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COMMITTE

THE ORGANIZING COMMITTEE OF INTERNATIONAL CONFERENCE ON"NEW DEVELOPMENT OF PHARMACEUTICAL CARE IN PHARMACOGENETIC

AND PHAR,TAACOGENOMIC APPROACH,,

Advisory Board

Steering Committee

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4.

Rector University Muhammadiyah of MalangDr. Muhajir Effendy., M.AP

1. Dean of Facufty of Health sciences University Muhammadiyah of Malang2. Assistant Dean I of Faculty of Health sciences university Muhammadiyah

of MalangAssistant Dean ll of Faculty of Health sciences university Muhammadiyahof MalangAssistant Dean lllof Faculty of Health sciences University Muhammadiyahof Malang

5. Expert staff of Department of pharmacy, Faculty of Health sciencesUniversity Muhammadiyah of Malang

Nailis Syifa' S.Farm., Apt., M.Sc

Siti Rofida., S.Si., Apt., M.FarmHeru Prabowo Hadi., S.Farm., AptDra. Uswatun Chasanah., Apt., M.Kes

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proceeding Editors and Event OrganizerHidajah Rachmawati., S.Si., Apt., Sp.FRSSiti Rofida., S.Si., Apt., M.FarmNikmatul lkhrom E.J.,S.Farm.,M.FarmKlimAsri Julia Sukmaningrum., S.Farm., AptDyan Retno., 5. Farm., Apt.Samiyah Abdullah., S. Farm., Apt.

LogisticSekar Uli Fatmarini., S.Farm., AptAkhmad Sobrun Jamil,, S.Si.,MpBunga Prahesti Septiyani

@ThelnternationalConferencePharmaceuticalCare

Ahmad FirdiansYah', S'Farm'' APt

Sendi Lia Yunita', S'Farm'' APt

Raditya Weka'Nugraheni' S'Farm'' Apt

Accommodation, Documentation and Conference Services

' Lukman HakimFerdiansYahAYu EvitasariDani Putra

URIC ACID INHIBITION ACTIVITY OF PLUCHEA INDICA L LEAF

EXTRACTED ETHANOL IN HYPERURICEMIA INDUCED MICE

Uswatun Chasanah. Myrna Dian RahmawatiDepartment of pharmacy, Faculty of Health, University of Muhammadiyah, Malang, lndonesia

ABSTRACT

Background : Gout is a clinical syndrome resulting from the deposition of urate crystals in joints

causing inflammation. lt has been known that the leaf of Pluchea indica L. contained flavonoid that

thought to have activity of anti-hyperuricemia by inhibit xanthine oxidase (XO).

Objective : This study was conducted to explore the activity of Plucea indica leaf extracted ethanol on

reducing the uric acid level in potassium oxonate treated male balb/c mice (7mgl20g body weight,

intra-peritoneal).

Methods : Male balb/c mice were exposed by Pluchea indica L. leaf extracted ethanol with the doses

of 4mg,L}g body weight, 15mg/20g body weight and 64mg/20g body weight, respectively per-oral at

60 minutes after induction of potassium oxonate, subsequently the observation was done at 60 minutes

after that.

Results : Analysis showed that p/uchea indica L leaf extracted ethanol can reduce the uric acid level

significantly at dose of 64mgl2og body weight (p=0.05, ANovA)'

Gonclusion : Therefore, the pluchea indica leaf is not potent to develop for hyperuricemic medicine-

Keyword : gout, Pluchea indica L, hyperuricemic mice

INTRODUCTION

Gout is one of the oldest and better understood of the rheumatic diseases. lt is characterized by

chronic hyperuricemia (> 450 pmol/l or > 7.0 mg/dl in men, and > 360 pmol/l or > 6'0 mg/dl in women),

recurrent attacks of acute arthritis provoked by release of sodium urate crystals into jolnts, and eventual

development in some patients of urate tophi, chronic tophaceous gouty arthritis, and urate nephropathy

(Kersley GD, 1971). Xanthine oxidase (xo) is an enzyme responsible for catalyzing the oxidation of

hypoxanthine to xanthine and of xanthine to formation of uric acid (Owen and Johns, 1999; Ramallo et

al., 2006). The treatment of gout entails the use of therapeutic agents such as xanthine oxidase inhibitors

(XOl) (Kong et al.,ZOAq Unno et at.,2004).XOl acts by blocking the biosynthesis of uric acid from purine

in the body (Unno et al., 2OO4) and it is believed that either by increasing the excretion of uric acid or

reducing the uric acid production helps to reduce the risk of gout (Umamaheswari et al., 2OO7).

