11961 - Special Methodologies in Pharmaceutical Analysis (A)

Learning outcomes

The aim of the course is to provide the knowledge about analytical methods (e.g., UV spectrophotometry, chromatography and titrimetry) in qualitative and quantitative determination of pharmaceutical compounds. In particular, attention will be paid to the pharmacopoeial assays. The course includes laboratory activity.

Course contents

Theory
Non-aqueous titrations. Classification of the solvents used in non-aqueous titrations. Autoprotolysis constant and dielectric constant of solvents used in non-aqueous titrimetry. Apparent pH (pH*). Solvolysis reaction of weak electrolytes. Pharmacopoeial applications of non-aqueous titrations.
Potentiometry. Theory: Nernst equation; reference electrodes and working electrodes. The glass electrode for pH measurement (pH-meter); ion-sensitive electrodes in pharmaceutical analysis. Potentiometric titrations. Diazotisation titration and Karl-Fisher titration.
Conductimetry. Theory: electrical conductivity of electrolytic solutions. Conductance cell. Specific conductivity and equivalent conductivity. Conductimetric titration techniques.
Ultraviolet and visible spectrophotometry. Keypoints. Factors governing absorption of radiation in the UV/vis region. Beer-Lambert law. Instrumentation. UV spectra of some representative drugs. Deconvolution of spectra from binary mixtures of drugs. Derivative spectrophotometry. Difference spectrophotometry. Spectrophotometric derivatization reactions: Ellman reagent; Bratton-Marshall method in analysis of sulphamidics. Applications in pharmaceutical analysis.
Chromatographic techniques. The chromatographic separation: stationary phases in gaschromatography and liquid chromatography. Chromatographic parameters (retention, selectivity, efficiency, resolution). Kinetics of the chromatographic separations. Detection in HPLC and GC. Calibration and quantitation in chromatographic analysis (calibration graphs, single-point calibration, external standardization). Mass spectrometry and hyphenated chromatographic techniques: HPLC-MS and GC-MS.
Capillary electrophoresis (CE). Keypoints. Separation in CE; mobility of ions (apparent and effective mobility). Instrumentation. The electroosmotic flow (EOF). Effect of pH on ionization and mobility of analytes. Effect of pH on EOF. Control of separation in CE. Micellar electrokinetic chromatography (MEKC). Application of CE in pharmaceutical analysis.
Chiral analysis. Importance of chirality of the drugs. Enantioresolution methods (indirect and direct enantioresolution). Direct analytical enantioresolution in HPLC, GC and CE. Chiral selectors: cyclodextrins, linear polysaccharides, proteins etc..
Stability of drugs. Degradation of drugs by: hydrolysis, decarboxylation, photo-oxidation, photo-reduction, racemization etc.. Analytical methods in stability studies and kinetic of degradation of drugs.

Laboratory activity

- Non - aqueous titrimetry. Determination of weak acids (sulphamidics) and weak bases (sodium acetate, lidocaine hydrochloride, alanine).
- Potentiometric titration of phosphoric acid.
- Conductimetric titration of sodium acetate.
- Diazotisation titration of sulphamidics (pharmacopoeial assay by biamperometric end-point detection).
- UV/vis spectrophotometry. Calibration of absorbance scale by using a bichromate solution (calibration graph).
- Spectrophotometric UV-vis analysis of Fe(II) in tablets by complexation with o-fenantroline.
- Spectrophotometric UV-vis analysis of procaine by derivatization with Bratton-Marshall reagent.
- Derivative UV spectrophotometric analysis of salicylic acid as related substance in aspirin.
- HPLC analysis of salicylic acid as related substance in aspirin.
- TLC separation and identification of steroids.

Readings/Bibliography

1) V. CAVRINI, V. ANDRISANO, Analisi Farmaceutica, Metodi di riconoscimento e di separazione. Società Editrice Esculapio, Seconda Edizione, Bologna 2009.

2) D. C. HARRIS, Chimica Analitica Quantitativa II Ed. Zanichelli, 2005.

3) Slides from lessons.

Teaching methods

The theoretical lessons are intended to provide the basic knowledge of instrumental techniques used in pharmaceutical analysis. In the laboratory activity the students will apply analytical procedures for identification and quantitation of active ingredients in pharmaceuticals.

Assessment methods

During the lab activity the students will perform self-assessment experiments. At the end of the lab course the students should sit for a laboratory examination consisting in the application of methodologies for the analysis of active ingredients in real pharmaceutical samples.

The final test will be an oral presentation on subjects dealt during the course.

Teaching tools

The theoretical lessons will be held by using slides.

The lab activity will be carried out in laboratories equipped with specific reagents and analytical instrumentation (UV- and FTIR spectrophotometers, HPLC and GC apparatus, polarimeter, refractometer etc.), suitable for the drug quality control.

Office hours

See the website of Roberto Gotti