Monday, 1 December 2014

iLearn, the e-Learning HPLC Course from MTS

iLearn, the e-Learning HPLC Course from MTS
Introduces the information required to understand the technique of HPLC and how it is applied to pharmaceuticals. Includes a discussion of: the different types of HPLC; types of stationary phases used to pack HPLC columns; the solvents and additives used in HPLC mobile phases; and the instrumentation used for HPLC analysis.

Suitable for:
This HPLC training course is aimed at beginners, but will also be useful for those with limited experience who wish to gain a better understanding of the technique.

This is an on demand open enrolment training course and is delivered online by e-Learning through e-MTS using a combination of videos, exercises and an assessment. The content is equivalent to 1 full day of training. All the training materials are available for 1 month and the learner decides when to watch the videos and complete the exercises and assessment.

iLearn, the e-Learning HPLC Course from MTS£125 + VAT per person. Includes: access to all the training materials through e-MTS; tutor feedback on exercises via email; a certificate of training on completion of the assessment; and a copy of the MTS training book, An Introduction to HPLC for Pharmaceutical Analysis.
More Information on this e-Learning HPLC Course (including booking form)

Visit the Course List page on our website for full details of all our available training courses.


Tuesday, 11 November 2014

Pharmaceutical Analysis Training Delivered at your site by Mourne Training Services

Pharmaceutical Analysis Training Delivered at your site by Mourne Training Services
On-site training courses are ideal when training is required for a group of learners. Significant savings may be made when compared to open enrolment training. Additionally, there are no expenses associated with travel or accommodation.Courses can be customised to include hands-on sessions using the laboratory equipment that the learners will operate after the training. All that is needed is a quiet room of a size suitable for the number of learners being trained, and access to any appropriate laboratory equipment.

The courses detailed on the MTS Course List are all available as on-site training. This includes:

Validation and Transfer of Methods for Pharmaceutical Analysis
How to Develop Stability Indicating HPLC Methods
How to Run HPLC Methods
How to Troubleshoot HPLC
How to Develop HPLC Methods - Part 1
How to Develop HPLC Methods - Part 2 

These may be customised and combined as required to provide a bespoke training programme which is both relevant and effective. We also offer a range of consultancy services related to training.

Attendees on all MTS courses are encouraged to bring along examples of problems, challenges etc. which may be relevant for discussion during the training. The MTS trainer is happy to enter into a confidentiality agreement should this be required. Consideration of real examples which have relevance to the learners will aid understanding and retention of information.

Contact us to discuss your in house training requirements and we will prepare a competitive quotation based on your specific needs.


Tuesday, 14 October 2014

Book: An Introduction to HPLC for Pharmaceutical Analysis

Book: An Introduction to HPLC for Pharmaceutical Analysis
Chemistry World review (May 2010):
 "A brief well-written guide and resource to assist the analyst in the use of HPLC in a pharmaceutical analysis environment."

This book is aimed at those who are new to HPLC. Whether you are a new starter in an analytical laboratory, an experienced scientist new to HPLC, a recent graduate, or a student, it provides an invaluable guide about how HPLC is actually used when analysing pharmaceuticals. Additionally, the book is full of practical advice on the operation of HPLC systems combined with the necessary theoretical knowledge to ensure understanding of the technique.

Key features include:
  • A thorough discussion of the stationary phase enabling the reader to make sense of the many parameters used to describe a HPLC column.
  • Practical advice and helpful hints for the preparation and use of mobile phase.
  • A complete overview of each of the different components which together make up a HPLC system.
  • A description of the contents of a typical HPLC analytical method and how to interpret these.
  • A step-by-step guide on how to follow a method and set up a HPLC analysis.
  • A discussion of system suitability criteria and how to interpret the values obtained during an analysis.
  • Explanation of the common methods of calibration and quantification used for pharmaceutical analysis
We understand the difficulty of buying a book online. You cannot just pick it up and leaf through the pages to help you decide if it is the right book for you. Therefore a preview made up of a selection of the pages in this book is provided to give a flavour of the content and style. You may also wish to visit the MTS training resources area where there are a selection of useful articles written by the author, Oona McPolin.

