Bicyclic diterpenoid lactone andrographolide is regarded as a “natural antibiotic” as it is known to exhibit a range of bioactivities including anti-inflammatory, antibacterial, antipyretic, antineoplastic, cardioprotective, hepatoprotective and hypoglycaemic, and is present in Andrographis paniculata. The aim of this article is to review the information on analytical methods for andrographolide in biological samples, pharmaceutical formulations and plant materials. This article includes various techniques such as Spectrophotometry, Chemiluminescence method, Electroanalytical method, Chromatography and various hyphenated techniques.

Objective: To develop a novel, accurate, precise and linear reverse phase high performance liquid chromatographic (RP-HPLC) method for simultaneous qualitative and quantitative estimation of curcumin, piperine and camphor in an ayurvedic formulation and validate as per international conference on harmonization (ICH) guidelines.Methods: In the present work, good chromatographic separation was achieved isocratically using a shim-pack HPLC C18 column (4.6 x 250 mm, 5μm) and mobile phase consisting of 0.02 M potassium dihydrogen orthophosphate buffer (pH adjusted to 3.5 with orthophosphoric acid) and acetonitrile in the ratio 40:60, at flow rate of 1 ml/min and column temperature maintained at 35 °C. The effluents obtained were monitored at 255 nm with UV-visible detector.Results: The retention time of curcumin, piperine and camphor was found to be 6.57 min, 7.32 min and 8.57 min respectively. Linearity of curcumin and camphor were found in the range of 4-8 ppm and that of piperine was found to be 5-9 ppm. The correlation coefficient for curcumin, piperine and camphor were 0.998, 0.99 and 0.994 respectively. The high recovery values (98 %-102 %) indicate a satisfactory accuracy. The low percent relative standard deviation (% RSD) values in the precision study reveals that the method is precise.Conclusion: The developed method is novel, simple, precise, rapid, accurate and reproducible for simultaneous quantitative estimation of curcumin, piperine and camphor in an ayurvedic formulation. Hence the developed method can be used for quantitative analysis and quality control of extracts and commercial samples of other species containing these three markers.

A simple, rapid, and specific reversed-phase HPLC method with DAD detection has been used for analysis of two flavonoids, quercetin and kaempferol, in Cissus quadrangularis Linn. The flavonoids were well resolved within 10 min, and quantification was achieved, on an endcapped C 18 column at 370 nm with acetonitrile-phosphate buffer (pH 3.4, adjusted with glacial acetic acid) 60:40 (v/v) as isocratic mobile phase at a flow rate of 1.0 mL min −1 . The method was validated for limit of detection, limit of quantification, linearity, precision, accuracy, and recovery. Linearity was demonstrated over the range 0.75 to 10 μg mL −1 for quercetin and 1.0 to 10 μg mL −1 for kaempferol with good correlation coefficients (r 2 > 0.998). Detection limits were 0.075 and 0.10 μg mL −1 for quercetin and kaempferol respectively. Recovery was 98.0-105.3% for quercetin and 95.0-101.1% for kaempferol. The method was successfully applied to analysis of two flavonoids in four samples of Cissus quadrangularis Linn.

Background: A tablet dosage form widely used in the treatment of cough and cold, containing phenylephrine hydrochloride, paracetamol, caffeine, and diphenhydramine hydrochloride as active pharmaceutical ingredient was selected for the development of a novel, rapid, simultaneous isocratic reversed phase-high performance liquid chromatography (RP-HPLC) method. Objective: The objective of this paper was to develop and validate a novel, rapid, simple, precise, accurate and reproducible RP-HPLC method for simultaneous estimation of phenylephrine hydrochloride, paracetamol, caffeine, diphenhydramine hydrochloride in bulk and pharmaceutical dosage form. Method: Optimized chromatographic conditions were an isocratic elution with prontosil C18-column (250×4.6mm,5µ), methanol and 20mM phosphate buffer (55:45 v/v, pH 3 ) as mobile phase, flow rate 1.0 mL/min and UV detector set at λ max 220 nm. The method was validated for specificity, precision, linearity, accuracy, sensitivity, and robustness as per the International Council for Harmonization guidelines. Result: The retention times of phenylephrine hydrochloride, paracetamol, caffeine, diphenhydramine hydrochloride were found to be 2.8 min, 3.3 min, and 4.0 min and 7.3 min, respectively. This novel method was found to be rapid, simple, linear (R2> 0.99), preciserelative standard deviation < 2.0 %), accurate (recovery 98-102%), sensitive and robust. Conclusion: The proposed novel isocratic RP-HPLC method is rapid (short run time below 10min), highly selective, precise, accurate , sensitive and robust. The method was successfully applied for the simultaneous analysis of phenylephrine hydrochloride, paracetamol, caffeine, diphenhydramine hydrochloride in a pharmaceutical dosage form.

Objective: The objective of this study was to develop and validate a novel, simple, rapid, precise and accurate reversed-phase high performance liquid chromatographic (RP-HPLC) method for simultaneous quantitative estimation of berberine, quercetin, and piperine in Ayurvedic formulation.Methods: The chromatographic separation was achieved using a stationary phase C18 shim-pack (150 mm x 4.6 mm, 5µ) column and mobile phase consisted of acetonitrile: 0.04 M potassium dihydrogen phosphate buffer (pH 3.0 adjusted using orthophosphoric acid) in a ratio of 65:35 v/v, with a flow rate of 1 ml/min and UV detection at 255 nm.Results: The retention time of berberine, quercetin, and piperine were found to be 2.7, 3.0 and 6.3 min respectively. Linearity for berberine, quercetin, and piperine were found in the range of 12-28 µg/ml. All calibration curve showed good linear correlation coefficients (r2˃ 0.999) within the tested ranges. Mean percent recoveries for berberine, quercetin, and piperine were found to be within the acceptance limits (98-120%). The percent relative standard deviation (% RSD) for precision was found to be less than 2% which indicates method is precise.Conclusion: The developed method is novel, simple, precise, accurate and can be used for quantitative analysis and quality control of the raw material as well as other commercial formulations containing these three markers.