitis, endophthalmitis, and retina diseases. Although much data on their effective- ness are available, much less is known about the intraocular pharmacokinetics of these drugs in patients. Studies on ocular pharmacokinetics are limited by the invasiveness of current techniques. There are currently no methods available to measure the intraocular concentration of ocular drugs other than by invasively sampling aqueous or vitreous humor. This limits the maximum number of measurements and does not allow for real-time monitoring of drug concentration. Real-time monitoring would allow the ophthalmologist to optimize drug treatment efficacy for each individual patient and more precisely determine dosage and optimum time for administration of the drug. An approach capable of continuously quantifying local drug concentration within the eye would allow monitoring of drug concentra- tions. Further, application of a noninvasive approach is de- sirable, because it can eliminate the risks involved with invasively acquiring samples. 1 In the biomedical field, Raman spectroscopy can be used to detect the presence of molecules that exhibit unique Ra- man signatures. 1–6 Raman spectroscopy measures the shift in wavelength of incident light as it is scattered by molecules in the tissue or media. This shift is characteristic for the inter- molecular and intramolecular vibrational frequencies. 7 The Raman spectroscopic technique has several advan- tages: a noncontact character, real-time response, both qualitative and quantitative analysis capabilities, a high de- gree of specificity, and simultaneous multicomponent detection. 8 The possibility of continuously repeating mea- surements, which allows monitoring of drug concentration, is another major advantage of Raman spectroscopy. The technique is limited by the weak intensity of the Raman scattered light and/or the occurrence of fluorescence. 9 This technique has been already successfully used to monitor the presence of various concentrations of the antiviral drug Maastricht University Medical Center, University Eye Clinic Maastricht, Maastricht, The Netherlands. JOURNAL OF OCULAR PHARMACOLOGY AND THERAPEUTICS Volume 00, Number 00, 2011 ª Mary Ann Liebert, Inc. DOI: 10.1089/jop.2011.0018 1