Cost: GBP £29.27
Author: Oona McPolin
ISBN: 978-0-9561528-0-0
Format: Paperback, 210 x 300mm (A4), 148 pages
Publisher: Mourne Training Services

Order a copy now on the MTS Website, click here.

Friday, 26 September 2014

Are Your Methods Fully Compliant with Regulatory Expectations?

  • Can you detect and quantify all potential degradation products during stability testing (as per ICH Q1)?
  • Is your new method validated correctly (as per ICH Q2(R1))?
  • Is your pharmacopoeia method verified correctly (as per EU GMP Chapter 6, FDA and USP <1226>)?
  • Is your validated method transferred correctly (as per EU GMP Chapter 6, FDA and USP <1224>)?
Find out how to develop, validate, verify and transfer analytical methods by attending one (or more) of the training courses from Mourne Training Services in Dublin, London & Berlin in 2014.

Choose from: (click on the course title for more information)
Validation of Analytical Methods for Pharmaceutical Analysis
1st & 2nd October 2014, Dublin
5th & 6th November 2014, London
3rd & 4th December 2014, Berlin
Transfer of Analytical Methods for Pharmaceutical Analysis
3rd October 2014, Dublin
5th December 2014, Berlin
How to Develop Stability Indicating HPLC Methods
13th & 14th November 2014, Dublin
8th & 9th December 2014, Berlin

The cost for the 2 day courses, method validation and developing stability indicating methods, is €1050 (+VAT) or £850 (+VAT), and the cost for the 1 day transfer course is €610 (+VAT) or £495 (+VAT). Full details of discounts for multiple bookings are available on the MTS website.


Thursday, 28 August 2014

MTS Recommends... Seven Common Faux Pas in Modern HPLC

MTS Recommends... Seven Common Faux Pas in Modern HPLC
Seven Common Faux Pas in Modern HPLC
By: Michael W. Dong
LCGC Europe
Volume 27, Issue 8, pp. 415–419
Aug 1, 2014

"The seven faux pas discussed here are common practices in high performance liquid chromatography (HPLC) that may no longer make sense because of improved technologies or other changes in modern HPLC. Therefore, careful consideration should be given to alternative approaches.
  1. Using columns packed with 5-µm particles
  2. Using 4.6-mm i.d. columns at 1 mL/min
  3. Filtering HPLC mobile phases
  4. Using buffered mobile phases
  5. Preparing fresh reference standard solutions with every assay
  6. Shaking HPLC sample vials
  7. Using stainless steel ferrules for column connections"

Wednesday, 20 August 2014

MTS Recommends... Data Integrity in the Analytical Laboratory

Pharmaceutical Technology, May 2, 2014

Data integrity in the analytical laboratory is an area of increasing focus for regulators such as FDA.

Monday, 11 August 2014

MTS Recommends... How Reversed-Phase Liquid Chromatography Works

How Reversed-Phase Liquid Chromatography Works

By: Mark R. Schure, Jake L. Rafferty, Ling Zhang, J. Ilja Siepmann
LCGC North America
Volume 31, Issue 8
Aug 1, 2013
How Reversed-Phase Liquid Chromatography Works
Described as follows by the authors:
"The keys to understanding reversed-phase liquid chromatography (LC) are provided at the molecular mechanism level as determined by high accuracy molecular simulation. The essential features of C 18 stationary-phase chains in contact with methanol–water and acetonitrile–water mixtures are discussed in the context of bonded-chain geometry, spatial distribution of alkane and alcohol solutes, retention mechanism, and retention thermodynamics. This tutorial is intended to be applicable to a wide audience ranging from occasional users of liquid chromatography to separation scientists."

MTS verdict: An interesting read for any HPLC method developer.


Monday, 4 August 2014

MTS Recommends... Complying with Revised Weighing Guidelines

Complying with Revised Weighing GuidelinesPharmaceutical Technology
Volume 37, Issue 8,

"USP s revised Chapters 41 and 1251 have new requirements for weighing, including balance calibration and testing."

