Summary. Densitometric HPTLC and HPLC have been used for quantification of
p-coumaric and protocatechuic acids in an ethereal fraction from a methanolic extract of
Aquilegia vulgaris L. HPLC analysis was performed on an RP-18 column with methanol–
water–formic acid 25:75:0.5 (v/v) as mobile phase. Thin layer chromatography was performed
on Si60 F254 HPTLC plates with mixtures of heptane, dichloromethane, diisopropyl
ether, formic acid, and water as mobile phases. Satisfactory separation of the phenolic
acids was achieved by use of the multiple gradient development technique. The
quantities of p-coumaric and protocatechuic acids determined by HPLC were 0.374 and
2.283 mg g−1 dry plant material, respectively; HPTLC results were somewhat higher –
0.396 and 2.584 mg g−1, respectively. The precision of both methods, expressed as relative
standard deviation, was satisfactory. The methods are useful for quality control of
Aquilegia vulgaris extracts.
Conclusion
We have established procedures for standardisation of Aquilegia vulgaris extracts
on the basis of the amounts of two phenolic acids present. The conditions
(mobile and stationary phases) used for separation of protocatechuic
and p-coumaric acids from other constituents of the ether extracts were optimised
for both techniques. Gradient multiple development (GMD) was
used in HPTLC and isocratic elution in HPLC. The advantage of the HPTLC
method over HPLC is that the former is simpler, less time-consuming, requires
less sample pretreatment, and uses less solvent. Because both methods
(HPTLC and HPLC) provide comparable results, both are appropriate
for standardization of the Aquilegia vulgaris extracts on the basis of the
amounts of the main phenolic acids.
References