Nature’s Treatment for Irritable Bowel Syndrome: Studies on the Isolation of (-)-Menthol from Peppermint Oil and Its Conversion to (-)-Menthyl Acetate Maeve Egan, E ́ ilis Margaret Connors, Zeeshan Anwar, and John J. Walsh* School of Pharmacy and Pharmaceutical Sciences, Trinity College Dublin, Dublin 2, Ireland * S Supporting Information ABSTRACT: A simple, robust, and reproducible method was developed for the isolation of (-)-menthol from peppermint oil and to study the effect of different types of leaving groups, catalysts, solvents, and tertiary base on the extent of esterification of (-)-menthol to (-)-menthyl acetate. In this experiment, students compare leaving group properties of acetate and chloride ions from the acylating reagents acetic anhydride and acetyl chloride, respectively. The extent of conversion is compared when pyridine and 4-(dimethylamino)pyridine are used as catalysts, when N,N-diisopropylethylamine is used as tertiary base, and when the solvent is changed from dichloromethane to diethyl ether to N,N-dimethylformamide. Students are assessed on the chromatographic/spectroscopic purity and yield of (-)-menthol isolated and on their understanding of the factors that affect its extent of conversion to (-)-menthyl acetate. Full spectral characterization of both compounds is also conducted. They also complete a series of answers to questions based on lecture material presented on this topic, complete a crossword as a formative assessment tool, and are required to present a PowerPoint slide to their peers on a particular aspect of the bench to bedside development of peppermint oil for the treatment of irritable bowel syndrome. KEYWORDS: Upper-Division Undergraduate, Laboratory Instruction, Organic Chemistry, Collaborative/Cooperative Learning, Hands-On Learning/Manipulatives, Chromatography, Esters, Medicinal Chemistry, Natural Products, NMR Spectroscopy ■ INTRODUCTION Many natural plant products have been used in the treatment of a diverse selection of ailments and diseases since ancient times. This experiment provides a valuable learning experience to students in the use of peppermint oil (PO) as an herbal medicine and the application of key analytical techniques frequently used in the laboratory. The primary isolation technique utilized in this experiment is flash column chromatography, allowing students the opportunity to master the skills required to achieve precise isolation of a substance of interest from a complex mixture. In converting (-)-menthol to (-)-menthyl acetate, students gain an understanding of the esterification process and the factors affecting its trans- formation. Students interpret IR, HRMS, and NMR spectra as a means to compare and contrast structural similarities and differences between (-)-menthol and (-)-menthyl acetate. This experiment complements many other laboratory based experiments on the isolation of Nature’s medicine including cinchonine and quinine from Cinchona calisaya, 1 valtrate from Centranthus ruber, 2 galantamine from Leucojum aestivum, 3 lovastatin from red yeast rice, 4 parthenolide from Tanacetum parthenium, 5 and hyperforin from Hypericum perforatum. 6 This experiment offers an interesting comparison to the chromato- graphic separation of (-)-menthol from consumer products 7 and also provides a new insight into a variety of chromato- graphic, spectroscopic and polarimetric analyses 8,9 previously conducted on PO and (-)-menthol. ■ BACKGROUND Peppermint (Mentha × piperita L.)(Figure 1) is a perennial herb first described in 1696 by John Ray and is cultivated in many parts of the world. 10,11 Background information on Mentha × piperita L., quality, efficacy, and safety standards of peppermint oil and, in particular, the significance of peppermint in the treatment of irritable bowel syndrome (IBS) and other therapeutic applications is highlighted using a cyclical model (Figure 2). ■ EXPERIMENTAL OVERVIEW The aim of this experiment is to isolate (-)-menthol from PO, to explore the esterification of (-)-menthol to (-)-menthyl acetate, and to confirm the identity of both compounds. The experiment and workshop are intended to complement the more theoretical aspects of the PO course delivered to our students. A comprehensive overview of the learning outcomes for the PO course, teaching methods and assessment criteria are shown in Table 1. The experimental component can be conducted over two, 3 h laboratory periods with a follow-on workshop on spectral assignment. It has been completed by 30 Laboratory Experiment pubs.acs.org/jchemeduc © XXXX American Chemical Society and Division of Chemical Education, Inc. A DOI: 10.1021/ed5007037 J. Chem. Educ. XXXX, XXX, XXX-XXX