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Biochinie 91(2000)1339-1341

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Short coIImunication

Alpha― glucosidase inhibitolv effect of anti― diabetic lnetal ions

and their complexes Yutaka Yoshikawa a,*,Ryoko Hirata a,Hiroyuki Yasuia,Hiromu Sakuraib aDttarmmrOfA,alylcal and ttofEorganた bFactlity ofPharmace2rcal sclenに

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ARTICLE INFO

ABSTRACT

■瀬cre

CI)efFect of metalions alnd their complexes wttch showed the We found alpha― glucosidase inhibitory(α― lll.」 h blood g:ucose lowe」 ng efFect in diabetlc lnodel aninnals.Tlle Cu(II)iOn and its complexes showed strong α― CI actlvity『 eater than clinically used acarbose in in vitro studies.Furdlemore, in in vivo experilnents,α ‐ GI actlon was newly discovered in llormal ddy■ lice.These results suggested that one of action mechanisllls of the and diabetic metal ions and complexes is related to the曖 ―CI ettects.

Listoリ

Received 13 February 2009 Acceptcd 5 June 2009 Available online 16 June 2009

Kツ ″OrdS:

0 2009 Elsevier Masson SAS.All Hghts reserved.

Alld diabejc nledicines

Metalions Metal complα es Alpha‐ 」ucosidase

1. Introduo慮 on

2. Methods

Diabetesmellitus(DM)is a metabolic disorder charactenzed bya congenital(type l DM)or aCquired(type 2 DM)inability to trans― port glucose from the bloodstream into cells.Since 2006,inJapan,

2.1. Ma“ riars

approximately 18 mllllon people have suffered from type l and type

2DM.Espedally type 2 DM is rapidly growing in several count五 es [1].ItiS believed thatl■ ost subieCtS developing type 2 diabetespass

alld allittals

ZnS04 CuS04.VOS04,pic01inic acid(pa),α ―glucosidasc sucrose, acarbose,and a Glucose Cll― Test lcit were purchased from lVako Pure Chelnical industries(OSaka Japan). Zn(pa)2, Cu(pa)2, and VO(pa)2 COmplexes were prepared as reported in Refs_12,8].The

through a phase of ilnpaired glucose tolerance. VVe have alsc

ddy mice were obtained from SIIIMIZU laboratory Supplies Co.,Ltd.

Previously repOrted tllat Zn,Cu,and VO complexes exhibit insull― nclmmeticactivity and anti― typel DM and― type 2 DM effects[2,31.

(Kyoto,」 apan).The animalstudy was approved by the Experlmental Animal Research Conlnuttee at Kyoto Pharmaceutical Un市 ersity

Recen■ らWe have proposed the insulinonumetic mechanism in rat adipo呼 tes[41.α ― GIs,a type ofOral anti‐ diabedc medicine,are used not only as nledicines but also as dietaw supplements 15].■ he

(KPU)and Was perfomed according to the Guidelines for Animal Expenmentation at KPU.

c‐ glucOsidase

22.In νねっα―α

in the smallintestille is involved in sugar absorption.

in,libito7 effed

Thus,α ― GIs would linlit tlle absotttiOn ofdietaw Carbohydrates and

in tum suppress postprandial hyperglb7cemia.■ ley are promising drug candidates in tlle treatment and Preventton of DM[61.■ lere

lVe ilnproved upOnthe rnethod ofDahlqvist and analyzed the α―

act市 ity;howeve■ studies on the relationship beれ veen inetals and

ml oftest compound(inal COncentration 300,30,10,3 and 03劇 M)in HEPES bufFer O,15M (pH 6.3)and O.l nJ ofalpha― glucosidase lSac“ aromyc“ sp.)5 units′ mlin HEPES burer

glucosidase action are quite limdted 171.ThuS,We examined the α‐

O.015 M were addedto O■ ml oFsubstrates(0■

inhibiton′ effects of anti― diabetic metal ions and their colllplexes

buffer o■

are severalrepolts on the relationship be“

agaillst

veenlnetals and ette

glucosidase. α…

Gl efFect i91.In briel o■

M suCOSein HEPES

5M)and then incubated at 37° C for 60 min.AFter incu―

bationi each reac● on was stopped in boiling water and each glucose

value was detennined by glucOse《

]1‐ test

Wako(TOkyo Japlln).For

companson,acarbose were added加 stead of test cornpOundζ .The キConespOndlng autllo■ Tel:+81755954630:fax:+81755954753_ 3mafI● ddressf yuraka@mLkyoto‐ phtacjp(V Yoshikawa) 0300‐ 9084′ S― see Flont mattcr 0 2009 Elsevier Masson SAS_A‖ doi:1010161」 .biOChi 2009_06005

ights resclvcd

"GI eFFect was calculated by the following fomula: α―GI efFect(%)=I(Ac― As)/AclxllXl

y yo壺 波c17c et a1/3bCi飾 た9r r20119,I1339-1341

ＯＦこ¨ ︵ ｏ ｏコ●０００ョ一 コでヽ 〇 〇〇０” ｍ ， .30

0

30

60

90

120

‐ 30

Time(min)

0

30

60

90

120

■me{min)

町 1.EfFect ofCuS04(0),Cu(pa12(□ )aCarbose〈 ◇),and vehに le(COntroL△ )on the postprandね l b100d glucose levelin sucmse(A)or gluCOSe(B),10aded nonllal ddy mice.Data are expressed as the mean■ 5D for準 5 mice.