This is an update to a previous article in Pharmaceutical Technology on the same topic (MTS Recommends... Complying with Revised USP Chapters on Weighing, Wednesday, 26 February 2014) and contains a useful extra section, "Common questions about the USP revisions".

Wednesday, 25 June 2014

MTS Recommends... Challenges in Analytical Method Development and Validation

BioPharm International,  

Experts give insight on method transfer, QbD, and regulations for analytical method development and validation for biopharmaceuticals.

Tuesday, 17 June 2014

MTS Recommends... Superficially Porous Particles: Perspectives, Practices, and Trends

MTS Recommends... Superficially Porous Particles: Perspectives, Practices, and Trends
Superficially Porous Particles: Perspectives, Practices, and Trends

By: Szabolcs Fekete, Davy Guillarme, Michael W. Dong
LCGC Europe
Volume 27, Issue 6
Jun 1, 2014

"Columns packed with superficially porous particles (SPPs) have created considerable excitement over the last few years. Indeed, this column technology manifests the advantages of fully porous material (loading capacity, retention) and some beneficial properties of nonporous particles (kinetic performance). This review provides an updated overview of the theory behind the success of SPP technology, trends, benefits, and limitations. It also summarizes the latest developments of sub-2-µm SPPs and instrumental constraints associated with their use. Finally, it describes several applications to illustrate the performance and the universal applicability of these newly engineered particles."

Wednesday, 11 June 2014

MTS Recommends... Is a Sample Size of n=6 a 'Magic' Number?

Pharmaceutical Technology
Volume 38, Issue 6

"Statistical analysis shows how much testing is needed to deliver a reliable estimate result."

Friday, 25 April 2014

EU GMP Chapter 6 Updated - Effective from 1st October 2014

The revision of EU GMP Chapter 6 on Quality Control has now been finalised and will be effective from 1st October 2014. The reasons for change on the new document are summarised as: "Inclusion of a new section on Technical transfer of testing methods and other items such as out of specification results."

The finalised document is mostly similar (a few of the proposed changes have not made it in) to the proposed draft that I reviewed in two blog posts last year.

The new section on method transfer is summarised in the first blog:
How does the Update to Chapter 6 of the EU GMP Guidelines Affect Your Lab?

The other updates in Chapter 6 are covered in this follow-up blog post:
How Does the Update to Chapter 6 of the EU GMP Guidelines Affect You? - Part 2

Ensure that you are up to date with the method transfer expectations of EMA, FDA and USP by attending the MTS training course, 'Transfer of Analytical Methods for Pharmaceutical Analysis'. Next opportunity is on the 2nd October 2015 in London. Check out our Course List for details of other dates and locations, and other courses, or contact us for more information.


Thursday, 17 April 2014

The Nature of 'Error' in Analytical Testing

It is important for anyone working in an analytical laboratory to have a through understanding of potential sources of error, and how the implementation of an effective laboratory quality system works to both minimise the effects of inherent sources of error and eliminate human error.

The meaning of 'Error' is very particular when applied to analytical testing. It is defined as the difference between an individual value and the true result and is regarded as having two components, namely, a random component and a systematic component. Random sources of error cause replicate measurements to fluctuate randomly around the mean, e.g., variability of HPLC injections. Systematic sources of error cause the result to be higher or lower than the true value, e.g., the assigned purity of a reference standard. Both types are inherent in the analytical method and are controlled by the elements in a typical quality system, such as calibration of instruments and equipment, and management of reference standards. They may also be introduced by the analyst, as human error, therefore suitable training in both correct procedures and the quality system is essential.

Mourne Training Services have now introduced a new training course to address this topic. It is day 2 of our 3 day course, 'Laboratory Data Integrity', and may be attended as a single day or as part of the full course.  

Brief course description:
Applying Data Integrity in the Laboratory; Minimising Analytical Error
This course deals with the effects of analytical errors on laboratory data integrity. This involves building an understanding of the nature and sources of analytical errors so that their effects can be minimised during testing, leading to high standards of data integrity and reduced numbers of OOS/OOE results due to laboratory errors.

