PFT871 - Method Validation By HPLC

Outline info
Last revision date 2018-07-20 11:50:00.422
Last review date 2018-07-20 11:50:30.555

Subject Title
Method Validation By HPLC

Subject Description
This subject deals strictly with HPLC method validation elements such as system precision, linear detector response, accuracy, limit of quantitation, limit of detection, ruggedness and robustness. (Lecture and Lab)

Credit Status
One credit toward the Advanced Pharmaceutical Instrumentation (HPLC) Recognition of Achievement

Learning Outcomes
Upon successful completion of this subject the student will be able to:

1. Demonstrate an understanding of analytical method validation elements outlined in ICH guidelines (Accuracy, Precision, Linearity, Specificity, Limit of Detection and Limit of Quantitation).

2. Demonstrate an understanding of the importance and structure of essential regulatory documents related to method validation: Method Validation Protocol and Method Validation Report.

3. Demonstrate an understanding of the importance of stability indicating methods.

4. Evaluate the importance of peak purity analysis.

5. Assess the critical elements of the method validation required for cleaning validation.

6. Systematically approach method development and apply acquired knowledge.

Cheating and Plagiarism
Each student should be aware of the College's policy regarding Cheating and Plagiarism. Seneca's Academic Policy will be strictly enforced.

To support academic honesty at Seneca College, all work submitted by students may be reviewed for authenticity and originality, utilizing software tools and third party services. Please visit the Academic Honesty site on for further information regarding cheating and plagiarism policies and procedures.

All students and employees have the right to study and work in an environment that is free from discrimination and/or harassment. Language or activities that defeat this objective violate the College Policy on Discrimination/Harassment and shall not be tolerated. Information and assistance are available from the Student Conduct Office at

Accommodation for Students with Disabilities
The College will provide reasonable accommodation to students with disabilities in order to promote academic success. If you require accommodation, contact the Counselling and Disabilities Services Office at ext. 22900 to initiate the process for documenting, assessing and implementing your individual accommodation needs.


Topic Outline
Introduction, discussion on student/teacher objectives and expectations

Review of subject outline, grading structure, review of laboratory safety regulations, requirement to have laboratory coat, safety glasses and no open toe shoes at all times during laboratory assignments

Validation of an analytical method as a next logical step following method development in a regulated industry (for example pharmaceutical)
Validation concept

Regulations and guidance related to method validation

International Conference on Harmonization (IHC) expectations

Validation parameters and their relationship to the method itself

Appropriate considerations prior to method validation

Method validation protocols, objective, content, approval

Method validation reports, objective, content, approval, reference and discussion on any relevant deviations.


Precision (Repeatability, Intermediate Precision, Reproducibility)
Laboratory work

Mobile phase preparation, filtration technique, HPLC system familiarization

Chemstation software familiarization

Buffers, their use in Mobile phase preparations

Limit of detection

Limit of quantitation
Laboratory work

Linearity test (assay)

Specificity and Selectivity

Forced degradation studies
Quiz # 1 (Covers theoretical material presented during weeks 1-5)

Laboratory work:

Linearity test for impurities

Limit of Detection test

Limit of Quantitation test



System suitability Concept


Laboratory work: Accuracy test

Concept of Cleaning Validation and Cleaning Verification

Method Validation for Cleaning validation / verification

Laboratory work:

Precision (Reproducibility) test

Laboratory work:

Precision (Reproducibility) test

Quiz # 2 (Covers theoretical material presented during weeks 6-10)

Laboratory work:

Robustness test
Laboratory work:

Ruggedness test
Review of the material covered during previous sessions

Preparation for Final examination

Mode of Instruction
Labs and lectures

Prescribed Texts

Reference Material

Required Supplies
All students must wear a laboratory coat and safety glasses while in the lab.  This requirement is mandatory and consistent with safe practice.  Anyone not having the mandatory requirements will be asked to leave the lab.

Metric Ruler, Calculator, Graph Paper, Laboratory Coat, Safety Glasses

Promotion Policy

Grading Policy
A+ 90%  to  100%
A 80%  to  89%
B+ 75%  to  79%
B 70%  to  74%
C+ 65%  to  69%
C 60%  to  64%
D+ 55%  to  59%
D 50%  to  54%
F 0%    to  49% (Not a Pass)
EXC Excellent
SAT Satisfactory
UNSAT Unsatisfactory

For further information, see a copy of the Academic Policy, available online ( or at Seneca's Registrar's Offices.

Modes of Evaluation

  • Assignments are due at the beginning of the class on which they are due.
  • A late penalty of 10% per day is assessed for late assignments, including those not handed in at the beginning of class when due.
  • Material will not be accepted after one week following the due date and/or when the marked material is returned to students, whichever comes first.
  • Assignments are to be prepared by computer.

Absenteeism and Exams
  • Students should be aware that absenteeism almost guarantees an inability to achieve satisfactory grades.
  • Students who are absent for an examination due to an emergency (e.g., motor vehicle accident, hospitalization or death in the family) may provide official documentation within five days of the missed exam and be provided a deferred exam at a later date.  Official documentation includes a death notice or an original doctor’s certificate identifying the date, length of time expected absence and the specific reason for the absence.  Examinations missed without official documentation and approval result in a grade of zero.
  • There are no deferred options for missed tests.  The value of missed tests, at the discretion of the Faculty, will be added to other evaluation components

English Proficiency
  • All written work should demonstrate the following characteristics for clarity and conciseness:
-writing is consistent with the rules of English grammar
-spelling and punctuation are correct
-sentences are structured correctly
-main ideas are supported with specific, relevant examples and reasons
-work flows logically through supporting statements/paragraphs
-work is arranged in correct format (e.g., as a report, essay)
-up to 10% of the final grade may be deducted on all work if the above English competencies are not met.

Format for Assignments
  • Students must use the standard, APA style for quoting sources.   Help is available at:

Laboratory Attendance
The laboratory component is essential and therefore it is strongly recommended  that you attend all labs.  Any missed labs must be supported with a legal document with three days of the lab.  Any student who fails to attend 2 scheduled laboratory classes for a 7 week subject and more than 3 laboratory classes for a 14 week subject will not pass the subject.    

Laboratory Safety
Students are required to review and understand the safety procedures and guidelines outlined on the first class and sign the sheet to this effect before beginning work in the laboratory.  Students must also wear a lab coat and safety glasses when conducting experiments.
A student who arrives without the proper safety equipment will not be permitted to participant in the lab but will be asked to leave the class.  The student will receive no grade for the lab missed.

Grading is based on the following marking scheme:
Lab work 25%
Homework 25%
Quizzes 20%
Final Exam    30%

Other Evaluation Considerations
The student is expected to comply with the Safety Rules for working in the laboratory, sign the safety contract, and WILL NOT UNDER ANY CIRCUMSTANCES WEAR CONTACT LENSES in the laboratory.  The student will know where all safety equipment is located in the laboratory and will be familiar with WHMIS concepts and signage.


Approved by: Denis Gravelle