PROPERTIES AND BIOCOMPATIBILITY OF ZINC OXIDE-OZONATED EUGENOL AND CONVENTIONAL ZINC OXIDE EUGENOL AS AN ENDODONTIC SEALER
– AN INVITRO STUDY
A Diss ertation sub mitted
in parti al fulfil men t of th e requirements for the degree of
MASTE R O F DENT AL SURGE RY
BRANCH – IV
CONSE RVATI VE DENTISTRY AND E NDO DONTICS
THE TAMIL NADU DR.M.G.R. ME DICAL UNI VERS ITY CHENNAI- 600032
2014 – 2017
DE PART MENT O F CONSE RVATI VE DENTISTRY AND ENDO DO NTICS
CERTI FICATE
This is t o certif y t hat DR.C.RAVI VARMAN, Post Graduat e student (2014 -2017) in the Department of Conservati ve Dentistr y and Endodontics, Adhi parasakthi Dent al College and Hospi tal , Melm aruvathur – 603319, has done this dissertation titled
“Comparative evaluati on of the physical prop erti es and biocompatibili ty of zin c oxid e -ozonated eugenol and con ven tional zin c oxide eu gen ol as an end odon tic sealer ” – An in vi tro s tudy under our direct gui dance an d supervi si on in part ial ful film ent of the regul ations l ai d down b y the Tami lnadu Dr.M.G.R Medical Univers it y,
Chennai – 600032 for MDS., (Branch-IV) CONSE RVATI VE
DENTIST RY AND ENDO DO NTICS degree ex amination.
Dr.S. Thil lainayagam MDS ., Principal
Guide
Dr.S.Thill ainayagam MDS ., Professor & Head
Co-Guide
Dr.A.Jayas enthi l MDS., Reader
I am ext rem el y grat eful to Dr.S.Th illai nayagam MDS., Guide, Professor and Head, Departm ent of Conservative Dentistr y and Endodontics, Adhi parasakthi Dent al College and Hospi tal , Melm aruvathur. Words cannot expres s m y gratitude for his qui et confidence in m y abilit y to do the stud y, his willingness to help clear the st umbli ng bl ocks along the wa y and his trem endous pati ence t ill the end o f t he stud y.
It i s m y dut y t o express m y thanks to m y Co -Guide Dr.A.Jayas enthi l MDS., R eader for his expert gui dance and moral support during t he compl eti on of thi s stud y. I consider m ys el f privileged, to have studi ed, worked and compl et ed m y dissert atio n under them i n t he departm ent.
M y sincere thanks t o our C orrespondent Dr.T .Ramesh MD., for his vital encouragem ent and support.
I am extrem el y thankful to m y t eachers Dr.D.S .Din esh MDS., Professor, Dr.N.Bh arath MDS ., R eader, Dr.S.Karthikeyan MDS., Senior lect urer, for t hei r val uabl e s uggestions , constant encouragem ent and tim el y help rendered throughout t his stud y.
I am extrem el y grat eful to Dr.Mohanku mar, Lab t esti ng, Ci pet , Guind y and Dr.K.T hirugnan asamband am, R esearch direct or, PC BS ,
I thank Dr.S.Pandian, Madras Vet erinary C oll ege and hospi tal, Veper y, Chennai, for helping m e with the stati sti cs i n the study.
I also wish to th ank m y post graduate col leagues and m y j uni ors for thei r hel p and support.
I owe m y grati tude to m y fat her (l at e) Dr.T.Chand ramohan MD., and m y m other Dr.R.Man oranji tham MS., who stood beside m e duri ng m y hard tim e and s acrifi ced s o much to m ake m e what I am toda y. A speci al thanks to m y wi fe Dr.R.J .Shali ni and m y son m ast er R.Dis haanchand for thei r s upport and encouragem ent t hroughout m y career.
My acknowledgement wouldn’t b e complete without mentioning m y t eaching and non -t eaching st affs , Offi ce s taffs, and l ibrar y st affs for being m y st rengt h, b y gi ving m e cont inual s upport all the time.
Before and aft er ever ything I t hank the Almi ght y for guidi ng m e through the ups and d owns in m y l ife. I humbl y bow before Hi m for hi s bles sings.