It is aimed at everyone working in an analytical laboratory, being both a perfect introduction for new starters and a useful refresher for more experienced analysts. Full details are available on the MTS website and if you would like us to deliver this course in your laboratory just Contact us for more information.


Tuesday, 15 April 2014

Help on: Do I need to revalidate my method if I change the HPLC column?

MTS Helpdesk - Send us your questions about pharmaceutical analysis
Do you have any problems relating to analytical chemistry for pharmaceuticals or training? Send your questions to the MTS helpdesk using our contact form.

"I would like to have a 'back-up' column for my HPLC method so that I am not reliant on one supplier, but how can I choose one that will be suitable and do I need to validate the method fully on the replacement column?"

"My preference is to develop a new HPLC method on a popular column from a reliable supplier. This gives me confidence that the manufacturing process is well established and the batch to batch variability of the columns is likely to be low. Although a back-up column may be reassuring, it is not really necessary at this stage. I normally write the phrase 'or equivalent' after the details of the column in the method, which will allow investigation of an equivalent column in the future should it be necessary.

In this scenario it is important that if a back-up column is selected at a later date, it does actually produce the same chromatography, and is therefore equivalent. I would suggest that the USP database ( would be a good resource to help choose a column which is likely to be equivalent. Scroll down to the 'PQRI database' and see if your current column is in the database. If so, you will be able to identify some potential candidates.

If you find that the retention times, and more importantly band spacing (or relative retention times) are much the same on the new column when compared to the original column, then an equivalency study should be sufficient to show that the columns are equivalent. This may be a simple repeatability study on each column combined with the system suitability requirements being met. The precedent for not embarking on a full validation study for the new column is that this is the approach used by the pharmacopeia.

If a back-up column is chosen which does not give the same chromatography as the original column then it is more likely that a full validation on the alternative column will be required since the methods are not equivalent. This is probably best judged on a case by case basis." you would like more advice on problems relating to HPLC or method validation, then why not come along to an MTS training course? Visit our Course List to browse our upcoming dates and locations. Or contact us for more information.


Friday, 4 April 2014

Training Courses on Method Validation and Transfer in London - May 2014

Training Courses on Method Validation and Transfer in London - May 2014
The next event in the MTS open enrolment training course schedule for 2014 will be held in London on the 28th to 30th May.

The courses on offer are:
Validation of Analytical Methods for Pharmaceutical Analysis, 28th & 29th May - early booking rate is £850 + VAT
Transfer of Analytical Methods for Pharmaceutical Analysis, 30th May - early booking rate is £495 + VAT

Check out the Course List page on the MTS website for more information on these training courses, including detailed course descriptions. More details about costs and available discounts for multiple course bookings and groups can be found on the booking forms (also accessed from the course list page).


Friday, 28 March 2014

MTS Recommends... Current Practices and Considerations for a Stability-Indicating Method in Pharmaceutical Analysis

Current Practices and Considerations for a Stability-Indicating Method in Pharmaceutical Analysis 

LCGC Europe

This article provides an overview of the requirements of a stability indicating method and discusses the process of SI method development and validation under three main steps: 
  1. Obtaining a suitable sample, 
  2. developing the method, including the separation technique and detector, and
  3. the final validation of the method. 
It includes a very useful literature review and thus the list of references should provide additional information on the topic.

If you would like more information on a practical approach to developing stability indicating methods then you may be interested in the MTS training course, 'How to Develop Stability Indicating HPLC Methods'. Upcoming dates include: 2nd & 3rd June in London and 8th & 9th December in Berlin.

Wednesday, 19 March 2014

New Method Validation Guidance from FDA

FDA has released a new guidance document for analytical method validation, "Guidance for Industry: Analytical Procedures and Methods Validation for Drugs and Biologics." This revised draft supersedes the previous guidance for industry "Analytical Procedures and Methods Validation", which has been in draft version from August 2000. When finalised, it will also replace the 1987 guidance for industry on "Submitting Samples and Analytical Data for Methods Validation". stated aim of this guidance is to provide "recommendations on how you, the applicant, can submit analytical procedures and methods validation data to support the documentations of the identity, strength, quality, purity, and potency of drug substances and drug products. It will help you to assemble information and present data to support your analytical methodologies."

