Certification Reevaluation for Control Reference Standards of

Certification & Reevaluation for Control Reference Standards of Herbal Medicines Do Hoon Kim, Keum Ryon Ze, Eun Kyung Kim, Ju Young Park, Jong Hwa Lee, Byong Hee Lee, Jin Sun Sin, Jong Hwan Kim, Rack Seong, Eun Hee Kim, Young Joong Kim*, Seung Yeup Chang Herbal Medicine Standardization Team Korea Food & Drug Administration, Seoul 122 -704, Republic of Korea *Pharmacy of Seoul National University, Seoul 151 -742, Republic of Korea To promote the quality control of herbal medicines, we have prepared and distributed control reference standards. These reference standards are prepared first by Botanical scientists and reviewed by KFDA staff. Reference standards is very important and essential quality control of herbal medicines. And the purity is important In reference standards. But reference standards can be change their properties and purity as time goes by. Therefore we have controled reference standards constantly. As one of constant management for reference standards, we’re reconfirmed reference compounds’ purity by identifical and analytical methods. In this study, we reviewed the process of making reference standards and introduce control refernce standards that we have distributed. [Keyword – Herbal medicines, Reference standards, Marker substance, HPLC] Introduction Reevaluations The Reference standards are very important and essential for quality control of herbal medicines. They are required for identification, assay and purity test etc. We have prepared reference medicinal plant materials and control reference standards and intended promote the quality control of herbal medicines. By botanical scientists, herbal materials are collected, identified its origin and then evaluated its quality and specification. Marker substances are prepared from medicinal plant materials through extraction, seperation and purification. All materials are tested and characterized in the accordance with the methods of the Korea Pharmacopoeia and the Korea Herbal Pharmacopoeia etc. KFDA staff scientists review and reevaluate the prepared control reference standards and then establish them as the official reference standards. In this study, we will introduce the example of making process for marker substances and the control reference standards. (Magnolol) Making Process & Certificaions Ø First, most important point is origin identification of herbal medicine. And then, it extracted by proper solvent. Next, it isolated from extract by chromatographic methods. Isolated compound was identified and evaluated by spectral data and analysis. Ø Isolation of Schizandrin* Schizandra Fructus(2 kg) was extracted by methanol for 90 min three times. It was obtained extract(200 g). And then it was suspended with water, n-hexane fraction(50 g) was obtained by solvent fractionation. Crude extract contained schizandrin was obtained by step gradient silica gel column chromatography with n-hexane, ethyl acetate. Then schizandrin was obtained by reverse phase column chromatography (Acetonitrile: Water=1: 2). And then schizandrin was purified by replication with mix solution(Acetonitrile : Water=1: 2). Ø Identification of Schizandrin* Schizandrin isolated from Schizandra Fructus was identified spectral data and reference data. This data was shown Schizandrin structure. Another data(ex. UV, IR, mp etc) was shown features of schizandrin. Ø Reevaluation We analyzed the purity of making and distributing control reference standards. First, we identified substance by HPTLC. And then analyzed by HPLC. We confirm purity of standards by area % of HPLC results. All test was based on experiment method of Korean Pharmacopoeia and Result report(A study on the preparation of Standard compounds from Herbal medicine* ). Ø Reevaluation - Identification by HPTLC - HPLC analysis v Magnolol - n-Bu. OH: Water: Acetic acid(glac. ) = 4: 2: 1 - Identify : Dragendorff Reagent - Purity calculation Method v We analyzed purity of Schizandrin and another standards(working standard, especially confrim the purity by certification). v Method 1 – Absolute Peak Area Percent Calculation This Method was calculated by area percent of HPLC analysis. As spectral data, the data was integrated and calculated by area percent. v Method 2 – Relative Peak Area comparision Calculation This Method was relative area comparision. Relative standards was used working standards(we used Chromadex standards) confirm by various analysis data(ex. HPLC analysis, NMR, Mass, UV etc). It is analyze d each standards by same instrument, methods. And then we compared the area of each standards. In this method, purity of standard is trustworthy. v Result – We tested purity by two methods. The Result was shown 97. 9%(Method 1), 98. 23%(Method 2). The result was shown similar values. So, we confirmed the purity of Schizandrin. Ø The purity of Magnolol was 99. 0 % in 2002. And now, the purity of Magnolol we reevaluated is 98. 0%. So, The purity of magnolol was downed 1% value. CONCLUSION To promote the quality control of herbal medicines, we have prepared and distributed control reference standards. Reference standard is very important and essential quality control of herbal medicines. But reference standards can be change their properties and purity as time goes by. As one of constant management for reference standards, we have reevaluated purity. In this study, we reviewed the making process, storage, distribution, reevaluation of control reference standards. Ø Certificate of Standards* Each standards reported each certification. This resulted from spectral data(UV, IR, NMR etc. ) and HPLC analysis. * Magnolol was made in 2002. In that time, the purity of magnolol was 99. 0 % * Young Joong Kim, Seoul National University. *Corresponding author: keumryon@kfda. go. kr We will manage reference standards constantly. As one of management, we will make out Certification of reference standards(Right Example). All cases will be applied to certification for control reference standards. * Certificatie of Analysis(Right Example) - Related Chemical compound - Related Adjusted purity Ø Distribute Standards (25 Standards) - Hesperidin, Ephedrine․HCl, Amygdalin, Berberine․Cl, Baicalin, Puerarin, Magnolol, Loganin, Schizandrin, Paeoniflorin, Geniposide, Gentiopicrin, Ginsenoside Rg 1, Icariin Paeonol, Poncirin, Ginsenoside Rb 1, Sennoside A, Sennoside B, Betaine, Eugenol, 6 -Gingerol, Saikosaponin A, TanshinoneⅡA, Decursin.