Original / Otros. Jian Zai-you 1,2, Xu Gui-fang 1, Chen Hong-zhi 1, Wang Hong-sheng 1 and Hu Xi-qiao 1

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Nutr Hosp. 2015;32(2):913-917 ISSN 0212-1611 CODEN NUHOEQ S.V.R.

1 Nutr Hosp. 2015;32(2): ISSN CODEN NUHOEQ S.V.R. 318 Originl / Otros Study on the differences of mjor phrmceuticl ingredients in different prts nd processed medicinl mteril of Epimedium Brevicornu Mxim in Tihng mountin Jin Zi-you 1,2, Xu Gui-fng 1, Chen Hong-zhi 1, Wng Hong-sheng 1 nd Hu Xi-qio 1 1 Henn Institute of Science nd Technology, Xinxing Henn. 2 Collortive Innovtion Center of Modern Biologicl Breeding, Henn Province (Chin). Astrct The im of this study is to evlute the medicinl vlues of different prts of Epimedium revicornu Mxim. nd the effect of processing on mjor phrmceuticl ingredients in it. The contents of icriin nd epimedin C in different prts nd processed medicinl mteril of E. revicornu in Tihng Mountin were determined with ultrsonic extrction nd RP-HPLC. The results indicted tht the contents of icriin nd epimedin C, respectively % nd %, in the lef re higher thn tht in other prts. The contents of icriin nd epimedin C, respectively % nd %, in the stem (include petiole) re the lowest. The contents of these ingredients in the root (include rhizome) re close to tht in the lef. The icriin nd epimedin C in ll prts of E. revicornu reduced fter processing. The content of icriin in the processed lef is out 59.5% of tht in unprocessed leves. The effect of prossing on the content of icriin in the stem is unovious. The content of epimedin C in the processed lef is out 33.7% of tht in unprocessed lef. The content of epimedin C in the processed stem (include petiole) is out 36.9% of tht in unprocessed stem. It is worth to exploit the stem nd petiole of E. revicornu ecuse there re certin contents of phrmceuticl ingredients in them. The firepower should e pid ttention to nd the temperture should not e very high to void the dmge on phrmceuticl ingredients in E. revicornu when process it. (Nutr Hosp. 2015;32: ) DOI: /nh Key words: Epimedium revicornu. Medicinl prts. Processing. Correspondence: Jin Ziyou. Henn institute of science nd technology, Huln rod, Xinxing city, Henn province (Chin). E-mil: jin @126.com Reciido: 9-III ª Revisión: 29-III Aceptdo: 3-V ESTUDIO SOBRE LAS DIFERENCIAS DE INGREDIENTES FARMACÉUTICOS MAYOR EN DIFERENTES PARTES Y PROCESADO MEDICINAL MATERIAL DE EPIMEDIUM BREVICORNU MAXIM EN TAIHANG MONTAÑA Resumen Este estudio pretende evlur los vlores medicinles de diferentes prtes de l Epimedium revicornu Mxim y el efecto de su procesmiento sore sus principles componentes frmcéuticos. El contenido de icriin y epimedin C en diferentes prtes y en mteril medicinl procesdo de Epimedium revicornu en l montñ de Tihng fue determindo medinte extrcción ultrsónic y RP-HPLC. Los resultdos indicron que el contenido de icriin y epimedin C, respectivmente 3,4524% y 0,5485%, en l hoj son myores que en otrs prtes. El contenido de icriin y epimedin C, respectivmente 0,1942% y 0,1342%, en el tllo (peciolo incluido) es más jo. El contenido de estos componentes en l ríz (rizom incluido) es similr l de l hoj. El contenido de icriin y epimedin C en tods ls prtes de E. revicornu se vio reducido después del procesdo. El contenido de icriin en l hoj procesd es proximdmente el 59,5% del de l hoj sin procesr. El efecto del procesdo sore el contenido de icriin en el tllo no es evidente. El contenido de epimedin C en el tllo procesdo es de proximdmente el 33,7% del de l hoj sin procesr. El contenido de epimedin C en el tllo procesdo (peciolo incluido) es de proximdmente el 36,9% de quel del tllo sin procesr. Vle l pen provechr el tllo y peciolo de l E. revicornu porque hy cierto contenido de componentes frmcéuticos en ellos. Hy que controlr l potenci de fuego y l tempertur no dee ser muy lt pr evitr dñr los componentes frmcéuticos de l E. revicornu. (Nutr Hosp. 2015;32: ) DOI: /nh Plrs clve: Brevicornu epimedium. Prtes medicinles. Trtmiento. 913