Allopurinol is one of the many known synthetic XOls, it's has an active metabolite Oxypurinol

(Alloxanthine) which works by inhibiting the enzyme xanthine oxidase which catalyzes the formation of

Table of Contens

Study of Clopidogrel Bisulfate Raw Material Properties of the Thermic TreatmentTitta Hartyana Sutarna, Fikri Alatas, Hestiary Ratih, Novi Mega L B7

Uric Acid lnhibition Activity ol Pluchea indica L Leaf Extracted Ethanol in Hyperuricemiainduced MiceUswatun Cha1q1gh, Myrna Dian Rahmawati .......... .t4O

The use of Furosemide ln Patients with chronic Kidney DiseaseNailis Syifa', Didik Hasmono, Fitri Surya Kartika Dewi .......... ................... .. lM

Analysis of Fatty Acid Levels and Microbial Contamination for Devetopment of VirginCoconut Oil Syrup in Tengkudak Village, Tabanan, BaliMade Ary Sarasmita, I Wayan Karta ......... 1g

Optimization of Derivative Beaction Atenotol with 1-Fluoro-2,4-Dinitrobenzene (FDNB)for Quantitative Determination Atenolof by HPLC-UVL.P.M.K. Dewi, 5. Martono, L. Hakim $z

lsolation and ldentification Lactic Acid Bacteria from the Goat Mitk and thisAntimicrobiaf Test to Escherichia coli and Shigella dysenteriaeMira Andam Dewi, Ririn Puspadewi, Agnes Dwi Aryani .......... 159

lmpact of lnsurance Status on Appropriate Utilization Antihypertensive MedicationsBased on Jnc-7 And clinical outcomes lnpatients of "x" Hospital BantutF. Rahmawati, Akrom, W. 5upadmi................. i63

Validation Of High Performance Liquid Chromatography Method For Determination OfSulfamethoxazole And Trimethoprim ln pharmaceutical Dosage FormEngrid Juni Astuti, Dian Ermawati, Harjana, Achmad lnoni .......... 1t"l

xi

Uric Acid lnhibition Activity of pluchea lndica... 141

uric acid from hypoxanthine via xanthine (Katzung, 2007) However, allopurinol generates superoxide (Berryand Hare, 2004) and some people develop rash as they are allergic to allopurinol (Wallach, 199g; Kong etal., 2OO0). Severe reactions also occur including liver function abnormalities (Fields et al., t9g6), a fitalcomplication known as "allopurinol hypersensitivity syndrome" (Umpie'rrez et al., 199g; Kong et at.,20OO)and adverse drug reactions such as Toxic Epidermal Necrolysis syndrome (TENS) pacher et at., 2OA6).

lndonesia has megabiodiversitas plant. Biodiversitas which is often used by humans who havebioactived content. Flavonoids has activity in inhibiting the enzyme xanthine oxidase to uric acidformation is inhibited (Cos et al., 1998).lt is reported that the flavonol class of flavonoids is quercetine,myricetine, kaempferole and flavones group is chrisine and luteoline had the greatest activity inlowering blood uric acid levels by inhibiting the enzyme xanthine oxidase (Nagao et at., 1999). Sonchusarvensis is a plant of the Asteraceae family, which has been shown to has good potential for theinhibition of the enzyme xanthine oxidase that can lower uric acid levels in the blood (Cos eta/.,199g).Sonchus arvensis been identif ied containing flavonoids kaempferol, quercetine and myricetine(Khan,2012). ln addition, Sonchus arvensis potential for an anti-inflammatory because containedsteroids and flavonoids (Lumbanraja, 2009).

Base of chemotaxonomic theory, that a plant has related taxonomically i.e has a family similarlywould produce compounds of the same type and also the possibility of having a similar activity (Kanget al-,2013). Plucea iridica and Sonchus arvensis have a same family, is Asteraceae. lt has been provi:nby Batari Q0O7) that Plucea indica contains flavonoids myricetine, quercetine, and kaempferole.Probably, the Plucea indica has pharmacological activity as well as Sonchus arvensis. Based on this, thisstudy was conducted to explore the activity of pluchea indica L.leaf extracted ethanol on reducing uricacid level in hyperuricemia mice.