To assist you in an assessment of the new guidance I have summarised the content under the following headings: What's new, what's stayed (nearly) the same, and what's gone?

What's new?
Section III on 'Analytical Methods Development' consists of three paragraphs focused on the importance of getting the method right at the beginning. To quote: "the choice of analytical instrumentation and methodology should be selected based on the intended purpose and scope of the analytical method." It goes on to list the validation characteristics which may be evaluated during method development: "specificity, linearity, limits of detection (LOD) and quantitation limits (LOQ), range, accuracy, and precision." The next two paragraphs are about method robustness and how this "should be evaluated because this characteristic can help you decide which method you will submit for approval."

I think that this emphasis on methods development and robustness of methods is a great idea. It seems so obvious that you might think it's not necessary in regulatory guidance. However, I am constantly surprised during my work, whether it is in training, auditing or consulting, by how little attention is paid to method robustness until it is too late, resulting in problematic method validation and transfer. No doubt FDA comes across this regularly. However, in the third paragraph it says that "To fully understand the effect of changes in method parameters on an analytical procedure, you should adopt a systematic approach for method robustness study (e.g., a design of experiments with method parameters)." I've no problem with that, it makes sense, but it goes on to say: "You should begin with an initial risk assessment and follow with multivariate experiments." In my opinion, multivariate experiments may be very useful for some methodologies, but are not always the best approach. 

Under section VI, Part C on 'Compendial Analytical Procedures', the section on method verification has been updated to make it clear that the verification should be generated under a verification protocol and the results should be included in the submission. Information required in the protocol is detailed but the actual choice of which validation characteristics to investigate is left to the applicant, it is implied (rather than stated) that the approach taken should be justified.

The new Section VIII on 'Life Cycle Management of Analytical Procedures' begins with four paragraphs on the concept of method life cycle management and how it relates to product life cycle. It says that "Trend analysis on method performance should be performed at regular intervals to evaluate the need to optimize the analytical procedure or to revalidate all or a part of the analytical procedure. If an analytical procedure can only meet the established system suitability requirements with repeated adjustments to the operating conditions stated in the analytical procedure, the analytical procedure should be reevaluated, revalidated, or amended, as appropriate." I like these sentences very much since it is advice that I regularly give to customers when they ask me that rather unscientific question - "How often should I revalidate my methods?"

Further on in this section, under Part B, 'Analytical Method Comparability Studies', the section from the previous draft guidance on 'Alternative Analytical Procedures' has been updated to include additional information on biological products and also more detail on the requirements for the comparability study, in particular with respect to stability indicating methods. There is a new section on 'Analytical Methods Transfer Studies' which consists of a brief paragraph containing a lot of information. A comparative approach is suggested where: "The comparative studies are performed to evaluate accuracy and precision, especially with regard to assessment of interlaboratory variability." The importance of using suitable materials for the transfer of stability indicating methods is stressed.
Finally, in part C of this section, 'Reporting Postmarketing Changes to an Approved NDA, ANDA or BLA' is discussed.  

You may also notice that the language used in the revision is different to the previous draft in that it addresses 'you' throughout, rather than 'the applicant'.

What's stayed (nearly) the same?
The 'Introduction' and 'Background' sections are similar to the previous version although there are new references to some of the content discussed above.

The section IV on 'Content of Analytical Procedures' is also quite similar although it has been rewritten and in my opinion is much improved with greater clarity. Of particular note:
  • The number of replicates in sample preparation for quantitative tests is mentioned. I think this is important since my opinion is that the required number of replicates should be considered for each individual method, based on the data obtained during validation.
  • Information on the stability of solutions and storage conditions is advised.
  • ASTM E29 is referenced with respect to use of significant figures.
  • The information provided is very focused on HPLC methods.
The section VII, 'Statistical Analysis and Models' is similar to the 'Statistical Analysis' section in the previous version but has been rewritten and includes reference to use of chemometric and multivariate models to reflect the increasing use of these methodologies in techiques such as NIR.