2 Introduction Epimedium revicornu Mxim is perennil herceous plnt 1. The lef of E. revicornu is kind of trditionl Chinese medicine. Mny of the medicinl chemicl components such s icriin, cohuoside, ohuoside, epimedin A, epimedin B, nd epimedin C re flvonoids 2,3. As trditionl Chinese medicine, the lef of E. revicornu is nmed epimedii folium with tonic, nti-rheumtic nd phrodisic effects. Epimedii folium cn e used to cure impotence, emission, osteomlci, rheumtism, poplexy nd so on 4. E. revicornu widely distriute in Chin. There is rich resources of E. revicornu in Tihng Mountin. The stem of E. revicornu is herceous nd its compound leves consist of nine leflets. Its petiole rnches nd form trifoliolte lef. The lef is thin nd like pper. The totl iomss of stem nd petiole is close to tht of lef lde. The prt of E. revicornu used s Chinese medicine is lef prescried in Chinese Phrmcopoei 4. Therefore, these stem nd petiole re wsted when hrvest E. revicornu. We determined the contents of mjor phrmceuticl ingredients in different prts of E. revicornu distriuted in Tihng Mountin to improve rtionl usge of the resources. In trdition, The processing of epimedii folium is roiling with suet oil to increse its effect. There re differences in the reports on the contents of mjor phrmceuticl ingredients in the processed epimedii folium. Therefore, the differences etween the contents of mjor phrmceuticl ingredients in the processed epimedii folium nd tht in unprocessed mterils were lso studied in our pper to promote the proper usge nd rtionl processing of epimedii folium. Mterils nd methods Regents nd mterils Instruments: HPLC instrument, Shimdzu, MS-2010, Lectronic Anlytic Blnce (precision: ), Ultrsontor nd Rotry Evportor were used in study. Regents: methnol (AR), ethnol (AR) nd cetonitrile (HPLC grde) were used in experiment. Stndrd icriin nd epimedin C (99.8 %) were purchsed from Ntionl Institute for the Control of Phrmceuticl nd Biologicl Products in Octoer Mterils: The E. revicornu mterils were collected in Gunshn, Xigou nd Jiulinshn of Xinxing city in Henn province in June plnts were rndomly dug in ech loction. The lef, stem (include petiole) nd root (include rhizome) were seprted nd drought to constnt weight t 45º C. Methods Process mterils: The leves of E. revicornu were cut into slices of 5-7 mm in width. The stems (include petioles) were cut into sections of mm in length. 10 g suet oil ws melted in pn with smll fire nd then 50 g E. revicornu mterils ws put into the pn. The mterils ws stired nd fried in the pn with smll fire. The fried mterils ws tken out when the suet oil completely infiltrted into the mterils nd these mterils ppered with glossy sheen. Preprtion of extrct: The E. revicornu mterils ws crushed nd sieved with 80 meshes sieve. 1 g mterils ws extrcted with 20 ml ethnol solvent (70%) in n ultrsonic th for 1 h. The mixture ws filtered nd the residue ws extrcted with sme solvent once gin. These filtrte ws merged nd dded to 40 ml. The extrct ws filtered using 0.22 µm memrne filter. Ech kind of mterils ws extrcted respectively three times. Determine the contents of icriin nd epimedin C in extrcts: The HPLC column used ws Dimonsil C18 reverse-phse column (5 µm, mm). The volume of extrct injected ws 10 µl, nd then elution ws performed with grdient moile phse consisting of cetonitrile nd wter. The content of cetonitrile in the grdient moile phse vries s elow (v/v): from 22 to 29 % in 0 12 min, 29 to 29.5 % in min nd 29.5 to 30 % in min. The flow rte ws 1 ml/min nd the temperture in HPLC column ws 35º C. A vrile wvelength recorder set t 270 nm ws used to detect ingredients eluted from the column. Stndrd icriin solutions were prepred t mg/ml, 0.15 mg/ml, 0.25 mg/ml, 0.5mg/ ml, 2.5 mg/ml nd 5 mg/ml respectively. Stndrd epimedin C solutions were prepred t mg/ml, 0.05 mg/ml, 0.15 mg/ml, 0.25 mg/ml, 0.5mg/mL, 2.5 mg/ml nd 5 mg/ml respectively. These stndrd solutions were nlyzed ccording to the ove HPLC method. Chromtogrphy pek res of icriin nd epimedin C in stndrd solutions were respectively recorded to prepre stndrd curves relting these pek res to their contents. All of those prepred extrcts were nlyzed ccording to the ove HPLC method. Chromtogrphy pek res of icriin nd epimedin C in extrcts were respectively recorded respectively. These contents of icriin nd epimedin C in extrcts were nlyzed ccording to their chromtogrphy pek res nd the stndrd curves. All of the dt were nlyzed with SPSS (Sttisticl Product nd Service Solutions). Results nd nlysis Stndrd curves of icriin nd epimedin C The HPLC chromtogrms of stndrd icriin nd epimedin C re showed in figure 1. These stndrd curves of icriin nd epimedin C were drfted ccording to their pek res nd their contents. Stndrd curve of icriin is y = x (x: content, y: pek re, R 2 = ). The retention time 914 Nutr Hosp. 2015;32(2): Jin Zi-you et l.

