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Chinese traditional medicine has been used in China for thousands of years for
their low toxicity and less side effect.
Nowadays, the constituent study of plant
medicine becomes more and more important for the bioactive ingredients in them may
provide model compounds for synthesizing some pharmaceuticals by chemical method.
Capillary electrophoresis (CE) has become a powerful technique for determining some
constituents in plant drugs.
Chinese traditional medicine, Scutellariae Radix, is the root of Scutellaria
baicalensis Georgi and can be use to clean away heat, purge fire and detoxify toxicosis.
Astragali Radix is the root of Astragalus membranaceus (Fisch.) Bge. Var. mongholicus
(Bge.) Hsiao or Astragalus membranaceus (Fisch.) Bge. It is mainly used to promote
vital energy. However, the two crude drugs are usually confused for their shape, color,
and the Chinese names are similar. For example, the Chinese names of Scutellariae
Radix and Astragali Radix are “Huang-Qin” and “Huang-Qi”, respectively. It would
bring about side effects if they were misused for their different therapeutical functions,
so it is necessary to establish some simple and accurate methods for their
differentiation.
For many bioactive constituents such as flavonoids in plant drugs are
electroactive, capillary electrophoresis with electrochemical detection (CE-ED) should
become an alternative technique in the study of Chinese traditional medicines. A novel
method for the differentiation of Scutellariae Radix from Astragali Radix based on
CE-ED has been established in this paper, providing a reliable technique for
pharmaceutical analysts. For Scutellariae Radix contains baicalin and baicalein that
can not be found in Astragali Radix, this differentiation method for both the crude drugs
was carried out by determining the baicalin and baicalein in them by CE-ED. The
electrochemical detection used in this paper provided a high degree of selectivity and
sensitivity to electroactive species. To our knowledge, no report on the application of
CE on the differentiation of Chinese traditional medicines or plants has appeared.
In order to improve the resolution and solubility of analytes, alkaline borate buffer
This journal is © The Royal Society of Chemistry 2000
was employed in this study for baicalein and bacalin and can form negative-charged
complex with boric acid in alkaline solution.
1
The adsorption of analytes on the
surface of working electrode and inner surface of capillary can be inhibited by using
alkaline borate buffer so that the reproducibility of peak current and migration time is
improved.
The effects of the acidity and concentration of running buffer, separation voltage
and injection time on CE are illustrated as in Fig.1.
Fig. 1 Effect of the acidity (A) and the concentration (B) of the running buffer and the
separation voltage (C) on the migration time of baicalein (a) and baicalin (b), effect of
injection time (D) on analyte peak current
The effect of the running buffer pH on the migration time of the investigated
analytes is shown in Fig. 1A. The running buffers were 100 mmol l1 BBs at five
different pH values (8.0, 8.5, 9.0, 9.4 and 10.0). As shown in Fig. 1A, when the running
buffer pH increases, the migration time increases with the resolution improved due to
the dissociation of the hydroxyl group for the analytes. Meanwhile, the peak current is
low and the peak shape becomes poor at pH value above 9.5. At pH 9.0, the analytes
This journal is © The Royal Society of Chemistry 2000
can be well separated within a relatively short time. Fig. 1B indicates that the migration
time and the resolution increases with increasing buffer concentration. However,
higher buffer concentrations (100 mmol l1) also have a negative effect on the
detection limits because the peak currents of both investigated analytes decrease and
the effect of Joule heat is more pronounced. So 100 mmol l1 BB (pH 9.0) is chosen as
the running buffer in this work in considering the peak current, resolution, analytical
time and the buffer capacity.
The influence of separation voltage on the migration time of the analytes is shown
in Fig. 1C. Increasing the voltage gives shorter migration time for both compounds, but
also increases the base line noise, resulting in poorer detection limits. It is found that
higher separation voltages are not beneficial to the resolution. However, too lower
separation voltage will increase the analysis time considerably, which in turn causes
peak broadening. Based on experiments, 12 kV was chosen as the optimum voltage to
accomplish a good compromise. The effect of injection time on CE separation was
investigated by changing the sampling time (2, 4, 6, 8, 10 and 12 s at a voltage of 12 kV,
as shown in Fig. 1D). The peak current increases with increasing sampling time, and
the breadth of the peaks increases simultaneously. In this experiment, 6 s (at 12 kV) is
selected as the optimum injection time.
Through the above experiments, the optimum conditions for determining baicalein
and baicalin are acquired.
Reference
1 Ph. Morin, F. Villard, M. Dreux, J. Chromatogr., 1993, 628, 161