Dr.C.Ravivarman Post graduate student
TIT LE OF THE DIS SERTATION “Comparative evaluation of the physical prop erti es and biocompatibili ty of zinc oxid e - ozon ated eugenol and conven tional zinc oxide eu gen ol as an endod onti c seal er ” – An invitro stud y
P LACE OF THE S TUDY Adhi paras akt hi Dent al C oll ege and Hospital, M elm aruvathur – 603319
DUR AT ION OF THE COURSE 3 years
NAM E OF THE GUIDE Dr.S.Thill aina yagam , MDS .,
NAME OF C O-GU ID E Dr.A.Ja yas enthil , M DS.,
I hereb y decl are that no part of the di ss ert ati on will be uti li zed for gai ning financial assist ance or an y promoti on wit hout obt aining prior permi ssion of the Pri nci pal, Adhiparas akthi Dent al C ol lege and Hospital, Mel maruvathur – 603319. In addition, I declare t hat no part of this work wil l be published eit her in print or in el ect ronic medi a without t he gui des who has been acti vel y invol ved i n dis sert ation. The aut hor has the ri ght to res erve for publis h work sol el y with the permiss ion of the Princi pal , Adhi parasakthi Dent al College and Hospital, M elm aruvathur – 603319
Co- G ui de Gui d e & H ead of depart m ent Si gnature of candi dat e
BACKGRO UND:The challenge to t he prognosis of endodonti c therap y lies on the qualit y of root canal fi lling. The zinc oxide eugenol (ZOE) bas ed seal ers were introduced i n Endodonti cs b y Gross man in 1936 and t he y have been used in dent ist r y over a l ong period of ti me. It has been report ed that eugenol rel eased from ZOE is an i rrit ant to the peri api cal tis sues and has c ytot oxic properti es . To overcome t hese disadvant ages , there is a continued res earch for chemical compounds with bett er ph ysi cal properti es and m ore biocom pat ibilit y. Ozone therap y has a long histor y of res earch and clinical appli cation wit h hum ans because of it s int eresting bi ologi cal properti es .
AI M:The aim of the st ud y is to eval uat e and compare the ph ysi cal properti es and bi ocompatibili t y of zinc oxide - ozonat ed eugenol and conventional zinc ox ide eugenol .
MATE RI ALS AND METHODS :Foll owing ph ysi cal properti es of conventional zinc oxide eugenol and zinc oxide-oz onated eugenol were evaluat ed and com pared . [Group I: Zi nc oxide – eugenol; Group II: Zinc oxide - ozonat ed eugenol]. Hardness of the mixed samples was measured using S hore hardness Durom et er t ype A. Sol ubili t y was t est ed as a percent age of t he mass of specimen rem oved from t he distil led wat er compared with the ori gi nal m ass of the speci mens usi ng M ettl er anal yti cal balance. p H of the fresh samples and the set s amples were measured using calibrat ed pH meter aft er predet ermi ned tim e intervals.
Biocompatibi lit y of the t est ed sam pl es was eval uated usi ng the M TT tet razol ium reducti on ass a y.
hardnes s and sol ubil it y of t he t est ed samples were almost similar in both the groups. pH and bi ocompati bilit y of the sam pl es i n Group II where eugenol i s ozonat ed was hi gher when com p ared t o the s ampl es in Group I. The proportional values obt ai ned were st ati sti cal l y anal ys ed b y unpai red t -test.
CONCLUS ION: Wi thin t he limit ati ons of the st ud y, it could be concl uded that o zone wit h i nteresti n g biol ogi cal properti es s eems to be a bett er treatm ent modalit y i n improvi ng the properti es of eugenol.
However further cl i nical t ri als and randomized cont rol studies are requi red i n t his fi eld to us e ozonated eugenol in combi nation with zinc oxide powder to us e it as an endodonti c s ealer i nvi vo.
S.NO TITLE PAGE No.
1. INTRODUC TION 1
2. AIM AND OBJ EC TIVES 5
3. REV IEW OF LITER ATURE 6
4. MATER IA LS AND METHODS 31
5. RESULTS 44
6. DISC USS ION 52
7. SUMM ARY AND C ONC LUS ION 60
8. REFERENCES 63
9. ANNEXUR E 73
Figu re No.