3 of icriin is min. Stndrd curve of epimedin C is y = x (x: content, y: pek re, R 2 = ). The retention time of epimedin C is min. Differences etween these contents of icriin or epimedin C in different prts of E. revicornu The peks of icriin nd epimedin C in extrct chromtogrms were identified ccording to their retention time in HPLC (Fig. 2). The contents of icriin nd epimedin C in extrcts nd unprocessed mterils were nlyzed ccording to their pek res nd stndrd curves lso (Tle I). The results showes tht there re very significnt difference etween these contents of icriin or epimedin C in different prts of E. revicornu. These contents of icriin nd epimedin C in stem (include petiole) re lowest nd tht in leves re highest. The content of epimedin C in root (include rhizome) is out 65.4% of tht in leves. However, the content of icriin in root (include rhizome) is only out 14.9% of tht in leves y = x R 2 = y = x R 2 = min Fig. 1. HPLC Chromtogrms of stndrd icriin nd epimedin C (: epimedin C, : icriin). (x100,000) min Fig. 2. HPLC chromtogrms of extrct from unprocessed E. revicornu leves (: epimedin C, : icriin). Study on the differences of mjor phrmceuticl ingredients in different prts nd processed medicinl mteril... Nutr Hosp. 2015;32(2):

4 Root (include rhizome) 1 Root (include rhizome) 2 Root (include rhizome) 3 Stem (include petiole) 1 Stem (include petiole) 2 Stem (include petiole) 3 Leves 1 Leves 2 Leves 3 Tle I The contents of icriin nd epimedin C in different prts of unprocessed E. revicornu (%) Prts Epimedin C (%) Iicriin (%) c Anov f= > f 0.01 =10.9 f= > f 0.05 =10.9 Note:, nd c men the results of multiple comprisons c Content vrinces of icriin nd epimedin C in mterils efore nd fter processing The representtive HPLC chromtogrm of extrct from processed E. revicornu mterils is showed in figure 3. The contents of icriin nd epimedin C in these extrcts of processed mterils were nlyzed ccording to their pek res nd stndrd curves lso (Tle II). The contents of icriin nd epimedin C in E. revicornu mterils decresed in different degree fter processing. The content of icriin in processed leves, out 59.5% of tht in unprocessed leves, oviously decresed. But there is not ovious difference etween the contents of icriin in processed stem (include petiole) nd tht in unprocessed stem (include petiole). The contents of epimedin C in E. revicornu mterils oviously decresed fter processing lso. The content of epimedin C in processed leves is only out 33.7% of tht in unprocessed leves. And the content of epimedin C in processed stem (include petiole) is out 36.9% of tht in unprocessed mterils. Discussion The utilized prt of E. revicornu is lef in trditionl Chinese medicine, which possily is relted to 2.00 (x100,000) min Fig. 3. Representtive HPLC chromtogrms of extrct from processed E. revicornu mterils (: epimedin C, : icriin). 916 Nutr Hosp. 2015;32(2): Jin Zi-you et l.

