Development and Validation of a Liquid Chromatography-Mass Spectrometry Based Analytical Assay for Determination of Cromolyn Sodium in Skin Permeation Studies
Faculty Sponsor’s Department
Pharmaceutical Sciences
Type
Oral Competitive
Project's Category
Chromatography
Abstract or Artist's Statement
Cromolyn sodium (CS) is a mast cell stabilizer which has been used to treat systemic mastocytosis, allergic- and exercise-induced asthma, and allergic reactions induced by atopic dermatitis. Presently, CS is administered orally and intranasally, and with a short half-life and poor absorption, 4 to 8 doses are required daily for treatment. Developing a transdermal product for CS would eliminate such drawbacks that lead to inconsistent patient dosing and provide sustained therapeutic effects when administered via skin. Our long-term goal is to determine the feasibility of delivering CS through skin. However, a prerequisite for evaluating the performance of any transdermal system is to have a sensitive analytical method that can selectively detect and quantify the drug without any interference from compounds that may leach from skin during permeation studies. Therefore, our preliminary goal was to develop and validate such a method that can be employed for transdermal studies. The optimized liquid chromatography-mass spectrometry (LC-MS) method utilized a chromatographic separation which involved an isocratic mobile phase (10mM ammonium bicarbonate, pH 8.0, 90% and acetonitrile, 10%) at a flow rate of 0.2500 mL/min. Detection involved direct MS/MS channels with m/z 467.0255 (precursor) and m/z 379.0517 (fragment) with argon as the collision gas. CS calibrants were prepared in phosphate-buffered saline (PBS), pH 7.4, for validation (0.1, 0.25, 0.5, 0.75, 1, and 2.5 μg/mL). To ensure no skin interference, dermatomed porcine ear skin was minced, placed in PBS, and shaken for 15 hours to extract any possible interfering components. The extract was filtered and analyzed with the optimized LC-MS conditions. Calibrants were also analyzed over 3 days with each day examining 6 injections (20 μL) of each sample. Peak areas determined by LC-MS were used to construct calibration curves for CS and to calculate % error and % RSD to elucidate accuracy and precision of the method. Results showed CS retention time to be around 4.4 minutes with no interfering peak from skin extract, and linearity was observed between 0.1-2.5 μg/mL. Inter- and intra-day accuracy and precision of the method were within the acceptable limit of ±20% at the lower limit of quantitation and ±15% at other concentrations. Future studies will involve using the validated method for quantification of CS in skin permeation studies to investigate our long-term goal.
Development and Validation of a Liquid Chromatography-Mass Spectrometry Based Analytical Assay for Determination of Cromolyn Sodium in Skin Permeation Studies
Cromolyn sodium (CS) is a mast cell stabilizer which has been used to treat systemic mastocytosis, allergic- and exercise-induced asthma, and allergic reactions induced by atopic dermatitis. Presently, CS is administered orally and intranasally, and with a short half-life and poor absorption, 4 to 8 doses are required daily for treatment. Developing a transdermal product for CS would eliminate such drawbacks that lead to inconsistent patient dosing and provide sustained therapeutic effects when administered via skin. Our long-term goal is to determine the feasibility of delivering CS through skin. However, a prerequisite for evaluating the performance of any transdermal system is to have a sensitive analytical method that can selectively detect and quantify the drug without any interference from compounds that may leach from skin during permeation studies. Therefore, our preliminary goal was to develop and validate such a method that can be employed for transdermal studies. The optimized liquid chromatography-mass spectrometry (LC-MS) method utilized a chromatographic separation which involved an isocratic mobile phase (10mM ammonium bicarbonate, pH 8.0, 90% and acetonitrile, 10%) at a flow rate of 0.2500 mL/min. Detection involved direct MS/MS channels with m/z 467.0255 (precursor) and m/z 379.0517 (fragment) with argon as the collision gas. CS calibrants were prepared in phosphate-buffered saline (PBS), pH 7.4, for validation (0.1, 0.25, 0.5, 0.75, 1, and 2.5 μg/mL). To ensure no skin interference, dermatomed porcine ear skin was minced, placed in PBS, and shaken for 15 hours to extract any possible interfering components. The extract was filtered and analyzed with the optimized LC-MS conditions. Calibrants were also analyzed over 3 days with each day examining 6 injections (20 μL) of each sample. Peak areas determined by LC-MS were used to construct calibration curves for CS and to calculate % error and % RSD to elucidate accuracy and precision of the method. Results showed CS retention time to be around 4.4 minutes with no interfering peak from skin extract, and linearity was observed between 0.1-2.5 μg/mL. Inter- and intra-day accuracy and precision of the method were within the acceptable limit of ±20% at the lower limit of quantitation and ±15% at other concentrations. Future studies will involve using the validated method for quantification of CS in skin permeation studies to investigate our long-term goal.