Liquid Chromatography Blog

Separation Science HPLC Solutions

Separation Science HPLC Solutions is produced in collaboration with John Dolan, best known as one of the world’s foremost HPLC troubleshooting authorities. He has been responsible for training of thousands of scientists over the last 30 years, and is also known for his ongoing research with Lloyd Snyder, resulting in more than 100 technical publications and three books.

Recent Posts

Acid in A and B

I had a follow-up enquiry about one of the recent articles. In 'Why Acid?', I discussed some of the reasons why acids such as formic or trifluoroacetic (TFA) are added to mobile phases for LC-MS work. The reader noted that in Figure 1 of that...

Injection Problems

Q: I am running an isocratic method with a 250 x 4.6 mm, 10 µm particle column at 1 mL/min with a 20% acetonitrile (ACN)/buffer mobile phase. I dissolve my sample in ACN and inject. Normally my compound of interest comes out at 7.7 min. I...

Frit Cleaning

In a recent email, a reader enquired about the best way to clean the frits used in the HPLC system. He indicated that he had been sonicating them in nitric acid, but wasn’t sure if that was the best way to approach the problem.

Retention Shifts

A reader complained about retention times that changed in an overnight HPLC run and wondered how to diagnose the problem source. The question left too many possibilities, and because there was no complaint about changing resolution, I’ll assume...

Enantiomer Separations

Recently, a reader sent a question about how to get started with a method that required separation of enantiomers by reversed-phase HPLC. Unfortunately, the problem is a non-starter in this context. Enantiomers cannot be separated by...

MP Lifetimes

A reader recently sent in this e-mail: “I am trying to find information that I can put in an SOP regarding how long a mobile phase can be used before it must be discarded. I’ve heard 1-2 weeks from some sources, whereas others say a month, and...

Why Acid?

Recently, I received an email from a reader asking why formic acid was specified as an additive for the mobile phase in an HPLC method he was using. Formic or trifluoroacetic acid at 0.1% concentrations are common, especially for LC-MS work....

PEEK Follow-up

PEEK (poly-ether-ether-ketone) tubing and fittings have become a standard part of most HPLC systems today. The flexibility and ease of cutting the tubing, coupled with the convenience of finger-tightened fittings make a combination that is hard...

Method Adjustment vs Change Part 7: Follow-up

In the previous articles in this series (Part 1Part 2Part 3Part 4Part 5, Part 6), we have explored the concept of the adjustment of HPLC methods. In particular, we have tried to determine the difference between method adjustment and...

Method Adjustment vs Change Part 6: Interactions

This is the last in a six-part series on allowed adjustments to HPLC methods. In the previous instalments (Part 1, Part 2, Part 3, Part 4, Part 5), we’ve looked at each of the specific variables listed in Table 1.

Method Adjustment vs Change Part 5: Volume, Column Temperature and Detector Wavelength

In the last few articles we've focused on the guidelines for method adjustment listed in the European Pharmacopoeia (EP) and United States Pharmacopoeia (USP). We looked at pH (Part 1), mobile-phase composition (Part 2), column dimensions (Part 3...

Method Adjustment vs Change Part 4: Flow Rate and Particle Size

In prior instalments of this series on method adjustment, we’ve looked at the guidelines from the European Pharmacopoeia (EP) and United States Pharmacopoeia (USP) for allowable adjustments in pH (Part 1), mobile phase (Part 2), and column...

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