Course Description
Separation, identification, and analysis of drugs by thin-layer, gas, and high performance liquid chromatography, supercritical fluid chromatography, and capillary electrophoresis.
Athena Title
Pharmaceutical Analysis I
Prerequisite
Permission of department
Semester Course Offered
Not offered on a regular basis.
Grading System
A - F (Traditional)
Course Objectives
a. To understand the basic principles of chromatography b. To define the techniques known as thin layer, gas and liquid chromatography c. To differentiate between the qualitative and quantitative aspects of thin layer, gas and liquid chromatography d. To calculate the parameters necessary for evaluation of the separation process e. To make changes in the chromatography process to enhance the separation f. To define the technique named supercritical fluid chromatography g. To apply the SFC techique to drugs separation and analysis h. To define the technique named capillary electrophoresis i. To apply the CE technique to drug separation
Topical Outline
1 Getting Started 1.1 Introduction 1.2 What Is Known Before Starting 1.3 Sample Pretreatment and Detection 1.4 Developing the Separation 1.5 Completing the HPLC Method 2 Basics of Separation 2.2 Resolution: General Considerations 2.3 Resolution as a Function of Conditions 2.4 Sample-Size Effects 3 Detection, Sensitivity and Selectivity 3.1 Introduction 3.2 UV Detection 3.3 Other HPLC Detectors 3.3.1 Universal Detection 3.3.2 Fluorescence Detection 3.3.3 Electrochemical Detection 3.3.4 Mass Spectrophotometer Detection (LC-MS) 3.3.5 Selecting the Mass Spectrometric Detector 3.3.6 Less Common Detectors 4 Sample Preparation 4.1 Introduction 4.2 Types of Samples 4.3 Preliminary Processing of Solid and Semi-solid Samples 4.4 Sample Pretreatment for Liquid Samples 4.5 Sample Pretreatment for Solid Samples 4.6 Column Switching 4.7 Derivatization 5 The Column 5.1 Introduction 5.2 Characteristics of Columns and Column Packings 5.2.1 Column-Packing Particles 5.3. Column Specifications 5.4 Column Problems and Remedies 6 Non-ionic Samples: Reversed- and Normal-Phase HPLC Part I: Reversed-Phase Chromatography 6.2 Retention in Reversed-Phase Chromatography 6.3 Selectivity in Reversed-Phase Chromatography 6.4 Optimizing the Separation of Non-ionic Samples in Reversed- Phase Chromatography 6.5 Non-aqueous Reversed-Phase HPLC Part II: Normal-Phase Chromatography 6.6 Retention and Selectivity in Normal-Phase Chromatography 6.7 Optimizing the Separation of Non-ionic Samples in Normal-Phase Chromatography 7 Ionic Samples: Reversed-Phase, Ion-Pair, and Ion-Exchange HPLC 7.1 Introduction 7.2 Acidic and Basic Samples 7.3 Optimizing the Reversed-Phase Separation of Ionic Samples 7.3.1 Initial Experiments 7.3.2 Controlling Selectivity 7.3.3 Special Problems 7.4 Ion-Pair Chromatography 7.4.1 Basis of Retention 7.4.2 Initial Experiments 7.4.3 Controlling Retention Range and Selectivity: Changes in % B, pH, and Ion-Pair Reagent Concentration 7.4.4 Other Changes in Selectivity 7.4.5 Special Problems 7.5 Ion-Exchange Chromatography 7.5.1 Basis of Retention 7.5.2 Method Development 7.5.3 Mixed-Mode Separations 7.5.4 Silica Columns 8 Systematic Approach to the Reversed-Phase Separation of Regular Samples 8.1 Introduction 8.1.1 Some Guiding Principles 8.2 Getting Started 8.2.1 Initial Conditions 8.2.2 Adjusting the Retention Range 8.2.3 Evaluating Peak Shape and Plate Number 8.3 Completing Isocratic Method Development 8.4 Alternative To Completing Isocratic Method Development 8.5 Completing Gradient Method Development 9 Biochemical Samples: Proteins, Nucleic Acids, Carbohydrates, and Related Compounds 9.1 Introduction 9.1.1 Primary Structure 9.1.2 Special Requirements of Biochemical HPLC 9.2 Separation of Peptide and Protein Samples 9.2.1 Reversed-Phase HPLC 9.2.2 Ion-Exchange HPLC 9.2.3 Hydrophobic Interaction Chromatography 9.3 Separation of Oligonucleotides 9.3.1 Ion-Pair HPLC 9.3.2 Ion-Exchange HPLC 9.4 Size-Exclusion Chromatography 9.4.1 The Basis of SEC Retention 9.4.2 Applications 9.4.3 Preferred Conditions for an SEC Separation 9.4.4 Common Problems and Remedies 9.4.5 Protein Folding 10 Chiral Separations 10.1 Introduction 10.1.1 Chiral Derivatization 10.1.2 Chiral Mobile-Phase Additives 10.1.3 Chiral Stationary Phases 10.1.4 Principles of Chiral Recognition 10.1.5 General Considerations of Chiral HPLC Method Development 10.1.6 Selecting a Chiral Column 10.2 Protein-Derived Chiral Stationary Phases for HPLC 10.2.1 Introduction 10.2.2 Background 10.2.3 Mechanism of Chiral Interactions 10.2.4 Characteristics of Protein-Based Chiral Columns 10.2.5 Adjusting Retention and Selectivity with the Mobile Phase 10.2.6 Experimental Parameters 10.2.7 Application and Special Techniques 10.2.8 Systematic Method Development 10.3 Polysaccharide (Carbohydrate) Columns 10.3.1 Introduction 10.3.2 Properties of Commercial Polysaccharide Phases 10.3.3 Mechanisms of Chiral Interactions 10.3.4 Experimental Parameters 10.3.5 Applications 10.3.6 Strategy for Method Development 10.4 Donor-Acceptor (Pirkle) Columns 10.4.1 Introduction 10.4.2 Properties of Commercial Donor-Acceptor CSPs 10.4.3 Mobile-Phase Conditions 10.4.3.1 Solvents 10.4.4 Method Development with Pirkle CSPs 10.5 Cavity-Type Columns 10.5.1 Introduction 10.5.2 Method Development for Separations Using Underivatized CD Columns 10.5.3 Method Development With Derivatized Cyclodextrins Notice: The course syllabus is a general plan for the course; deviations announced to the class by the instructor may be necessary. Principal course assignments: Each student is required to present a 20-30 minute class presentation on some aspect of chromatography with drug systems. Specific course requirements: There will be two (2) take-home exams with 75% of the course grade. The remaining 25% of the grade is based on the classroom presentation. Grading Policy: The standard A - F grading system will be followed. Attendance Policy: A student is expected to be present at each scheduled class period. The student must notify the instructor prior to the class period by email or telephone. Failure to notify the instructor will result in a drop of one letter grade in the final course. Text: PRACTICAL HPLC METHOD DEVELOPMENT, 2nd Edition, Snyder, Kirkland, and Glajch, Wiley-Interscience Supplemental Text: Analytical Supercritical Fluid Chromatography and Extraction, edited by M Lee and K Markides, Chromatography Conferences Inv., Provo, UT 1990. Supplmental Text: High Performance Capillary Electrophoresis, Theory, Techniques and Applications, edited by MG Khaledi, Volume 146 in Chemical Anlaysis, Wiley-Interscience, 1998. Policy for make-up examinations: Make-up exams will be allowed provided the student notifies the instructor prior to the exam period. The exam will be rescheduled soon after the illness or other outside event has concluded.