TITLE Page No.
1.
Zinc oxide powder, Eugenol, Gl as s sl ab and st ainl es s st eel spatul a.
39
2.
Stainl es s st eel ring of dim ensi on 1.5mmx10m m.
39
3. Ozonator m achine. 39
4.
Eugenol ozonat ed b y bubbli ng gases through it.
40
5. Shore hardness Duromet er t ype A. 41
6. Calibrat ed di git al pH met er. 41
7.
Sampl es s haped using st ainl es s steel molds .
42
8.
Mettl er AE 260 Delt a range Anal yti cal Bal ance.
42
9. MTT R eagent 43
10. Micro -pl at e R eader 43
11. Biocompatibi lit y Tes ting Group I 43
12. Biocompatibi lit y Tes ting Group II 43
Table No.
TITLE Page No.
1.
Mean hardness values of the test ed samples
45
2. Mean pH values of t he tested s amples 46
3.
Mean solubilit y values of t he t est ed samples
47
4.
Mean biocom patibili t y values of the test ed samples
48
Chart
No. TITLE Page No.
1. Hardness values of test ed sam pl es after
12hrs 49
2. Hardness values of test ed sam pl es after
24hrs 49
3. Mean hardness val ues of the tested s ampl es 50
4. Mean pH values of t he tes ted samples 50
5. Mean s olubilit y values of t he t est ed
samples 51
6. Mean biocom patibili t y values of the test ed
samples 51
ADA : Ameri can dent al associ ation
Al : alumi nium
ANS I : Ameri can nat ional st andards ins titut e ATP : adenosine t riphosphate
COX : c ycloox ygenase
DNA : deox yribose nucl ei c acid DUW Ls : dental uni t wat er li nes IC : inhibit or y concent rat ion IL-β : Int erleukin -1β
IR M : interm edi at e restorat ive mat eri al LDH : lacti c deh ydrogenase
LPS : Li popol ys accharide
MTT : (3 -(4, 5-dim eth yl thi azol -2 - yl )-2, 5 - diphen yl tetrazoli um brom ide)
NaOCl : sodium h ypochl ori te
NF : nuclear factor
O3 : ozone
OSHA : occupati onal safet y and health admini strat ion ZOE : zinc oxide eugenol
PDGF : plat el et derived growth factor PDT : photod ynami c therapy
PRC L : prim ar y root cari ous lesi ons pH : pot enti al of h ydrogen
ppm : parts per million RNA : ribose nucl ei c acid TNF : tumour necrosi s fact or TGF-β : trans forming growth fact or
VEGF : vas cul ar endotheli al growth factor
INTRODUCTION
Success in Endodont ic therap y i s based on diagnosis, t reatm ent planning, knowl edge of toot h anat om y and the traditi onal concepts of debridem ent and obt uration. Schild er describes the fi nal obj ective of endodontic procedure as “the complete obturation of the root canal space”. It is the sealing of complex root canal system from the periodont al bone t hat ens ures t he health of att achm ent apparat us against the breakdown of e ndodonti c ori gi n¹.
Endodontic seal ers are used in t he obturation of root canal s yst em t o achi eve a flui d ti ght seal throughout the canal incl uding the canal i rregulariti es . An i deal root canal sealer s hould provide excell ent seal when s et, shoul d have di m ensi onal stabilit y, bi ocompati bilit y and hel p to res olve peri apical l es ions².
The Zinc Oxide Eugenol (ZOE) bas ed seal ers were introduced in Endodontics b y Gross man in 1936. ZOE seal ers have a histor y of success ful us e over extended peri od of t ime and has both anaestheti c and antimi crobi al properti es³. However t he eugenol rel eas ed from ZOE is report ed to be an irrit ant t o periapi cal tiss ues and sai d to have c yt otoxic properties4. Ever y att empt made t o overcome t hes e disadvant ages and t o replace ZOE com plet el y has fail ed, s o there is continued res earch for chemi cal com pounds with bett er ph ys i cal properti es and biocompati bilit y.