MATERIALS

Plant materials : Fresh Plucea indica leaves were obtaided from Materia Medica, Batu, lndonesiaAnimals : Male balb/c mice with body weight of 20-30 g were used in the study.Potassium oxonate : Potassium oxonate was obtained from Sigma, USA; for induction of gout in mice.Allopurinol : Allopurinol is Allopurinol tablet was obtained from Kimia Farma, lndonesia, for positivegroup control.

METHODS

Preparation of plant extract : Around 300 g of Plucea indica leaf powder was macerated withethanol 96% for overnight. From this, a number of 23,4 g crude extract was obtained.Determination of flavonoid content of plant extract : Some extracts reconstituted with 2mtmethanol that had been heated in a test tube. Then added a few drops of concentrated HCI and 0.2gof Mg powder. The positive results containing flavonoids showed arising orange for 3 minutes.Preparation of dosage form : Ethanol extract of Pluces indica leaf dispersed in water using the sameamount of CMC Na (0,5%). For control, the vehicle was prepared through dissolving the same amountof CMC-Na in water.

Design of experimental study for anti-hiperuricemia activity : The anti-hyperuricemia activity ofPlucea indica leaf extracted ethanol was evaluated in male balb/c mice. Twentylfir" male balb/c micedivided into five groups, each comprised 5 mice. Two groups served as control (one positive control andthe other was gouty control), where mice received no extracts. The other three groups were the testgroups. At the start of experiment, the blood of all mice were withdrawn from tail vein to determinationof uric acid healty mice, and then they were injected intraperitoneally with potassium oxcnate (7mgt2Ogbody weight) for induction of gout [Yonetani et al., 1987]. An hour later, mice of different test gioupswere given different oral dose (4mgl20g mice body weight , 4mgl20g mice body weight, 64mg/20g micebody weight) through an esophagus tube of Plucea indica leaf extracted ethanol. Mice of the goutycontrol groups were given only the vehicle and positive control groups were given allopurinol 0,09mg/

142 EitatdffrtttrI{ The I ntern ati o na I Co nf e rence Ph a rmace utica I Ca re

20g mice body weight, intra-peritonial). After t h from treatment, blood samples were withdrawn from

tail vein for the determination of uric acid using Easy Touch@ Uric Acid Meter.

statistical analysis : The results obtained were expressed as the mean+sE. Test groups were compared

with control gouty group and positive group. The significance of values was analysed by One way

ANOVA by p>0.05.

RESUUTS AND DISCUSSION" Before treatment uric acid serum level of the mice were determined. ln this study, uric acid levels

of normal mice were in the range of 1.1 - 1.6mg/dL. After induction with potassium oxonate the uric

acid levels of the mice was increase roughly, in which all mice have uric acid levels above 1.6m9/dl,

on average of 3.5 t O.8mg/dL. lt can be said, that all experimental mice are in hyperuricemia condition.

One hours after treatment with Plucea indica leaf extracted ethanol the uric acid level of hyperuricemia

mice were determined and the result was presented in Table 1'

Table 1. Uric Acid Levels of Hyperurisemia Mice

Treatment GrouPHyperuricemia AlloPurino

(ms/dL)

0.9m9/209BW(ms/dL)

Extra ct dose

of 4mgl20gBW(mg/dL)

Extract dose

of 16mgl20gBW(mg/dL)

Extract dose

of 64mgl20gBW(mg/dL)

I

ll

tlr

IV

V

3.0

3.9

2.92.3

5.1

3.5

3.4

3.4

2.9

3.0

1.4

1.9

2.0

1.2

1.9

1.3

1.1

1.1

1.4

1.2

3.7

3.5

2.9

4.1

4.2

Mean 3.4sD 1.0

1.2

0.1

3.7

0.5

3.20.3

1.7

0.3

Values significanly differ between treatment and control group, indicate by p<0.05

ln this study we obtain that for positivg control group treated with allopurinol, their uric acid levels

become 1.2*0.3mg/dL. For the varies extract treatment, i.e. dose of 4, 16, and 64 mg/2OgBW, the uric

acid levels obtained are 3.71{.5mg/dL, 3.2f0.3mg/dl and 1J*l,2mgldl, respectively. The result showed

the dose of 64 mgl2og BW produced the highest reduction in uric acid level in plasma of hyperuricemia

mice, while the doses of 4mgt20g BW and l6mgDag BW were not reducing the uric acid level (p<0.05)-