What's gone?
The new draft document is 14 pages long and the previous draft is 33 pages long so it follows that quite a lot of content has been removed. The sections in question are (all referenced by the section number in the year 2000 draft):
  • III. 'Types of Analytical Procedures'. 
  • IV. 'Reference Standards' - sections on characterisation have been removed.
  • VII. 'Methods Validation for NDAs, ANDAs, BLAs, and PLAs - Validation characteristics are listed in the new version but the details on the required methods validation information has been removed. Also the table defining recommended validation characteristics is gone.
  • X. 'Methods Validation Package: Contents and Processing' - replaced by a short section which directs that necessary information will be provided by FDA if required.
  • XI. 'Methodology' - details on particular methodologies such as HPLC, GC, etc. has been removed.
  • Attachments
Find out more about how to validate, verify and transfer analytical methods on an MTS training course. Check out the Course List for details of dates and locations, or contact us for more information.


Thursday, 6 March 2014

MTS Recommends... Life Cycle and Quality by Design for Chromatographic Methods

LCGC Europe,
Volume 27, Issue 2,

"A recent stimulus to the review process article by the United States Pharmacopoeia (USP) expert committee proposes a major change to the way regulated laboratories develop, validate, and control analytical procedures. Is this Quality by Design (QbD) for the chromatography laboratory?"

Wednesday, 26 February 2014

MTS Recommends... Complying with Revised USP Chapters on Weighing

An overview of the USP's revised Chapters <41> and <1251>, which became official in December 2013, and have new requirements for weighing, including balance calibration and testing.

Friday, 7 February 2014

MTS Recommends... Drug-Excipient Interactions

Drug-Excipient Interactions

by Patrick Crowley and Dr Luigi G Martin, in Pharmaceutical Technology Europe, March 2001

I think that this article from back in 2001 will be a useful resource if you are performing forced degradation studies on drug products. It provides an excellent overview of potential drug-excipient interactions to consider.

Thursday, 6 February 2014

Help on: For how long should I equilibrate my HPLC column?

Do you have any problems relating to analytical chemistry for pharmaceuticals or training? Send your questions to the MTS helpdesk using our contact form.

"While operating an HPLC system my manufacturer told me to equilibrate my system for approx. 10 minutes. Though I came to know from available literature from various sources that equilibration time depends on column dimensions (which is coming out to be quite large). So what is the purpose of equilibration and how do we determine the time of equilibration? And also do I need to equilibrate every time I run my system?"

"Equilibration of an HPLC system is necessary to achieve reproducible chromatography and therefore quantitative results. It would seem that equilibration results in a stable and even distribution of the mobile phase within the stationary phase in the column and thus it is required every time a system is set up for analysis. It has been found that for best results, when an HPLC system is first set up, a minimum of 20 column volumes is required. We measure it in column volumes because the time taken will vary depending on the dimensions of the column (as you have discovered). There is a formula that you can use to determine how long a time will be required for 20 column volumes and I refer you to a blog that I have written on this subject: Equilibration of HPLC columns

I also provide a calculator that will work it out for you on my website which you may find useful: HPLC Calculator

Typical symptoms of incomplete equilibration are drifting baselines and retention times. You will also need to equilibrate your system between each injection if you use a gradient method, 10 column volumes is usually recommended."

If you would like more advice on equilibration and other HPLC-related issues, then why not come along to our HPLC courses on the 10th to 13th June 2014 in Dublin? You can choose from: 
  • How to Run HPLC Methods (10th June)
  • How to Troubleshoot HPLC (11th June)
  • How to Develop HPLC Methods - Part 1 (12th June)
  • How to Develop HPLC Methods - Part 2 (13th June)
Visit the MTS Course List for more information including course descriptions and course fees.


Tuesday, 4 February 2014

Free Aide Mémoire for Analytical Method Transfer is Updated!

The aide mémoire provided by MTS in the course, Transfer of Analytical Methods for Pharmaceutical Analysis, which assists in the review of analytical methods prior to transfer has been updated to include more considerations for transfer of HPLC methods.

The aide mémoire is a document in progress, if you have any ideas about what you think should be included then please send your suggestions to me using our Contact Us page.

Click on the image to the right, or click here, to access the download page for the free method transfer tool.


