High Spikes in Internal Standard DHBA
Posted: Wed Mar 07, 2012 3:55 pm
I am having trouble with my internal standard reading about 3-4 times higher than the amount that was put in during the extraction procedure. I am adding 50 pg of DHBA to the sample and eluting with 250 ul of perchloric acid. I am injecting 20 ul of eluent into the HPLC, so the most internal standard that could possibly be on column is 4 pg of DHBA, yet I am getting 3-4 times this amount.
Also, it does not happen everytime. Sometimes I get a good DHBA response and other times it is a large spike in DHBA.
My thoughts are that:
1) There is some component in plasma that is binding to DHBA and causing the ECD to read it at higher than what it should be.
2) I need to switch the internal standard to something else.
Are there alternate internal standards that could be used for plasma catecholamine extractions that may not give erroneous results?
Thanks!
Also, it does not happen everytime. Sometimes I get a good DHBA response and other times it is a large spike in DHBA.
My thoughts are that:
1) There is some component in plasma that is binding to DHBA and causing the ECD to read it at higher than what it should be.
2) I need to switch the internal standard to something else.
Are there alternate internal standards that could be used for plasma catecholamine extractions that may not give erroneous results?
Thanks!