Part 7: Preparation of Samples
In an ideal situation, samples and standards are prepared in exactly the same way, using exactly the same procedures and solutions. In the real world, things may be different.
If your method uses a "process standards" method (usually because it is a derivatization procedure), then both samples and standards must be treated the same way. In most other methods, however, samples will require additional steps.
Oh, the lucky ones who have a "dilute and shoot" method. Dissolve in mobile phase, filter (don't forget that), and you are ready. Unless your sample has insoluble components that float up to the top in your volumetric flask. Where do you "fill to the mark," at the top of the 'floaties' or at the bottom where the bulk liquid starts? I have heard analysts defend either method. Without taking sides on this one, be consistent; always do it the same way.
"Sample preparation" to separate the analytes from other parts of the sample may be required, using such techniques as liquid-liquid, liquid-solid, or sold phase extraction. As with any other part of the method, you must be as precise as necessary in describing these procedures.
Other thoughts:
Do:
- Remember to list all reagents, accessories, and solutions in the appropriate section.
- Describe or list the masses and volumes needed for each sample. As most samples are usually solids, measuring out an adequate mass (approximate target amount) on an appropriate balance (specify number of decimal places) will be required.
- The method should prepare the sample for whatever final analytical technique is used. Ensure that your final dilution solvent is compatible with the instrument.
- Reversed phase methods will require a solution which is aqueous-organic, at the proper pH. Note the the organic part must match the LC method. Use the maximum concentration of aqueous solution that will dissolve the components.
- HILIC methods are the same but you should maximize the concentration of the organic phase.
- GC methods require a single solvent, which is similar in polarity to the column being used.
- Other analytical instrumentation will have their own unique requirements, but in every case, the solvent environment for both samples and standards should be as similar as possible.
Do Not:
- Assume other users will understand how to do a procedure, unless it is described in a document somewhere, or unless you require that they be trained in that technique in or to complete the procedure (update Scope as required).
- Specify that an exact mass be measured. This is nearly impossible to achieve with any practical accuracy or effort, so you should specify a range only (e.g., 100 +/- 2 mg).
- Assume that the user will or should know what you mean. There should be no confusion.
- Step back from your method and ask yourself if you could properly follow this procedure if you were seeing it for the first time.
- One test of this idea might be to make a list of every piece of equipment and glassware that you used (look in the sink before you begin cleaning", and make sure that each is mentioned in the procedure.
- (yes, I used this last month, but it was such a good suggestion that I decided to keep it for this month too!)
Next Month: Analysis Procedures
Would you like us to evaluate your methods, or comment about this topic? Contact us.
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Getting Help in the Lab!
Two recent lab visits highlighted a common situation in most laboratories today. Each visit involved training on basic and advanced techniques for LC and GC, and each visit was requested and configured by the laboratory manager. In each case, the lab manager wanted the training because "they didn't have enough time to help everyone learn these techniques. The staff needed to be able to work more independently."
Now, to be clear, these two labs are the exception, not the rule. The problem is that in most labs, the staff have little or no training, so whoever has to manage these labs will spend most of their time "fighting fires" or helping the lab staff solve all their problems. If the manager is not technical, then this job falls to whomever appears to know the most (whether they actually do or not).
A majority of labs that I visit suffer from a lack of internal resources at the "expert" level. Training the staff will help, but I cannot transfer all the knowledge gained in my 30+ years of experience to a new analyst in a day, or two, or three. Learning takes time, and happens more quickly when there is a "mentor" available to help with the process.
In the two labs that I mentioned earlier, those mentors were the lab managers themselves, and they realized, correctly and to their credit, that they could not help everyone all the time. Getting the staff some initial training to get them started along the learning road was the only way to get the lab to function efficiently. I wish more labs had leadership like this.
These are good models for organizing your lab:
- Assign someone as a "mentor" in the lab, and make sure that they have the time to actually do some mentoring,
- Get the less experienced staff some training so that the mentor can spend time solving the difficult problems, rather than wasting time on basic level training.
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Quote for the Day
"All good science is accumulative; no one
can get everything right the first time."
– Stephen Jay Gould,
Eight Little Piggies: Reflections in Natural History,
Norton, New York, 1993, p. 32
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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:
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Vendor Seminars
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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 (almost full)
December 2 - 5, 2014 (seats available)
Learn all the features of this new software program in a hands-on setting.
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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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