In search of other t herapeuti c m odaliti es, ozone therap y has a long hist or y of research and cl ini cal appl ications with hum ans because of its int eresting biologi cal properti es. The word ozone was fi rst introduced b y S chon bein in 1840 when he s ubj ected ox ygen t o el ect ric discharge and noted the odour of el ectric matt er5. Ozone i s s ai d to have antim icrobial, im mune -m odul at or y, i mmune-stimul at ing, anti - infl amm ator y, anal gesi c and haem ost ati c propert y. Ozonat i on is an energetic proces s that res ults i n an ox ygen m ol ecule being split int o singl et ox ygen, the s ingl et ox ygen then combines with diat om ic ox ygen to form ozone. Three s ys t e ms of generating ozone are: 1.Ultraviolet s yst em 2.C old plasm a s ys tem 3.Corona di scharge s ys tem . The modes of applicati ons of ozone can be in gas eous form , aqueous sol ution or in the form of ozonat ed oils .
Anti mi crobial action: Bei ng a superoxide, ozone i s 5 ti mes more effective than chlorine. It affects onl y the unhealth y cell s such as cancer cells, organi sms like bact eri a, virus es , fungi, parasi tes whi ch have los t their prot ective m echanism and those devoid of anti -oxidants and free radical scavengers.
Anti -inflammatory and Analgesi c action: Ozone hel ps in the s ynthesi s of biol ogi call y acti ve substance such as i nt erl eukins, leukot riene and pros tagl andins t hereb y reduci ng the inflamm ation and pai n.
Anti -cariogeni c action: The strongest nat urall y occurrin g acid produced b y acidogenic bact eri a is p yruvic acid. Ozone has t he abilit y
to decarbox yl at e p yruvi c acid to aceti c acid and sai d t o enhance the rem inerali sati on of t eeth.
Antih ypoxi c action: ozone brings about the ris e of parti al press ure of ox ygen and acts as super ox ygenat or b y bringing ox ygen t o tissues, assi sting in its natural heali ng process6.
In dentist r y, ozonated oil appli ed on herpes l abial is i nfection and osteom yeliti s has demonst rat ed fast er healing rat e. It has been al so used in the t reat ment of avascul ar ost eonecrosis of the j aw. Ozone has strong bact erici dal activi t y against pl aque bi ofi lm and can be us ed to disinfect denture bases from C andida albicans. Zinc oxide -ozonated sesam e oil when us ed as a root filli ng m at eri al has demons trat ed good success rate(93.3%) when compared to ZOE(63.3%)7. In restorative dentist r y, gaseous ozone has been used in the t reatm ent of inci pi ent lesi ons si nce it caus es reduction in the number of mi croorganisms8. In Endodontics, ozonat ed water had nearl y t he s am e ant imicrobi al activit y as 2.5% NaOCl wit h fewer side effects when us ed as an irri gant9.
Ozonator D -400 us es a hi ghl y effi ci ent corona dis charge s ys tem rat ed at 400m g/hr. of const ant ozone out put. It can be us ed to ozonate wat er and oil. Due t o the hi ghl y unst abl e nature of ozone i n aqueous form, we opted use ozone in the form of oils. Eugenol is a member of phen yl propanoids, a yellow oil y liquid extracted es peci all y from cl ove oil ( 80-85%), whi ch can be ozonat ed t o overcome its dis advant ages like irrit ati ng action on peri api cal tissues and c yt ot oxic properties.
Hence, the purpos e of the s tud y is to eval uat e and compare the ph ysi cal properti es and bi ocompatibi lit y of ozonated ZOE and conventional ZOE for being used as an endodonti c s eal er.
AIM AND OBJECTIVES
Ai m of the s tudy:
To evaluat e and com pare the ph ysi cal properti es and biocompati bilit y of zinc oxide -ozonated eugenol and that of conventional zinc ox ide eugenol .
Objectives of the study:
1. To evaluate the ph ysical properti es s uch as hardness , sol ubil it y and pH of zinc oxide - ozonat ed eugenol and conventi onal z inc oxide eugenol.
