NATURAL QUINOLONE ANTIMICROBIAL AGENTS FROMANTIMICROBIAL ... · NATURAL QUINOLONE ANTIMICROBIAL AGENTS FROMANTIMICROBIAL AGENTS FROM MALAYSIAN RUTACEOUS SPECIES Natural Medicinal

NATURAL QUINOLONE NATURAL QUINOLONE ANTIMICROBIAL AGENTS FROMANTIMICROBIAL AGENTS FROMANTIMICROBIAL AGENTS FROM ANTIMICROBIAL AGENTS FROM

MALAYSIAN RUTACEOUS SPECIESMALAYSIAN RUTACEOUS SPECIES

Natural Medicinal Products CentreKulliyyah of Pharmacy

International Islamic University Malaysia

INTRODUCTIONINTRODUCTIONINTRODUCTIONINTRODUCTION



QUINOLINE ALKALOIDS OF QUINOLINE ALKALOIDS OF

• Quinoline alkaloids of the Rutaceae plant family

THE RUTACEAETHE RUTACEAE• Quinoline alkaloids of the Rutaceae plant family

- an anthranilic acid-derived alkaloids.

• Typical and unique chemical constituents of the Rutaceae plant family (Grundon,1988).

• Main classes :Simple Quinolines 2(1H)-quinolinonesp Q ( ) q

4(1H)-quinolinones dihydrofuroquinolinesfuroquinolines pyranoquinolinesfuroquinolines pyranoquinolines



• A considerable number of antimicrobial active• A considerable number of antimicrobial activequinoline alkaloids have been isolated from variousspecies of Rutaceae.

• The 4-oxygenated quinolone and 2-substituted 4-quinolone alkaloids which have similar basic structureto that of 4 quinolone antimicrobial agents have beento that of 4-quinolone antimicrobial agents have beenproved to be candidates for further study as potentialantimicrobial agents.



Antimicrobial active quinoline alkaloids from Rutaceae speciesq p

OCH3

DubamineHapplophyllum sp

OCH3

OCH

GalipineG officinalis

N

O

OHapplophyllum spGrundon, 1988 N

OCH3

OCH3

G. officinalisHoughton et al., 1999

OO

N

CH3

(CH2)m (CH2)nCH3

Evocarpine, m = 7, n = 3 Evodia rutaecarpaRho et al., 1999 N

CH3

IsodictamnineDictamnus albusTowers et al., 1981

Quinoline alkaloids with DNA-binding activity (Hanawa et al., 2004)

N O

OCH3

HO N OH CO

OCH3

N OH CO

OCH3

H3COO

N O

OCH3

HO N OH3CO

OCH3

N OH3CONH

O

Haplopine Skimmianine Kokusaginine Haplopine



SELECTED RUTACEOUS SPECIESSELECTED RUTACEOUS SPECIES(1) Selected Rutaceae

species(1) Glycosmis pentaphylla(1) Glycosmis pentaphylla

(Retz.) DC./Nerapan

(2) Lunasia amara Blanco

(3) Ruta angustifolia (L.)Pers./ Aruda/Garuda

6



Quinoline alkaloids of Glycosmis pentaphyllaAlkaloid Occurrence References2-(1H)-QuinolinonesGlycolone Leaves

Root barksBhattacharyya and Chowdhury, 1985Kumar et al., 1986

GlycophyloneGlycosolone

SeedsLeavesRoot barks

Bhattacharyya and Chowdhury, 1984Bhattacharyya and Chowdhury, 1985Das and Chowdhury, 1978

Homoglycosolone Root barks Kumar et al., 1986FuroquinolinesDictamnine Mester, 1973γ-FagarineKokusaginineSkimmianine

StemsLeavesStems

Mester, 1973Ito et al., 1999Chaterjee and Majumdar, 1954Ito et al 1999Stems Ito et al., 1999



Quinoline alkaloids of Lunasia amara

Simple Quinolines4 Methoxy 2 phenyl quinoline

FuroquinolinesKokusagine

(Goodwin et al., 1959c; Hart and Price, 1966; Mester, 1983)

4-Methoxy-2-phenyl-quinolineGraveolinine

KokusagineSkimmianine

2(1H)-QuinolinonesL idi

DihydrofuroquinolinesL iLunacridine

LunidonineLunidine( ) H d l idi

LunacrineLunine(-)-O-Methylluninium ionLunasine(+)-Hydroxylunacridine

Hydroxylunidine

Lunasine(-)-HydroxylunacrineHydroxylunine

4(1H)-Quinolinones Pyranoquinolines( )1-Methyl-2-phenyl-4-quinolone5-Hydroxy-1-methyl-2-phenyl-4-quinoloneEduleineLunasia I

y q(-)-Lunacrinol

Lunasia ILunamarine



Quinoline alkaloids of Ruta angustifoliaQuinoline alkaloids of Ruta angustifolia

Alkaloid Occurrence References4-(1H)-QuinolinoneGraveoline Aerial parts

RootsWh l l

Del Castillo et al., 1984Ulubelen and Terem, 1988V d d L k 1968Whole plants Vasudevan and Luckner, 1968