Thursday, 30 January 2014

HPLC Calculator is Updated!

The free HPLC Calculator from MTS has been updated following a suggestion from a user. The flow rate has been moved to the gradient table to provide more flexibility.

Please send us any suggestions for improvement or additional calculations that would be helpful.


Friday, 10 January 2014

20% Discount on All Bookings by LinkedIn Followers!

20% Discount on Training Course Bookings until 14th Febnruary 2014
MTS have a very special offer for you: Any bookings made on or before 14th February 2014 (St. Valentine's day) will be eligible for a 20% discount, provided you are a LinkedIn follower of MTS.

The offer is in addition to existing offers such as early booking discount, group bookings, and multiple course bookings.

All you have to do is click below to follow MTS in LinkedIn.

(Note: you will need to have set up an account in LinkedIn to become a follower.)

Then choose which course or courses you would like to attend. Visit the MTS website for a full list of available courses and dates, or click here for the 2014 calendar. Topics include: Analytical method validation and transfer; stability indicating method development; and basic, intermediate and advanced HPLC.

The offer also applies to our training books:
Validation of Analytical Methods for Pharmaceutical Analysis, and
Introduction to HPLC for Pharmaceutical Analysis.


Thursday, 9 January 2014

MTS Recommends... Calibration of instruments: Is Your UV Spectrometer Accurate Enough

Calibration of instruments: Is Your UV Spectrometer Accurate Enough
Pharmaceutical Technology
This article takes a statistical look at the calibration requirements for a UV spectrometer.

Wednesday, 8 January 2014

Replicates in Analytical Methods - When and How Many?

Do you prepare your samples/standards in duplicate without really thinking about it? If challenged on why, would your answer be something like, "it's good practice, isn't it?" The use of replicates in analytical procedures is commonplace but it is important to understand why you are doing it and also what the results are telling you.

There are different types of replicates in analytical methods, which are used for different purposes. Most commonly it comes down to two types, either preparation replicates or measurement replicates. In the case of preparation, these are samples or standards that are prepared from the beginning to end of the procedure in the same way but separately (often referred to as replicate weighings). In the case of measurement replicates, a single sample/standard is measured more than once (an example would be multiple injections in the case of HPLC).

Standard Preparation
Replicates in calibration standard preparation are often used to provide confidence in the initial weighing of the standard material. The idea is that if the response is corrected using the actual amount which was weighed out, then the response factors will be the same if the weighing has been performed correctly.

This may apply in the case of single point calibrations, where a single concentration of standard is used for the calibration, or in multi-level calibration where the original weighing is diluted to prepare a series of standards over a range of concentrations.

The difficult part is setting an acceptance criteria for the required agreement between these replicate standards. Duplicate preparation is common since it requires the least amount of extra work but two results cannot be compared using common statistical tools, such standard deviation. Usually the agreement is expressed as a percentage of one result compared to the other. How close this should be to 100% will depend on the inherent error in the method and the considerations are similar to those in setting acceptance criteria during method validation.

Sample Preparation
When replicates are used in sample preparation it is more likely that the reason is related to variability of the method caused by the inherent errors in the sample preparation steps. When the sample is very simple, such as a pure substance then the replication may be used a weighing check as discussed for standards previously. For samples which are composed of mixtures of compounds it would be expected that the sample preparation will involve more sources of error than for the standard preparation.

When the variability is evaluated in method validation then number of replicates prepared is usually 6. This number is high enough to express the variability of the method using statistics such as standard deviation and thus confidence intervals. (In out of specification testing, where retesting is required to determine the actual value of a suspect sample with a high level of confidence, the number of replicates is usually somewhere between 5 and 8 for the same reason.)

However, in routine analysis, 6 replicates per analysis would be unacceptable in terms of the time taken for a single analysis. A decision has to be made regarding how many replicates will be enough for an acceptable result. Usually the best source of information for this decision is the precision data in the validation report.

