Chromatography Forum

I'm doing method modification of Cefuroxime in Human Plasma. Just wanna make sure of its stability in stock solution. I dissolved the compound in Phosphate buffer 0.01 M, pH 6.5 because I've read from literature that it is stable from pH 4.5-7.3. Would this be practical or will i just use deionized water? I observed that any change in pH in the solution also affected it's RT, the reason why i used buffered diluent. though a lot of methods only used deionized water or MilliQ water as diluent. Would it prolong the stability of cefuroxime as well? Haven't conducted any stability test yet as of the moment. Any feedback would be appreciated. Thanks!

Ghie

I'm doing method modification of Cefuroxime in Human Plasma. Just wanna make sure of its stability in stock solution. I dissolved the compound in Phosphate buffer 0.01 M, pH 6.5 because I've read from literature that it is stable from pH 4.5-7.3. Would this be practical or will i just use deionized water? I observed that any change in pH in the solution also affected it's RT, the reason why i used buffered diluent. though a lot of methods only used deionized water or MilliQ water as diluent. Would it prolong the stability of cefuroxime as well? Haven't conducted any stability test yet as of the moment. Any feedback would be appreciated. Thanks!

Ghie

What is you mobile phase? You need to add some organic solvent to your diluent.

Diluent can be mobile phase, any organic solvent.

Using mobile phase (but of less solvent strength avoids peak splitting) as diluent provides good result in terms of validation of developed method. If your peak is not tailed, sharp and no ghost peak is appearing, I think you should not worry for the stability. Using deionised water may result in peak spliting since water is of higher solvent strength.


A change in pH of your buffer will affect RT. You need to see your pH meter is calibrated to provide reproducible results.

A simple test. Keep your solution of drug in buffer for a day (24 hours)and analyze. If no ghost peak appears it is stable. A variation is RT may occur don't worry with that they are usual LC trouble shooting problems.


Good Luck!

Regards,

Amary.

Thanks for your reply. I'm worried of the long term stability of the stock for a period of at least 28 days. Our practice here is to prepare 1 stock solution for the whole period of analysis of the plasma samples from a clinical study which usually takes us 2 to 3 weeks. We analyze the samples after each period using the same stock solution. That's why I am really inquiring of the stability of cefuroxime in stock solution and where can I dissolve it to prolong its stability. Have anyone read or studied Cefuroxime's stability in buffer, acetonitrile or MeOH? Again, thank you very much!

-ghie-

These reference related to stability of cefuroxime may be useful:

1. Cefuroxime hydrolysis kinetics and stability predictions in aqueous solution. J Pharm Sci. 1994 Apr;83(4):577-81.

Below is some text from this reference in PUBMED:

A. Acetate, phosphate, and borate buffers did not catalyze degradation.
B. Maximum stability was observed in the pH-independent region from pH 4 to 7, where the time during which cefuroxime concentration exceeded 90% of its initial concentration at 25 degrees C was 1.2 days.

According to above, the degrdation is specific acid-base catalysed rather than general acid-base catalyzed.

However, there is contradiction with anothere reference:
2. The relationship of diastereomer hydrolysis kinetics to shelf-life predictions for cefuroxime axetil. Pharm Res. 1991 Jul;8(7):893-8.

Buffer catalysis was observed in acetate and phosphate buffers.
No significant kinetic effect was observed for ionic strength in the range mu = 0.1-1.0.
..........exhibiting maximal stability in the pH range 3.5 to 5.5.

3. Stability of cefuroxime sodium in some aqueous buffered solutions and intravenous admixtures.
J Clin Hosp Pharm. 1986 Feb;11(1):47-54.

Both buffered and unbuffered solutions followed first-order decomposition. In 5% dextrose and 0.9% sodium chloride injections, cefuroxime was stable for 1 day (more than 90% potent) at 25 degrees C and for at least 30 days at 5 degrees C. At -10 degrees C, there was negligible decomposition after 30 days. The pH values of the solutions stored at 5 degrees C and -10 degrees C remained in the maximum stability range and the solutions were clear even after 30 days of storage.

1. Since ionic strength have no effect, deionized water does nt have advantage. Buffered solution is advocated to maintain the pH (in maximum stability range, based on two studies pH range can be narrowed to 4-5.5). Buffer of species reported to not catalyzing the degradation.

2. 28 days storing of stock solution may be possible at low temperature (stability need to be established in hand). I will not prefer doing this and would change the practice and make fresh solution everytime.

3. Chances of finding information on stability in Methanol and Acetonitriles are remote (at least at first place).

Thanks! t'was such a big amount of information. It will really be of help. I have a new problem though.

1. We run system suitability first before proceeding with any validation run and in the result, the RT of cefuroxime is repeatable with RSD of less than 2.
2. During the validation run, we noticed a change in RT for both cefuroxime and IS sulfamethoxazole.
3. RT changed from 6.2 to 6.6 minutes for Cefuroxime and 7.5 to 8.0 minutes for sulfamethoxazole.

M.P.: MeOH/ 0.005 M KH2PO4 buffer (pH 3.6), 30:70
Sample treament: Analytes are in plasma, precipitated with Acetonitrile and back-extracted with DCM/Hexane (1:1)
Stock and Working Solutions are dissolved in 0.01M KH2PO4 buffer, pH 6.5

Is this change in RT significant? Is there any part in the sample preparation or Mobile Phase that can influence the change in RT?

I will appreciate any feedback!

Thank you.
Ghie Malig

Since both IS and analyte time is shifted poptionately. May be lack of Equlibrium in column during system suitability test.