32.■ lhibiloッ eFe由

rable l Estllnated IC50 Values for tlle alpha―

叫 呻 “

compounds

」ucosidase inhibltory efFect of Cu,Zn,and V0

IC50(画 )

005・・

Zn(pab

128± 154±

5・

VO(pa)2

>1(mM)

Acarbose

747■ 7

0.78■ 0.01‐

470士 201±

・

ll t16・

Cu(pa)2

1.6・ `

・,く 001ν

The Cu lon and its picolinic acid ccmple、 which exhibited the highestin ν,鮨oα ― GI ad巨 宙￨ュ was Selected fortlle ttll vivo expeliment

in which α―GI effects were observed in nomal ddy n■ ice.The elevation in the blood glucose levels in normal ddY mice that were

adlninistered disacchande, sucrose,was signiflcantly suppressed after a singie oral adlninistration of Cu ion and its complex aζ

Sucrose(al M)was uSed as thc substrate le nlean± 5D oF3 expenntents Siglullcance:‐ pく 0001,

Date are represented as」

Of metari。 ぉ 。 m`he blood glacose

raveL in carbollydロ セー loaded mice

'acarbose

compared with the control■ 1lce that were ad■ linistered vehicle (Fig.lA).MoreOVe■ Cu conlpounds had the same efFect as the aCaFbOSe. On the other hand, no changes were observed in the blood`JucoSe levelin inice administered lnOnosacdlaride,31ucose,

Ac:production glucose concenttadon of cOntrol(alpha―

gluco―

sidase and substrate)

As: production glucose concentration of the subiect(alpha_ glucosidase,substrate and test compounds).

2ユ rn vivo α_Gr

illれ

ibi10ry qttCt

after Cu iOn and its complex or velucle was adl■ inistered(Fig.lB).

These results suggested tllat Cu lon and Cu(pab inhibited the alpha― glucosidase action in the epithelium of the sman intestine and reduced disacchaHde diges● o■ In conclus,on,we observed the,It vltro and,■ ν:ν οinhibitory ― e■ ects ofillsulinol■ umetic metalions on α glucOsidase acti宙 ty ln f■ .

vitio expe貢 ments,cu ion alld Cu(pa)2 StrOllgly inhibited l← gluco¨

Male 6 week old ddy n亘 ce in eadl anilnal expe亘 ment were fed l labOratory diet and glven vvater ad libi檀 ビn.They were lloused _n an air‐ conditicned room at a temperature of 23± 1 ° C and a humidity of 60%± 10%,M五 th lights on from 8:00 to 20:00.Each mice was unfed for 12 h before a single oral administra●

on ofthe sample by gavage. At 30 H通 n after the sample administration (0.24 mmOl′ kg BllV),2gノ kgいV substrate(glucOse and sucrose)

sidase activity and their erects were apprccmately 1000 tilnes higher than that of clinically used acarbose.On tlle other hand,the inhibitory modes ofyeast alpha― glucosidase by Cu lon and Cu(pa)2

were studied. Iineweaver― Burk p10ts sllowed that Cu ion and Cu(Pa)2 aCted as a non― competitive inhibitor against yeast alpha―

glucosidase(data nOt showll).Furtllet we evaluated the change in

solution was adnlinistered tO dle lllice. Blood samples were

the blood glucose levels in■ orlllal ddy nlice.Cu ion and cu(pa)2 suppressed the elevation in the blood glucose levels in■ orlnal ddy

obtained witl■ out anesthesia ttOn■

mice when dley were admillistered disaccha五 de(sucrOSe).lhese

the tail vein at O,15,30.60,90,

and 120 Πlin after the substrllte adnunistratlon, and the blood

ゴuCOSe Lvels were measured wtth a Glucocard(た kray Co.,Kyoto,

results wlll be useful when they are clinicallyapplied to DM therapy ln the mture.

Japan).

3_ Rttults and`題

s― sion

AclmowledgelEllent lllis research was Partially supported by Grallt‐

g.7.Inhibitory■

8LcFs ofme女 J iOns ο,α glucosid“ e acttfヶ illゼ ■νf″Os如 ″ The inhibitow efFects of acarbose(pOSitive control),Zn 10rL Cu

lon,Vo ion1 2n(pab,Cu(pa)2,and VO(pa)20n α― glucosidase activity were concentratton dependent.From the result,the apparent IC50 values of the test compOunds with respect to the glucosidase α‐ activity were estimated(Table l).In cOmparisOn亜 th acaぬ ose,all test cOmpounds except for Vo(pa)2 ShOWed strOng GI erect. α¨

Scientists(B),17790042,2005-2006■

in―Aid

for Young

om the Ministry of Eductl‐

tion,Sdence,Sports and Cuiture to,rι

References

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盤讐 群瑚凛l憮_‐sT[潔厨 環

d tteatnl。

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1と

121 N、 Yasumatsu,Y Yoshlkawa.Y Adachi,H Sakur」 ,AlJdiabetic coppe