The process of catabolism of purines into uric acid in human is transforming adenosine and

guanosine into uric acid. Adenosine initially converted into inosine by adenosine deaminase. Then the

"nryr" xanthine oxidase catalyzes hypoxanthine and xanthine into uric acid ' ln mammals , except the

higher primates (humans) , uratase (uricase) converts uric acid into allatoin , it is highly soluble in water

(Murray et a1.,2009). potassium oxonat used to increasing uric acid levels in the blood by inhibiting

the activity of the enzyme uricase, from the experiments on mice, the levels of uric acid in the blood

increased by 159% of control after administration of potassium oxonate(Watanabe et al., 2006)'

Allopurinol was applied in this study as a positive control, since this medicine is a cure for

hyperuricemia case. ln low dose this compound has an ability to inhibit the formation of xanthine

oxidase enzyme.

Uric Acid lnhibition Activity of Pluchei lndica... 143

coNcLUstoNsThis study investigates the application of natural plant, Plucea indica as a cure of hyperuricemia

on experimental mouse. The mouse was induced to become hyperuricemia by inducing of potassiumoxonate. ln this study, the result showed that only the dose of Plucea indica leaf extracted ethanol at64mgl20g BW mouse decrease uric acid level significanly. lt is so high, so it's not effective to developas cure for hyperuricemia.

REFERENCE

Batari, R., (2007) ldentifikasi Senyawa Flavonoid pada Sayuran tndigenous Jawa Barat, Skripsi Bogor:Skripsi lnstitut Pertanian Bogor.

Berry, C. E. and Hare, J. M. 2004. Xanthine oxidoreductase and cardiovascular disease: The molecularmechanisms and pathophysiological implications. The Journal of Physiology 555 (3): 589-606.

cos, P., Ying, L., calomme, M., Hu, JP., cimanga, K., poel, BV., pieters, 1., vlietinck, AJ., Berghe, DV.1998. Structure-Activity Belationship and Classification of Flavonoids as lnhibitorsof XanthineOxidase and Superoxide Scavengers, J. Nat. Prod, 61;71-76.

Gonzllez, A. G., Bazzocchi, l. L., Moujir; 1., Ravelo, A.G., Correa, M. D. and Gupta, M. p. I995. Xanthineoxidase inhibitory activity of some Panamanian plants f rom celastraceae and lamiace ae. The Journalof Ethnopharmacology 46 (1): 25-29.

Kang, F.N., Lifongo,L.L., Mbaze, L.M., Ekwelle, N., owono, L.c., Magnassan, E., Judson, p.N., sippl, w., Efange5.M., 2013. Cameroonian medicinal plants: a bioactivity versus ethnobotanical survey ard dremotaxenomic classification, Biomed Central Complementary and alternative medicine., nol- 't1 tp-147, pp-l-l&

Katzung, BG., 2007. Basic and clinical Pharmacology. 1Oth ed. san Fransisco USAKersley GD. A short history of gout. ln: Gutman AB, Fessel WJ, Hall AB et al, editors. GouE a clinical

comprehensive. New York: Medcom, lnc; '1971:8-13.

Khan, RA., 2012. Evaluation of Flavonoids and Diverse Antioxidant Activities of Sonchus arvensis.Chemistry central journal, 6:126

Koirewoa, YA,. Fatimawali, Wiyono, Wl., 2012. lsolasi dan ldentifikasi Senyawa Flavonoid dalam DaunBeluntas (Pluchea indica L.). Sulawesi Utara:Universitas Sam Ratulangi Manado.

Kong, L. D., Zhang, Y., Pan, X., Tan, R. X. and Cheng, C. H. K. 2000. lnhibition of xanthine oxidase byliquiritigenin and isoliquiritigenin isolated from Sinofranchetia chinensi. Cellular and MolecularLife Sciences 57: 500-505

Lumbanraja, LB., 2009. Skrining fitokimia dan Uji Efek Antiinflamasi Ekstrak Etanol Daun Tempuyung(Sonchus arvensis L.) Terhadap Radang pada Tikus. Skripsi Fakultas farmasi Universitas Sumatera Utara.

