Vancomycin from tissues
Posted: Fri Dec 18, 2020 11:50 am
Dears
Does anyone have experience with Vancomycin (glycopeptide antibiotic) extraction from animal tissue?
The molecule is large (1450 Da) and highly polar. LC/MS/MS analytics is no problem with LOQ < 1 µg/L in water.
I tried extraction from tissues (liver, kidney and subcutaneous tissue) with the following methods:
1.) QuPPe-AO method => Recovery 0 %
2.) QuEChERs (buffered) => Recovery 0 %
3.) Homogenisation with excess water, centrifugation, filtration => Recovery < 10 %
3.) Homogenisation with lower amounts of methanol/water => Recovery < 10 %
4.) Homogenisation with metaphosphoric acid/methanol => Recovery < 5 %
5.) Homogenisation with water, cooking in microwave => Recovery 50 % (one-time lucky shot, sadly)
6.) Homogenisation with 0.3% metaphosphoric acid, cooking in microwave => Recovery < 20 %
6.) Homogenisation with less water, repeated cooking / resuspension of pellet in water bath at 80 °C => Recovery 20 - 30 %
7.) Addition of NaCl to 6. => Recovery 20 - 30 %
I tested for matrix effects => not significant, spiked blank extracts are OK.
I tested cooking stability => stable during mild cooking but not in the microwave
I have also tried to remove fat and debris with LLE (cyclohexane and dichloromethane) and dSPE (C18, PSA, diatomaceous earth) => no improvement
I hoped for literature because Vancomycin is an old molecule, but found only methods for plasma, urine and water.
Any further suggestions?
Thanks a lot
Jörg
PS.: I know from literature that Vancomycin is not metablised in liver, so I don't think that we see enzymatic degradation or similar biological effects.
Does anyone have experience with Vancomycin (glycopeptide antibiotic) extraction from animal tissue?
The molecule is large (1450 Da) and highly polar. LC/MS/MS analytics is no problem with LOQ < 1 µg/L in water.
I tried extraction from tissues (liver, kidney and subcutaneous tissue) with the following methods:
1.) QuPPe-AO method => Recovery 0 %
2.) QuEChERs (buffered) => Recovery 0 %
3.) Homogenisation with excess water, centrifugation, filtration => Recovery < 10 %
3.) Homogenisation with lower amounts of methanol/water => Recovery < 10 %
4.) Homogenisation with metaphosphoric acid/methanol => Recovery < 5 %
5.) Homogenisation with water, cooking in microwave => Recovery 50 % (one-time lucky shot, sadly)
6.) Homogenisation with 0.3% metaphosphoric acid, cooking in microwave => Recovery < 20 %
6.) Homogenisation with less water, repeated cooking / resuspension of pellet in water bath at 80 °C => Recovery 20 - 30 %
7.) Addition of NaCl to 6. => Recovery 20 - 30 %
I tested for matrix effects => not significant, spiked blank extracts are OK.
I tested cooking stability => stable during mild cooking but not in the microwave
I have also tried to remove fat and debris with LLE (cyclohexane and dichloromethane) and dSPE (C18, PSA, diatomaceous earth) => no improvement
I hoped for literature because Vancomycin is an old molecule, but found only methods for plasma, urine and water.
Any further suggestions?
Thanks a lot
Jörg
PS.: I know from literature that Vancomycin is not metablised in liver, so I don't think that we see enzymatic degradation or similar biological effects.