FuroquinolinesDictamnine Roots Vasudevan and Luckner, 1968Dictamnineγ-Fagarine

Roots Vasudevan and Luckner, 1968Kong et al., 1984



Antimicrobial activity of quinolone antimicrobial agentsy q g

345

6

O OCOOH

OCOOHF

OF COOH

NH

2

36

7

8 1N

C2H5

H3C

NHN N NH3CN N

F

CH2F

H2C

4-quinolone Nalidixic acid Ciprofloxacin Fleroxacin4 quinolone Nalidixic acid Ciprofloxacin Fleroxacin

• Nucleic acid-targeted drugs. • The functional target is the enzyme DNA gyrase that exists only in bacteria (Shen, 1993; Kreuzer, 1998). • Inhibit the replication and transcription activities of bacterial cell’s genetic code (Hooper and Wolfsonbacterial cell s genetic code (Hooper and Wolfson, 1993).



METHODSMETHODSMETHODSMETHODS



BIOASSAYBIOASSAY--GUIDED ISOLATION OF GUIDED ISOLATION OF ANTIMICROBIAL ACTIVEANTIMICROBIAL ACTIVE ALKALOIDSALKALOIDSANTIMICROBIAL ACTIVE ANTIMICROBIAL ACTIVE ALKALOIDSALKALOIDS

Extraction Crude extracts

Bioactivity screeningInactive extracts

Active extractsFractionation

TLC analysis

F1F2F2F3F4F5

Bioautographicscreening

Active fractions

Isolation &Purification

Antimicrobial active

alkaloidsF5

Inactive fractions Spectroscopic

identificationNatural Medicinal Products Centre

Kulliyyah of PharmacyInternational Islamic University Malaysia

ANTIMICROBIAL STUDYANTIMICROBIAL STUDY

(b) Chequerboard AssayA ti i bi l C bi ti Eff t

(a) Broth Microdilution Assay for - MIC & MBC

- Antimicrobial Combination Effects

*



RESULTS & OBSERVATIONSRESULTS & OBSERVATIONSRESULTS & OBSERVATIONSRESULTS & OBSERVATIONS



AAntimicrobialntimicrobial active alkaloid from active alkaloid from R. R. angustifoliaangustifoliaC. albicans

O

45

NO

1

2

364a

7

8

8a

1'5'

6'

AAntimicrobialntimicrobial active alkaloid ofactive alkaloid of GG pentaphyllapentaphylla

S. aureusO

CH3 2'

3'4'

O

45

S. aureusAAntimicrobialntimicrobial active alkaloid of active alkaloid of G. G. pentaphyllapentaphylla

N

N

CH21

2

34

6

7

8 1"

5"6"4a

8a

4"1'

CH3

1

2" 3"

E. coli ArborineNatural Medicinal Products Centre


Antimicrobial Antimicrobial activity of alkaloiactivity of alkaloidsds

Microorganism MIC/MBC/MFC of Rutaceous alkaloidsLunidonine Graveoline ArborineLunidonine Graveoline Arborine

E. coli ATCC 35213 - 250/250 125/125S. aureus ATCC 25923 125/125 125/250 250/250C. albicans ATCC 90028 - 500/500 NR



Synergism antimicrobial Synergism antimicrobial effectseffects

CombinationMIC (μg/ml)

FIC FICICombinedeffect

alone combined

E. coli ATCC 35213

alone combined

Ampicillin 3.91 0.98 0.25 0.38 Synergy

Arborine 125 15.63 0.13


Graveoline 250 31.25 0.13


Quinoline 250 31.25 0.13


Odds, 2003

p y gy

Quinaldine 250 31.25 0.13

Odds, 2003

FICI <0.5 – Synergy

0.5<FICI>4.0 – No interaction

FICI>4.0 - AntagonismNatural Medicinal Products Centre


g

CONCLUSIONSCONCLUSIONS• 3 alkaloids which were firstly detected to have

antimicrobial properties have been isolated from Lantimicrobial properties have been isolated from L.amara, R. angustifolia and G. pentaphylla, identifiedas lunidonine, graveoline and arborine, respectively., g , p y

• The alkaloids especially graveoline and arborine,which produce effect against Gram-negative bacteriawhich produce effect against Gram-negative bacteriamay be suitable candidates for use in combinationantimicrobial therapy.py

INTERNATIONAL ISLAMIC UNIVERSITY MALAYSIANatural Medicinal Products Centre


ACKNOWLEDGEMENTACKNOWLEDGEMENTACKNOWLEDGEMENTACKNOWLEDGEMENTPROF DR MUHAMAD BIN ZAKARIAPROF. DR MUHAMAD BIN ZAKARIA

UNIVERSITY OF MALAYAINTERNATIONAL ISLAMIC UNIVERSITYINTERNATIONAL ISLAMIC UNIVERSITY

MALAYSIAMOHE (FRGS GRANT)MOHE (FRGS GRANT)



THANK YOUTHANK YOUTHANK YOUTHANK YOU



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NATURAL QUINOLONE ANTIMICROBIAL AGENTS FROMANTIMICROBIAL ... · NATURAL QUINOLONE ANTIMICROBIAL AGENTS FROMANTIMICROBIAL AGENTS FROM MALAYSIAN RUTACEOUS SPECIES Natural Medicinal