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Analytical Solutions To Analytical Problems"


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Vol. 3, No. 8. August, 2014
 In this issue...

Writing the Perfect Method
Part 8 - Analysis Procedures
HPLC Method Development Class
Writing the Perfect Method!

To view previous sections, view our
Newsletter Archive.

Part 8: Analysis Procedures
 
Your perfect method now must instruct the user on how to actually analyze samples.  Current practice in regulated laboratories generally requires a far more formal set of procedures.  In other environments, the procedures have historically been less demanding - e.g.., a standard followed by your samples.  However, with the availability of autosamplers and other automation, it is usually advisable to do more to improve the integrity of the data.

All methods should discuss the following activities, and provide some guidance on injection order.  The differences between regulated and non-regulated labs will usually be the amount of effort required for each step, and whether or not some of the activities can be combined in a single injection.

  • Instrument equilibration
  • Method verification/system suitability
  • Blanks
  • Calibration
  • Samples
  • Continuing/check standards
  • End of sequence verification/calibration
  • Instrument/column shutdown

Specific comments are listed below:
  • Instrument equilibration is important for all analytical instruments.  However, longer is not always better.  Provide general guidance for time (e.g., a minimum of 15 minutes).
  • Check the system with injection of "conditioning injections" before you begin the formal analysis.  Typically you want to inject one or more standards to verify that retention times and peak size are consistent.  It is very common for the first one or two injections to be "different" than the rest.  Avoid this problem.  (Note: this does not need to be a formal procedure as this information is never reported.  This is an informal activity as part of equilibration.)a
  • System suitability is an important part of validated methods, since this step verifies the performance of the instrument.  The amount of effort required will be related to the importance of the results, but at least some evaluation is required, even if it is only related to retention times.
  • Blanks and calibration injections demonstrate the response of the system, and both must be included.  Blanks are especially important for trace analysis methods.  There are some known situations where there is some background quantity of analyte or interference.  You must know how much is present.  Do specify exactly how many standards are are required and at what levels.  I personally believe that we spend too much time calibrating in modern methods, but once the method is finalized, it is too late to change the procedures.
  • Indicate whether samples are always analyzed singly or in duplicate or triplicate.  Specify when you run check standards and spikes, and how often, or under which conditions they are included.
  • You must always end the sequence of injections with a calibration verification.  (more on this next month when we discuss QC)
  • Instrument shut down and column storage are not critical for method performance, but they will improve both instrument and column lifetimes if they are performed correctly.
Do Not:
  • Specify an exact equilibration time (e.g., 15 minutes), as this is not technically accurate and can cause problems with compliance auditors.  Specify a reasonable range; do not burden the analyst with demanding requirements that do not help the quality of data.
  • Ignore the need to verify system performance.  This does not require extensive evaluation, but should be present in every method.
  • Assume that the blanks are negative.  You must verify, even if it takes only one injection.
  • Do not let the analyst decide on how many replicates to inject, or how often to run check standards.  Demanding procedures are not required, but something must be specified in the method.
  • Ignore the need to prepare both the instrument and column for storage/shutdown.  Typically you will want to remove any dissolved solids from the system (buffers, etc.) and clean the column before storage.  Failure to do this will cause early failure of both column and instrument.
Next Month: QC


Would you like us to evaluate your methods, or comment about this topic?  Contact us.
HPLC Method Development

October 6 - 10, 2014
ChromAfrica Laboratory
Nairobi, Kenya

Our next international event is a week long class in HPLC method development sponsored by ChromAfrica LLC and Chromacademy

This will be a highly interactive class featuring both conventional lectures and online training using the Chromacademy online content.  Students will also have an opportunity to work on case studies in method development.  All students will also receive a complimentary one year Premier Membership in Chromacademy, providing unlimited access to the largest chromatography training resource in the world!

If you would like more information on this event, please contact Mr. John Angira, or visit the ChromAfrica web site.

Seat are still available and we are also looking for additional sponsors.

We will have a summary in a future newsletter.
Quote for the Day

"Statistics is like taking cross sections of the trunk of a tree. It can tell you a lot about its history. You can figure out how healthy it is, how much volume the whole tree has, how much is root and how much is branch. But what it can't tell you is where the tree will branch, and which branches will become major, which minor, and which will rot and fall off and die."

– Orson Scott Card,
"The Originist," in Foundation's Friends,
M. H. Greenberg, Ed., Tor Books, New York, 1989

In This Issue

Writing the Perfect Method
Part 8 discusses your operating conditions.

HPLC Method Development

October 6 - 10, 2014, Nairobi, Kenya.

Quote for the Day
Tree lover.

Dowload Recent Presentations

Get your own copy of two recent presentations.  Both cover our recent work on HILIC columns.  These publications describe a simple way to evaluate multi-mode mechanisms on HILIC columns.

Pittcon, March, 2014:

Mixed Mode Mechanisms in LC: Curse or Cure?

Minnesota Chromatography Forum, May, 2014:

Vendor Seminars

The following courses are offered by Agilent Technologies at their Atlanta, GA training center.

Infinity HPLC Maintenance and Troubleshooting
October 28 - 29, 2014 (seats available)
December 16 - 17, 2014 (seats available)
Learn how to maintain your 1220, 1260, and 1290 instruments.

HPLC OpenLab ChemStation Operation
September 23 - 26, 2014 (full)
December 2 - 5, 2014 (seats available)
Learn all the features of this new software program in a hands-on setting.

 

Contact Us

Merlin K. L. Bicking, Ph. D.
ACCTA, Inc.
3534 Jessie Ct
Saint Paul, MN  USA  55125

Email: info@accta.com
Internet: www.accta.com

Phone:651-731-3670




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