2. To eval uat e the biocompatibili t y of zinc oxide -ozonat ed eugenol and conventional zinc oxid e eugenol .
3. To com pare and correlate the ph ysi cal properti es and biocompati bilit y of zinc oxide - ozonat ed eugenol and conventional zinc ox ide eugenol .
REVIEW OF LITERATURE
Ro ot cana l t herap y dep e nds o n t he int eg ra lly re lat ed t reat me nt phas es: micro bia l co nt ro l, c le a ning a nd s hap ing, a nd e ffect ive s ea ling o f t he ro ot cana l s yst e m. The succ ess o f t reat me nt dep e nd s o n t he e xecut io n o f t he fina l p ha se i.e. sea ling o f t he ro ot cana l. E ndo do nt ic filling mat er ia ls ma y be co ns idere d like t rue imp la nt s a s t he y t o uch a nd are based in v it a l t issu es o f t he bo d y. T he ma in co mpo ne nt s o f a ro ot filling are: a so lid co re mat er ia l a nd a sea ler. ZOE se a lers ha ve a hist o r y o f s ucc es s fu l u se o ver e xt ended per io d o f t ime. It ha s bo t h a nae st het ic a nd a nt imicro bia l pro pert ie s1 0.
To rabin ejad M (1993)1 1 st at ed t hat sea ler s he lp t o preve nt le akag e, reduce t he p o ss ib ilit y o f r es id ua l ba ct er ia fro m t he c a na l t o invade t he per iap ic a l t is sue s a nd t o reso lve per ia p ic a l le s io ns.
M cM ichen FR et al (2003)1 2 st at ed t hat the ha nd ling pro pert ies o f ro ot ca na l s ea ler s a nd it s c lin ica l be ha vio ur ma y be c har act er ised by la b t est s o n t he ir p h ys ic a l pro pert ies. T he se inc lu de flo w, so lu bilit y, wo rking t ime a nd set t ing t ime. He a lso st at ed t hat t hese mat er ia ls s ho u ld po sse s s ade quat e wo rk ing t ime fo r c lin ic a l ha nd ling a nd t reat me nt pro cedures.
ISO standa rd s: ISO 6876: 200 11 3
Flo w o f e ndo do nt ic se a ler sho u ld ha ve a d ia met er no t le ss t ha n 20 mm.
Wo rk ing t ime o f t he sea ler t o be le ss t ha n 30 min
Sett ing t ime o f t he s ea ler sho u ld be gr eat er t ha n 30 min a nd upt o 72hrs.
Film t hic k ne ss s ho u ld no t be mo re t ha n 50 µ m
D ime ns io na l c ha ng e s ho u ld no t t o exce ed 1.0 % in s hr inkag e o r 0.1% in e xpa ns io n.
So lu b ilit y o f t he se a ler s ha ll no t exceed 3 % o f mas s fra ct io n.
Rad io -o pac it y s ho u ld be eq u iva le nt t o o r no t le ss t ha n 3 mm o f a lu min iu m.
Gros sman st at ed t hat Zinc o xid e eu ge no l se a ler ha ve a hist o r y o f succ ess fu l u se o ver a n e xt e nd ed per io d o f t ime. An ide a l ro ot cana l sea le r s ho u ld po sse ss fo llo w ing pro pert ies 1 4.
1. It sho u ld be t ack y w he n mixed, t o be a ble t o pro vide ad he s io n/ sea ling bet wee n t he ca na l wa lls a nd t he co re o bt urat ing mat er ia l.
2. Flo w a bilit y in it s no n-s et st at e t o pro mo t e lut ing.
3. It sho u ld be no n- irr it at ing to t he per iap ic a l t is su es.
4. It sho u ld po sse s s a her met ic se a ling a bilit y.
5. It sho u ld be rad io p aque.
6. It sho u ld po sse s s min ima l s et t ing s hr inkag e.
7. Sho u ld be no n-st a in ing o n t he de nt in / t oot h st ruct ure.
8. It sho u ld be id ea lly bact er ic ida l o r at lea st co nt ro l t he bact er ia l gro wt h.
9. Sho u ld ha ve lo ng wo rk ing t ime.
10.Inso lu ble in t issue flu ids.