Murray RK, Granner DK, Mayes PA, Rodwel VW.,2009. Harper lllustrated Biochemistry 27th ed, LangeMedical Books/McG raw-H i I I

Nagao, A., Seki, M., Kobayashi, H., 1999. lnhibition of Xanthine Oxidase by FlavonoidsOwen, P. L. and Johns, T. 1999. Xanthine oxidase inhibitory activity of northeastern North American

plant remedies used for gout. Journal of Ethnopharmacology 64: 149-6A

Pacher P., Nivorozhkin, A. and 5zab6, C. 2006. Therapeutic effects of xanthine oxidase inhibitors:Renaissancehalf a century after the discovery of allopurinol. Pharmacology Reviews 58 (l): S7-114.

Ramallo, l. A.. Zacchino, S. A. and Furlan, R. L. E. 2006. A rapid TLC autographic method for the detectionof xanthine oxidase inhibitors and superoxide scavengers. Phytochemical Analysis 17 (1):15-19.

Umpie'rrez, A., Cuesta-Herranz, J., De Las Heras, M., Lluch-Bernal, M., Figueredo, E. and Sastre, J. 199S.Successful desensitization of a fixed drug eruption caused by allopurinol. Journal of Allergy andClinical lmmunology 101: 286-287

wallach, 5. L. 1998. The side effects of allopurinol. Hospital Practice 33:22Yonetani Y., lwaki K., Ogawa Y., Decreasing effect of allantoxanamide, a hypuricemic agent on renal

functions in rats. Jpn. J. Pharmacol., 1987, 45,37-43.

ffiUNIVERSITAS MUHAMMADIYAH MALANG

FAKULTAS ILMU KESEHATANProgram Studi D3 & Sl Keperawatan, Profesi Ners dan SI Farmasi

Kampus II : JL. Bendungan Sutami No. 188-A Tlp. (03a1) 552443 - Pst (144-145)Fax. (0341) 582060, Malang 65145

1.

2.

J.

3.

4.

Pejabat yang memberi tugas

Nama yang diberi tugas

Jabatan

Alamat / Kedudukan

Yang bersangkutan diberi tugas

Tugas tersebut dilaksanakan pada

Keterangan lain

SURAT TUGASNo. E.2. e/ 364 qtrr'xEs-uMMllv/20

1 4

Dekan Fakultas Ilmu Kesehatan

Universitas Muhammadiyah Malang

Dra. Uswatun Chasanah,Apt.,M.Kes

Dosen Program Studi Farmasi Fakultas llmu

Kesehatan Universitas Muhammadiyah Malang

Di Malang

Sebagai Oral Presenter pada ac&ra

International Conference Pharmaceutical Care

"New Development of Pharmaceutical Care in

P harmaco genet ic and P harmacogenomic

Appr o ach", di Universitas Muhammadiyah

Malang

19 April2014

Tunaikan tugas dengan penuh tanggung jawab

sebagai amanah

5.

6.

07 AprilItas Kesehatan

i'r!.

L

.Kep.,Sp.Kom

Tembusan:1. Pembantu Rektor I (pemberitahuan)

2. PD I, II & III FIKES- UMM3. Yang bersangkutan4. Arsip

ffiUNIVERSITAS MUHAMMADIYAH MALANG

FAKULTAS ILMU KESEHATANProgram Studi D3 & 51 Keperawatan, Profesi Ners dan Sl Farmasi

Kampus II : JL. Bendungan Sutami No. 188-A Tlp. (03a1) 552443 - Pst (144-145)Fax. (034I) 582060, Malang 65145

1.

2.

J.

3.

4.

Pejabat yang memberi tugas

Nama yang diberi tugas

Jabatan

Alamat / Kedudukan

Yang bersangkutan diberi tugas

Tugas tersebut dilaksanakan pada

Keterangan lain

SURAT TUGASNo. E.2. ei 364 qtrr'.KE S-UMIWIV/20 1 4

Dekan Fakultas llmu Kesehatan

Universitas Muhammadiyah Malang

Dra. Uswatun Chasanah,Apt.,M.Kes

Dosen Program Studi Farmasi Fakultas Itnu

Kesehatan Universitas Muhammadiyah Malang

Di Malang

Sebagai Oral Presenter pada acara

Intemational Conference Pharmaceutical Care

"New Development of Pharmaceutical Care in

P hormaco genet ic and P harmacogenomic

Approach", di Universitas Muhammadiyah

Malang

19 April2014

Tunaikan tugas dengan penuh tanggungjawab

sebagai amanah

5.

6.

07 April 2Al4Kesehatan

L

.Kep.,Sp.Kom

Tembusan:1. Pembantu Rektor I (pemberitahuan)

2. PD I, II & III FIKES. UMM3. Yang bersangkutan4. Arsip