Standard Measurement
For many analysts in the pharmaceutical analysis laboratory, the most familiar use of replicates in standard measurement is the 6 injections performed as part of system suitability testing for HPLC methods. The purpose of this test is to show that the variability of the peak area for the same standard injected multiple times does not vary significantly, and thus the combination of the amount injected and the integration applied is the same for every injection. This is critical when using external standard calibration where separate solutions of standard and sample are being compared.

When it has been shown that the variability is acceptable then a decision still has to be made about which standard measurements will be used to construct the calibration curve. This may vary quite significantly with some analysts preferring to use all injections of standard during the analysis (in the case of a chromatographic run where there are regular injections of standard in between injections of samples) and others using injections of standard prior to or bracketing the samples being analysed. Whichever approach is used should be justified and also applied during the method validation studies so that the actual method is being evaluated.

Sample Measurement
Measuring a sample more than once implies that there is a level of variability in the measurement which needs to be accounted for to produce a reasonable estimate of the true value for the sample. If the method validation investigation for a method has shown that the variability in a method is related to the sample preparation and very little to the measurement then there is little benefit in including replicate measurement in the routine use of the method.

It is tempting to use replicate measurements instead of preparations as a short-cut since there is less work involved but if this approach is taken it should be because the aim is to account for measurement variability and not sample preparation variability.

Occasionally I see acceptance criteria for HPLC methods based on 4 individual results which comprise of duplicate sample preparation combined with duplicate injections. The problem with this approach is that when it fails you don't know whether it is because of the injector/HPLC system or the sample preparation. The remedial action will be quite different for each since in one it would be a problem with the HPLC system which needs to be corrected, and in the other it would be a problem with the weighings, dilutions, etc. involved in preparing the sample test solutions.

In summary, replication is used to increase confidence in the integrity of data generated by analytical methods. The use of replication needs to be considered carefully and the acceptance criteria relating to the agreement of the replicates should be defined clearly in the method.

Although there is no explicit requirement for doing so, I would recommend that the decision regarding replication for a particular method is discussed and justified in its associated validation report. This seems to me to be the most appropriate document since it contains the evidence which demonstrates that the analytical method is suitable for its intended use.


Tuesday, 7 January 2014

Training Courses on Method Validation and Transfer in Dublin - March 2014

The first events in the MTS open enrolment training course schedule for 2014 will be held in Dublin on the 26th to 28th March.

The courses on offer are:
Validation of Analytical Methods for Pharmaceutical Analysis, 26th & 27th March - early booking rate is €1050 + VAT
Transfer of Analytical Methods for Pharmaceutical Analysis, 28th March - early booking rate is €610 + VAT

Check out the Course List page on the MTS website for more information on these training courses, including detailed course descriptions. More details about costs and available discounts for multiple course bookings and groups can be found on the booking forms (also accessed from the course list page).


Thursday, 2 January 2014

Help on: How long should it take to purge the lines on my HPLC system?

Do you have any problems relating to analytical chemistry for pharmaceuticals or training? Send your questions to the MTS helpdesk using our contact form.

"When I am setting up the HPLC systems in the laboratory where I work, the relevant procedure directs that the new mobile phase is flushed through to the purge valve on the pump for 5 minutes at a flow rate of 5mL/min. Is this enough? How do you work out the most suitable purge for an HPLC system?"

"When setting up an HPLC system, the aim of the purge is simply to flush through all the lines so that any remaining solvent in them from a previous analysis or wash is replaced with the new mobile phase. As you would expect, the amount of new solvent required depends on the volume of the tubing in the system. The major contribution to the volume of tubing is usually the on-line vacuum degasser, if installed. The design of these modules is such that they require a relatively large volume to perform well. For example, a typical Agilent 1200 degasser has a volume of ~12 mL for each line. Agilent recommends a purge of at least 30mL in total for the 1200 systems.

Therefore, to estimate how much volume is required for a purge, check the volume specification of the inlet tubing, the degasser and the pump that you are using, then double it to give an approximate purge volume. You may find that the manufacturer of your particular instrument has a recommended amount to use. A flow rate of 5mL/min is commonly used for standard HPLC systems.

If you have a mixture of different brands of HPLC instruments (and degassers) within one laboratory and want the procedure to cover them all then find out which requires the longest purge and use that as the default."