11.It sho u ld be ea s y t o remo ve if ne ce ssar y.
12. It sho u ld be ne it her mut age nic no r carc ino ge nic.
13. It sho u ld no t pro vo ke an immu ne rea ct io n.
Grossman‟s original formula for zinc oxide eugeno l based sealers co nt a ined ZnO, st abylit e res in, bis mut h s ub c ar bo nat e, bar iu m su lp hat e a nd so d iu m bo rat e w it h e uge no l a s t he l iq u id co mpo ne nt .
Roth’s sol sealer (Roth‟s pharmacy, Chicago, in c.) is essent ially t he same as Grossman‟s orig inal formulat ion wit h the subst itut ion o f bis mut h su b- nit rat e fo r bis mut h su b- car bo nat e.
Ricket’s formula market ed a s Kerr‟s pu lp ca na l s ea ler ( S ybro n e ndo / Kerr, Ora nge, C A) ha s lo ng bee n acc ept ed mat er ia l. But it s u se has bee n re st r ict ed fo r st a ining o f t oot h st ruct ure fro m t he s ilver us ed fo r rad io -o pac it y.
Tub lis ea l(S ybro ne ndo / Kerr) is a t wo past e s yst e m co nt a ined in t wo separ at e t ubes de ve lo p ed as no n -st a in ing a lt er nat ive w it h bar iu m su lp hat e fo r rad io pac it y. It ha s t he d isad va nt age o f rap id set t ing t ime.
Wach’s cement(Ro t h int ernat io na l I nc. C hic ago ) co ns ist s o f po wder o f ZnO, bis mut h s u b- io d id e, bis mut h su b- nit rat e, mag nes iu m o xid e a nd ca lc iu m p ho sp hat e. It ha s t he ad va nt age o f s mo o t h co ns ist e nc y a nd t he Ca na da ba ls a m mak e t he s ea ler t ac hy1 4.
Hauman CH et a l (2003)1 5 repo rt ed t hat t here are ma n y fo r mu lat io ns a nd bra nds o f sea ler t hat h a ve z inc o xid e a s pr ima r y ingr ed ie nt , d iffer ing o n ly b y o t her co mpo ne nt s ad d ed t o t he s ea ler. Zinc o xid e
sea le rs a llo w fo r t he add it io n o f ot her c he mic a ls, suc h a s para fo r ma lde h yde, ro s in, C a nad a ba ls a m a ll o f w hic h ma y e nha nc e t he ir pro pert ie s. Endo do nt ic mat er ia ls ca n be bro ad ly cat ego r ize d as t ho se used t o ma int a in pu lp vit a lit y a nd t ho se used in pu lp ca na l t herap y fo r d is infe ct io n o f t he p u lp spa ce ( irr iga nt s a nd int ra -ca na l me d ica me nt s) a nd ro ot cana l filling ( s o lid mat er ia ls a nd s ea ler s).
B io co mp at ibilit y o f t he se e ndo do nt ic mat er ia ls is c haract er ized b y ma n y p ara met ers suc h a s g e no to xic it y, mut age nic it y, c arc ino g e nic it y, c yt o to xic it y, hist o co mp at ibilit y o r mic ro bia l e ffect s. He nce it is impo s s ib le t o bio lo g ica lly c haract er ize t he mat er ia ls b y a s ing le t est met ho d a lo ne a nd t he ir pro pert ies ne ed t o be inve st igat ed by a ser ie s o f var io us in vit ro and in vivo t est s ina st ruc t ured appro ach.
Candei ro et a l (201 2)1 6 repo rt ed t hat no wada ys var io us k ind s o f e ndo do nt ic se a lers are a va ila ble suc h as s ea ler s bas ed o n g la s s io no mer, ca lc iu m hydro xide, silico ne, res in bas ed a nd bio c era mic mat er ia ls.
