Chromatography Forum

Hi All

Has anyone experienced any problems with the method for Monodechlorovancomycin in Vancomycin HCl. When we start the analysis we have a symmetrical peak with a tailing factor of 1.0. After about 50 injections the peaks start to front and the tailing factor is about 0.8. We have tried a Waters Xterra RP 18 (one recommended by the USP), as well as a BDS Hypersil. Both columns showed the same phenomenon with the peak starting to front after about 50 injections.
USP conditions:
Mobile phase: Dissolve 2.2 g of 1-heptanesulfonic acid in about 500 mL of water, add 125 mL of acetonitrile and 10 mL of acetic acid, and dilute with water to 1 L.
Mode: LC
Detector: UV 280 nm
Column: 4.6 mm × 25 cm; packing L1
Column temperature: 60
Autosampler temperature: 5
Flow rate: 1.5 mL/min
Injection volume: 50 µL

Any advice or recommendation would be greatly appreciated.

Mike

Hi Mike,

We're now working with this method, too, six years' later. I've tried the Zorbax Eclipse XDB C18 as well. Not too happy with the results obtained with this column from jump. Advice from Waters is to try out the XBridge equivalent to the Xterra RP 18 (so a polar-embedded phase). Maybe it's not such a surprise that Eclipse and Hypersil Gold behave differently than a polar-embedded phase to begin with?

OK then. I think we're going back to the future to try an Xterra RP 18, don't know if the boss is getting the 3.5 um or the 5 um particle size, 4.6 x 250 mm, I'll see what happens with that in my hands. Like you (back then ), we're trying to verify the USP method for limit of monodechlorovancomycin.

Cheers!

Hi Matt

We eventually settled on an Xterra RP 18, 250mm x 4.6mm, 5um. Column stability is not great, however, we are not testing many batches /year. The monodechlorovancomycin is a process related impurity, therefore, we only perform this test on the API.

Regards

Hi Michael,

Thank you very much for your guidance--I greatly appreciate your help! Was kind of afraid that this would be the case. I'm also looking at NovaPak C18 (USP method again) with a 50:50 (v/v) 18 mM or so sodium phosphate buffer @ pH 8.0 mixed with ACN for Ropivacaine HCl. So far method is "stable", don't know how long that will last w.r.t. the stationary phase. That said, bupivacaine and 2,6-dimethylaniline are both resolved in injections of the API.

Hopefully We'll be able to bear out your experience with monodechlorovancomycin...gee whiz, there are two "monodechlorovancomycins" anyway, aren't there, as well as a di-dechlorospecies. What a mess!