Instrumental analysis of drugs A - L
Academic Year 2022/2023 - Teacher: Angelo SPADAROExpected Learning Outcomes
Applying knowledge and understanding. The student must be able to use the knowledge acquired in the context of instrumental analysis to develop and validate analysis protocols for the analysis of active ingredients in pharmaceutical forms and biological matrices.
Making judgments: The student will have to interpret the experimental results obtained relating to the analysis of active ingredients present in complex matrices.
Communication skills. At the end of the course the student will have to explain what has learned using appropriate terminology for a correct scientific presentation regarding the analytical techniques and the analysis of compounds of pharmaceutical interest.
Lifelong learning skills. The student will have to developed useful theoretical/practical skills in order to update and expand his knowledge with the aim of face new analytical problems.
Course Structure
Theoretical lessons (35 hours) in the classroom with the aid of video projection systems of slides and videos. Group exercises (12 hours) for small groups of students (10-12) on high performance liquid chromatographs (HPLC, HPLC-IEC).
If the teaching is given in a mixed or remote mode, the necessary changes with respect to what was previously stated may be introduced, in order to comply with the program envisaged and reported in the syllabus.
Information for students with disabilities and / or SLD
To guarantee equal opportunities and in compliance with the laws in force, interested students can ask for a personal interview in order to plan any compensatory and / or dispensatory measures, based on the didactic objectives and specific needs. It is also possible to contact the CInAP contact person (Center for Active and Participatory Integration - Services for Disabilities and / or SLD) of the Department of Pharmaceutical and Health Sciences.
Required Prerequisites
Attendance of Lessons
Detailed Course Content
LECTURES
Introduction to instrumental methods of analysis: classification and selection of analytical methods. Chromatographic techniques. General principles of the chromatographic process and chromatographic parameters. Distribution constant. Retention time. Capacity factor. Selectivity. Efficiency: plate theory; van Deemter's equation; extra-column factors. Asymmetry of chromatographic peaks. Resolution. Liquid chromatography: stationary phase, mobile phase. Liquid-solid chromatography, liquid-liquid chromatography, ion exchange chromatography, ion pair chromatography, molecular exclusion chromatography, affinity chromatography. Column chromatography, thin layer chromatography (TLC). High performance liquid chromatography (HPLC): instrumentation. Gas chromatography (GC): instrumental aspects, types of stationary phase, detection in GC. Enatioselective chromatography separation: direct method with use of enantioselective stationary phases; indirect method with derivatization with defined stereochemical reagents. Capillary Electrophoresis (EC): instrumentation, theoretical principles, analytical parameters. Combined techniques: liquid chromatography and UV-VIS photodiode detector (HPLC-DAD), liquid chromatography and mass spectrometry (HPLC-MS), gas chromatography and mass spectrometry (GC-MS). Qualitative and quantitative analysis in HPLC (external and internal standard methods). Sample preparation and extraction methods. Traditional extraction methods (liquid/liquid and liquid/solid). Extraction with supercritical fluids (SFE). Solid phase extraction (SPE). Preparation of samples from complex biological matrices. Validation of the analytical method: linearity, precision, accuracy, reproducibility, limit of detection or detention (LOD) and the limit of quantification (LOQ). Development of analytical methods for qualitative and quantitative identification of drugs in quality control and in pre-clinical and clinical research. Determination of drug/protein interactions, using high performance affinity chromatography (HPAC), useful for the development of new drugs.
LABORATORY PRACTICAL CLASSES
High performance liquid chromatography: development of an HPLC method for the separation of drugs in complex matrices. Study of efficiency, selectivity and resolution. Calculation of chromatographic parameters (capacity factor, selectivity factor, resolution, number of theoretical plates, peak asymmetry). Qualitative/quantitative HPLC determination of drugs in pharmaceutical forms present on the market. Extraction, purification and characterization of an active ingredient from a complex matrix.
Textbook Information
- V. Cavrini. Principi di Analisi Farmaceutica, terza edizione. Società Editrice Esculapio, Bologna.
- Veronika R. Meyer. Practical High-Performance Liquid Chromatography, 5th Edition. Wiley, New York (consultation text)
- J.F. Holler, S.R. Crouch. Fondamenti di chimica analitica di Skoog e West. EdiSES, III edizione 2015
- Teaching material provided by the teacher available on Studium.
Course Planning
| Subjects | Text References | |
|---|---|---|
| 1 | Introduction to instrumental analytical methods. Classification of instrumental analytical methods. Choice of the analytical method. Chromatographic techniques. General principles of the chromatographic process and chromatographic parameters. Constant of distribution. Retention time. Capacity factor. Selectivity. Efficiency: plate theory; van Deemter equation; extra-column factors. Chromatographic peak asymmetry. Resolution. | Book 1, 2, 3, and teaching material |
| 2 | Liquid chromatography: stationary phase, mobile phase. Liquid-solid chromatography, liquid-liquid chromatography, ion exchange chromatography, ion pair chromatography, molecular exclusion chromatography, affinity chromatography. Column chromatography, thin layer chromatography (TLC). High performance liquid chromatography (HPLC): instrumentation. | Book 1, 2, 3, and teaching material |
| 3 | Enatioselective chromatographic separation: direct method using enantioselective stationary phases; indirect method with derivatization with defined stereochemistry reagents. Gas chromatography (GC): instrumental aspects, types of stationary phase, detection in GC. Capillary electrophoresis (EC): instrumentation, theoretical principles, analytical parameters. | Book 1, 2, 3, and teaching material |
| 4 | Combined techniques: liquid chromatography and photodiode UV-VIS detector (HPLC-DAD), liquid chromatography and mass spectrometry (HPLC-MS), gas chromatography and mass spectrometry (GC-MS). | Book 1, 2, 3, and teaching material |
| 5 | Qualitative and quantitative analysis (external standard method, internal standard method). Sample preparation and extraction methods. Classic extraction methods (liquid / liquid and liquid / solid). Extraction with supercritical fluids (SFE). Solid phase extraction (SPE). Preparation of samples from complex biological matrices. Validation of the analytical method: linearity, precision, accuracy, reproducibility, limit of detection (LOD), limit of quantification (LOQ). | Book 1, 2, 3, and teaching material |
| 6 | Development of chromatographic methods for quality control and in pre-clinical and clinical research. Determination of drug / target protein interactions, by high performance affinity chromatography, for the development of new drugs. | Book 1, 2, 3, and teaching material |
| 7 | Laboratory exercises: Development of HPLC methods for the separation of drugs in complex matrices. Study of efficiency, selectivity and resolution as the stationary phase, the composition of the eluent phase and the flow vary. Calculation of chromatographic parameters (capacity factor, selectivity factor, resolution, number of theoretical plates, peak asymmetry). Qualitative / quantitative determination of molecules of pharmaceutical interest in pharmaceutical forms on the market. | Book 1, 2, 3, and teaching material |
Learning Assessment
Learning Assessment Procedures
Examples of frequently asked questions and / or exercises
Theoretical principles of chromatographic separations: approaches to improve resolution in liquid chromatography. Theoretical principles and applications of capillary electrophoresis. Sample preparation and extraction methods. Validation of the analytical method.