Acetaminophen and codeine Phosphate Dissolution USP monograp

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Hi, Does anyone have any experience with the Codeine Phosphate and Acetaminophen monograph for dissolution?

I validated the dissolution procedure and cut the column down from a 250mm column to a 150mm column. At the time we got the required resolution (NLT 2) (Actual was something like 6). When we tried opening brand new columns, the peaks elute almost right on top of each other (Resolution 1.7)

Method Parameters:
Mobile phase: Monobasic potassium phosphate pH 2.35 containing 2mL of Triethylamine/L 92:8 Buffer Methanol.
Diluent/dissolution media: 0.01N HCl
Column: 250mm X 4.6mm, 5um L1 - USP column recommendation says Luna C18 - I tried running with a 150mm Luna C18(2) and with one column we got adequate resolution, and now with a brand new column we're having problems, both the USP and Phenomenex say that this is an appropriate column for Acetaminophen products.

Injection volume: 30uL
Column Temperature: ambient
Flow rate: 1.5mL/min
UV Detection: 214nm
I assume you have checked the obvious suspects:
- was the mobile phase prepared correctly? For Rs to drop by that much would require a big change in retention (k') or selectivity (alpha). Did the k' values of both peaks decrease significantly? If so, you may simply need to adjust the buffer/MeOH ratio (2% organic is not very much).
- were the new columns fully equilibrated?
- are the plate counts on the new columns within spec?

The next possible suspect is the column itself; did you develop / validate with a 150 mm column using a new column or did it have previous use (i.e., is there a chance the "old" column's properties had changed)?

Were all of the "new" columns from the same lot? With all due respect to Phenomenex, lot-to-lot variation can happen to any vendor. Ask them nicely if they can send you a column from a different lot to check.
-- Tom Jupille
LC Resources / Separation Science Associates
+ 1 (925) 297-5374
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