EMPLOYMENT OF ATR-FTIR AND HPLC-UV METHOD FOR DETECTION AND QUANTIFICATION OF ANDROGRAPHOLIDE Original Article OKTAVIA INDRATI 1,2 , RONNY MARTIEN 1 , ABDUL ROHMAN 3 , AKHMAD KHARIS NUGROHO 1* 1 Department of Pharmaceutics, Faculty of Pharmacy, Universitas Gadjah Mada, Sekip Utara, Yogyakarta Indonesia 55281, 2 Department of Pharmacy, Faculty of Mathematics and Natural Sciences, Universitas Islam Indonesia, Jl. Kaliurang Km 14,5 Sleman Yogyakarta Indonesia 55584, 3 Received: 23 Jul 2018, Revised and Accepted: 04 Sep 2018 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah Mada, Sekip Utara, Yogyakarta Indonesia 55281 Email: oktavia.indrati@uii.ac.id ABSTRACT Objective: The purpose of this present study was to describe the employment of infrared (IR) spectroscopy and high-performance liquid chromatography with ultraviolet detection (HPLC-UV) method in determining the identity and purity of bulk material containing andrographolide. Methods: Attenuated total reflectance (ATR)-Fourier transform infrared (FTIR) spectroscopy was carried out in transmittance mode to investigate the molecular vibration of the bulk material component and the similarity of functional groups between bulk material and standard andrographolide. Meanwhile, the liquid chromatographic analyses were performed on a reversed-phase C18 column and under UV detection at 224 nm to determine the andrographolide content of the bulk material. Results: The obtained ATR-FTIR spectra indicated that functional group of the bulk material were in close similarity with those of standard andrographolide. The linearity of the evaluated method achieved over a concentration range of 1 to 60 μg/ml with a high correlation coefficient (0.999). By using the studied HPLC-UV method, the andrographolide of bulk material was found to be 98.12% (retention time of 2.58 min). Conclusion: The studied HPLC-UV method of andrographolide determination is accurate, precise, selective, and brief in terms of analysis time. The studied method, therefore, provides a rapid and reliable assessment for identifying and determining the purity of andrographolide bulk material. Keywords: Andrographolide, Bulk material, Determination, HPLC-UV, Identity, Purity © 2018 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open-access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/) DOI: http://dx.doi.org/10.22159/ijap.2018v10i6.28691 INTRODUCTION Andrographolide (fig. 1) is a diterpenoid substance isolated from the medicinal plant of Andrographis paniculata. This compound becomes the point of interest since A. paniculata herb is widely used as a traditional medicine in several Asian countries such as India, Indonesia, China, Malaysia, and Thailand [1, 2]. A large number of studies were conducted to isolate, identify, and quantify this substance [3–5] as well as assess its pharmacological activities [6-8], and its safety [9, 10]. Andrographolide is reported to exhibit various biological activities, including antidiabetic [11], anticancer, anti-inflammatory [6], antivirus, and hepatoprotective activities. Despite the many pharmacological activities, it is reported that andrographolide has poor aqueous solubility and is extensively metabolized leading to its low oral bioavailability [2, 12]. Hence, numerous studies were performed to develop andrographolide containing dosage form in order to enhance the acceptability, solubility, and bioavailability of this bioactive compound. Numerous dosage forms such as microcrystal [13], microemulsion [14], micelle [15], nanoparticle[16], solid lipid nanoparticle [17], spray-dried [18] and suspension [19] of andrographolide have been successfully developed. Fig. 1: The chemical structure of andrographolide The quality of the bulk material is an important factor that has to be considered during a dosage form development process and routine analysis before drug manufacturing. Bulk material quality affects the quality of the finished product. Several authors have studied the quantification of andrographolide isolated from Andrographis paniculata herbs [3, 4, 20, 21], but none of them reported the determination of andrographolide bulk material’s identity and purity. ATR-FTIR spectroscopy offers a rapid and direct analytical technique without sample preparation as the advantage [22, 23]. Meanwhile, HPLC-UV method is a simple and rapid technique for quantification of drug substance compared to other techniques [24], which becomes the method of choice for bulk and dosage form analysis [25-28]. Therefore, this study aims to describe the determination of andrographolide content in bulk material employing infrared spectroscopy and HPLC-UV method. MATERIALS AND METHODS Chemical and reagents Andrographolide bulk material was purchased from Sinobright Pharmaceutical, China while andrographolide standard (98%) was obtained from Sigma. Methanol for HPLC (J. T Baker) was used as the mobile phase. ATR-FTIR(Attenuated total reflectance-Fourier transform infrared) The powder of bulk material and standard andrographolide were investigated in ATR-FTIR to determine the molecular functional groups of the component and the similarity between these two samples. IR spectra were recorded by using Spectrum Two FTIR spectrometer (Perkin Elmer, USA) with a single reflectance horizontal ATR cell. The analyses were made in transmittance mode with a small amount of sample deposited on the ATR crystal. The transmittance was measured in the frequency range from 400 cm -1 to 4000 cm -1 with resolution of 4 cm -1 . International Journal of Applied Pharmaceutics ISSN- 0975-7058 Vol 10, Issue 6, 2018