Lace y S (2 005)1 7 st at ed t hat a s yet no r oot ca na l s ea ler s has a ll t he id ea l pro pert ies a nd mo st le ak w it h t ime, e it her t hro ugh po o r init ia l adapt at io n t o t he ca na l wa lls o r due to solu bilit y a nd d is int egrat io n o f t he sea ler. T he y a lso st at ed t hat su cce ssfu l o bt urat io n is a c hie ved by t he ad hes io n a nd st a bilit y o f t he sea le r to t he c a na l wa lls w hic h, a mo ng st ot her fact o rs, is a ffe ct ed b y it s rheo lo g ic a l pro pert ie s. The y repo rt ed t hat narro wer t ube s pro duced increa sed ve lo c it y, w hic h wa s sig nific a nt fo r a ll sea le rs, a nd reduc ed vo lu met r ic flo w, w h ic h w as
significant for all sealers except Grossman‟s sealer. Reduct ion in powder: liquid rat io o f Grossman‟s significant ly increased flow in narro w t ube s a nd at hig her st ra in rat e.
Eugeno l (4- a c yl-2- met ho xy p he no l) is a n e xt ract o f c lo ve o il w id e ly used in de nt ist r y as a t hera peut ic a ge nt , mo st co mmo nly a s a co mpo ne nt o f z inc o xid e eu ge no l ce me nt a pp lied a s a ba se o r t empo rar y dre ss ing t o dent ine, o r as a root ca na l s ea ler1 0.
Lindquist et a l (1981)1 8 repo rt ed t hat euge no l t hat le ac hes o ut o f z inc o xide eug e no l bas ed ro ot cana l sea le rs, ma y part ic ip at e in t he de ve lo p me nt o f per iap ic a l infla mmat io n o r t he co nt inua t io n o f a pre- e xist ing per iap ic a l le s io n. T he y co ndu ct ed a st ud y t o chec k t he a mo u nt o f euge no l re le as ed fro m fo ur d iffer e nt de nt a l mat er ia ls imme rs ed in pho sp hat e bu ffer me as ured b y g as liqu id c hro mat o graph y. M a xima l re lea se o f eug e no l fro m z inc o xide -e uge no l c e me nt (ZOE) a nd IR M wa s att a ine d w it hin 5 hr s a nd co rrespo nded t o 5% a nd 4%, resp ect ive ly, o f t he tot a l a mo u nt o f eug e no l in ea c h mat er ia l. Euge no l app lie d t o gro wing cu lt ure s o f hu ma n d ip lo id fibro bla st s re duce d t he nu mber o f ce lls reco vered t o a va lu e w h ic h wa s 4 % o f t hat fo u nd fo r co nt ro l cu lt ure s w hic h grew t o fo rm mo no la yer s. Co nflu e nt mo no la y ers o f H- Ur id ine la be le d c e llcu lt ur es w hic h wer e incu bat ed fo r 1 hr in t he prese nce o f 4 mm eug e no l lo st appro xima t ely 100% o f t his c yt o p la s mic la be l, ind icat ing t ot al c e ll deat h.
B ran stette r J (1982)1 9 st at ed t hat endo d o nt ic se a ler s are use d in t he o bt urat io n o f root cana l syst e m t o achie ve a flu id t ig ht o r her met ic se a l
t hro ugho ut t he ca na l inc lu d ing t he ap ica l fo ra me n a nd ca na l irregu la r it ie s.
B edoukian (1986)2 0 repo rt ed t hat eugeno l wa s fir st iso lat ed in 1929 a nd co mmerc ia l pro duct io n co mme nced in t he U nit ed St at es in t he 1940s. Euge no l c a n be pro duced synt het ica lly, t he mo st pract ic a l met ho d be ing t he a llylat io n o f gua ia co l w it h a llyl c hlo r id e. Ho we ver, euge no l is predo mina nt ly pre pared fro m nat ura l o il so urce s by mix ing t he e ss e nt ia l o il w it h a n e xce s s o f aqu eo us so d iu m (3%) o r pot ass iu m hydro xide so lut io n a nd s hak ing, le ad ing t o t he fo r mat io n o f a p he no lic a lka li s a lt . The inso lu ble no n -p he no lic po rt io n is t he n e xt ract ed w it h a so lve nt o r via st ea m d ist illat io n. T he u n - d isso lved po rt io n is re mo ved, t he a lk a li so lut io n is ac id ifie d at lo w t emp erat ures a nd t he liberat ed euge no l pur ifie d b y fra ct io na l d ist illat io n.