5 Ingredients nd processing Tle II The contents of icriin nd epimedin C in processed nd unprocessed E. revicornu (%) Stem (include petiole) 1 Stem (include Stem (include Averge Leves 1 Leves 2 Leves 3 Averge petiole) 2 petiole) 3 Epimedin C Unprocessed Processed Anov f=88.267>f 0.01 =34.1 f=590.32>f 0.01 =34.1 Icriin Unprocessed Processed Anov f=0.310<f 0.05 =10.1 f=17.958>f 0.05 =10.1 the high contents of medicinl ingredients nd evident efficcy. The result of our investigtion showes tht the iomss of stem nd petiole in E. revicornu is equl to (out 90%) tht of leves. Although the content of icriin in stem (include petiole) is lower thn tht in leves, ut the contents of epimedin C in them re pproximtive. Therefore, there is high medicinl vlue in the stem nd petiole of E. revicornu. We cn extrct medicinl ingredients from the stem nd petiole. The stem, petiole nd root re utilized s Chinese medicine in some plce in Chin 5. Although the contents of medicinl ingredients in the root nd rhizome of E. revicornu re very high. But excvting the root nd rhizome is serious destructive to E. revicornu resources nd lso hrmful to sustinle utiliztion these resources. Therefore, the root nd rhizome of E. revicornu should not e utilized. Some icriin nd epimedin C, specificlly epimedin C, cn e dmged in structure when E. revicornu mterils is processed. The structure of epimedin C is more complicted thn tht of icriin nd more decomposed thn icriin in processing 6,7. All of these cuse the contents of icriin nd epimedin C to decrese. The trditionl Chinese medicine ws decocted in wter efore it e tken s medince. The soluility of these medicinl ingredients in wter is lower thn tht in orgnic solvents. The suet oil is used s ccessories in processing E. revicornu mterils. There is some suet oil in the E. revicornu mterils fter processing. The suet oil is lso kind of orgnic solvent, which increse the soluility of these medicinl ingredients. Owing to these fctors, the contents of medicinl ingredients in processed E. revicornu mterils re higher thn tht of unprocessed ones. In ddition, there re some medicinl ingredients in suet oil 8. Therefore, the efficcy of processed E. revicornu mterils is evidenter thn tht of unprocessed ones. Since processing cn dmge some medicinl ingredients, The firepower should e pid ttention to when E. revicornu mterils is processed. The temperture should not e too high to mke some medicinl ingredients decompose. Conclusion There re ovious differences etween these contents of icriin or epimedin C in different prts of E. revicornu. The content of icriin or epimedin C in the lef is the highest nd tht in the stem (include petiole) is the lowest. The contents of icriin nd epimedin C in ll prts of Epimedium revicornu reduced more or less fter processing. The decrese of epimedin C is ovious. Interests Declrtion The uthors of this rticle declre tht they hve no conflict interests. References 1. Flor of Chin (volume 27). Beijing, Chin, Science Press, 1979: LI Yu-o, MENG Fn-ho, LU Xiu-mei, LI F-mei. Chemicl constituents from her of Epimedium revicornum. Chin Journl of Chinese Mteri Medic 2005, 30(8): MENG Ning, KONG Ki, LI Shi-weng. Advncesin Studieson Chemicl Constituents nd Phrmceuticl Activitiesin Species of Epimedium. Act Bot. Bore1 Occident. Sin 2010, 30(5): Chinese Phrmcopoei (2010 edition, prt I). Beijing, Chin, Chemicl industry Press 2005: JIA Xin-sheng, WU Ji-qi, MAO Qing. Study on the chemicl constituents in the root of Epimedium cumintum Frnch. Chin Journl of Chinese Mteri Medic 1998, 23(3): ZHANG Hi-yn, KANG Jin-xun, DONG Jin-jun, ZHAO Tin-zeng, WU Ming-jin. NM R Study on Epimedin C. Journl of Zhengzhou University (Engineering Science), 2013, 34(4): CHENG Yn, W ANG Ni-li, W ANG Xin-lun, ZHANG Dwei, HUNA G W en-xiu, YAO Xin-sheng. Chemicl constituents from Epimedium korenum Nk. Journl of Shenyng Phrmceuticl University 2006, 23(10): SUN E, WEI Ying-jie, ZHANG Zhen-hi, CUI Li, XU Fengjun, JIA Xio-in. Processing mechnism of Epimedium fried with suetoilsed on sorption nd metolism of flvonoids. Chin Journl of Chinese Mteri Medic 2014, 39(3): Study on the differences of mjor phrmceuticl ingredients in different prts nd processed medicinl mteril... Nutr Hosp. 2015;32(2):

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