Page 70 - Application Handbook - Liquid Chromatography

LAAN-A-LC-E276

Application High Performance Liquid Chromatography

News Analysis of Aflatoxin B1, B2, G1 and G2 in Kakkonto

Using Nexera-i and RF-20AXS
No.L500

Aflatoxins are a type of mycotoxins that cause severe
and acute toxicity. They are also carcinogenic, and O O O
testing for aflatoxins is required for crude drugs O
produced from natural plants or preparations that O O O
contain crude drugs. "Analytical methods for aflatoxins O O
in crude drug and its product" was published in O O O O
Japanese Pharmacopoeial Forum as a proposed revision CH3 CH3
for the 17th Edition of the Japanese Pharmacopoeia (as Aflatoxin B1 Aflatoxin G1
of July 2015). This test method proposal proposes a TFA TFA
reference level of ≤10 µg/kg for total aflatoxins (sum of
aflatoxin B1, B2, G1 and G2). O O O O
This Application News introduces an example analysis O
of the complex crude drug Kakkonto based on the HO HO O O
proposed revision to the Japanese Pharmacopoeia 17th O O
Edition. The proposed revision explains a method of O O O O
analyzing aflatoxins using a fluorescent detector after CH3 CH3
derivatization with trifluoroacetic acid (TFA). This Aflatoxin B2a Aflatoxin G2a
Application News describes an example analysis
performed using this method, and another example
analysis performed without derivatization but with O O O O
direct fluorescence detection. O O O

O O
Aflatoxins in food are subject to regulation all over the O O
world, and in Japan, a regulation* and notification test O O
1)
2)
method* for aflatoxins have been published. See CH3 CH3
previous Application News L351, L422, L428, L430 and Aflatoxin B2 Aflatoxin G2
L435 for example analyses of aflatoxins in food
performed using these test methods.
Fig. 1 Structures of Aflatoxin B1, B2, G1 and G2 and Aflatoxin
Structures After TFA Derivatization (B2a and G2a)
n Analysis with Trifluoroacetic Acid Derivatization
When in a polar solvent, aflatoxins B1 and G1 are known
to have a lower fluorescence intensity compared to
aflatoxins B2 and G2. Possible methods of increasing the
fluorescence intensity are derivatization with a
photochemical reactor, TFA derivatization, and
electrochemical derivatization. This Application News
uses the TFA derivatization method that appears in the
proposed test method. The TFA derivatization reaction
is commonly known as the pre-column method, and
involves derivatization of the analytical sample before
HPLC analysis. The structures of each aflatoxin before
and after TFA derivatization are shown in Fig. 1.