Thomp son et a l (1990)2 1 st at ed t hat t he euge no l ind uce d c yt o to xic it y is in flue nce d by t he fo r mat io n o f g lut at hio ne co nju gat es dur ing o xidat io n o f eug e no l by micro so ma l fract io n o f rat liver a nd lu ng.
Jeng et a l (1994)2 2 inve st igat ed t he e ffect o f e uge no l o n mo us e fibro bla st at w it h a co nce nt rat io n hig her t ha n 3 mmo l/L, a nd re po rt ed t hat euge no l wa s c yt o t o xic t o o ral mu co sa l fibro bla st s in a co nce nt rat io n a nd t ime -d epe nd e nt ma nn er. The y repo rt ed t hat ce ll deat h wa s a sso c iat ed w it h int ra ce llu lar d ep let io n o f g lut at hio ne. Mo st o f t he g lut at hio ne w as d ep let ed pr io r t o t he o nset o f ce ll d eat h. At co nce nt rat io ns o f 3 mmo l/ L a nd 4 mmo l/L, euge no l dep let ed a bo ut 45%
a nd 77% o f t he c e lls a ft er o ne - ho ur inc ubat io n. I n add it io n, euge no l
decrea se d ce llu lar ATP (Ade no sine tr i-p ho sp hat e) le ve l in a co nce nt rat io n a nd t ime d epe nd e nt ma nner a nd t he y co nc luded t hat euge no l is c yt o to xic t o ora l muco sa l fibro bla st s.
Tai et a l (2 002)2 3 st at ed t hat z inc o xide e uge no l ba sed s ea ler s irr it at ed per iap ic a l t iss ue s inv ivo a nd e xhib it ed c ytoto xic e ffe ct s e xvivo . I n t his st ud y, t et razo liu m bro mid e redu ct io n assa y, DN A prec ip it at io n as sa y, a nd DN A fr ag me nt at io n a na lys is w ere per fo r med t o inve st igat e t he c yt o to xic a nd ge no t o xic e ffe ct s o f fo ur differe nt root cana l se a ler s o n cu lt ured V79 ce lls. T he se find ings su g gest t hat ro ot cana l se a ler s co nt a ining fo r ma lde h yde a nd bis p he no l - A d ig lyet her pro ved as no t o nly c yt o to xic but a lso geno to xic. The y a lso st at ed t hat ma mma lia n t est s yst e m mig ht be u sed ro ut ine ly fo r e va luat io n o f t he ge no to xic it y o f de nt a l mat er ia ls in fut ure invest igat io ns.
Ito et a l (200 6)2 4 invest igat ed t he a nt i-o xid a nt act ivit y o f euge no l a nd it s iso mer iso eug e no l us ing iro n- me d iat e d lip id pero xid at io n a nd a ut o - o xidat io n o f Fe 2 +. The inh ib it o r y e ffe ct o f e uge no l o n lip id peroxidat io n was less potent with an IC 50 value o f ≈80 μm, which was e ig ht -fo ld t he va lu e o f iso e uge no l. T he mec ha nis m o f act io n o f t he t wo co mpo und s wa s inve st ig at ed a nd it wa s suggest ed t hat t he a nt i-o xid a nt capa c it y o f e uge no l co u ld be e xp la ined by t he fo r mat io n o f co mp le xe s w it h reduc ed met a ls. The po t ent inh ib it o ry e ffe ct o f iso euge no l o n lip id pero xidat io n ma y be re lat ed t o t he dec r ease d fo r mat io n o f t he per - ferr yl io n o r iro n-o xyg e n c he lat e co mp le x as t he in it iat ing fact o r o f lip id p ero xidat io n.
van Zy l et al (200 6)2 5 co nduct ed a st ud y in w hic h t we nt y e s se nt ia l o il co nst it ue nt s were e va lu at ed fo r t he ir a nt ima lar ia l a ct ivit y aga inst t he c hlo ro qu ine-res ist a nt st ra in P. f alcipa rum (FCR-3). Euge no l e xhib it ed so me a ct ivit y w it h a n I C 50 va lu e o f 753 μ m w hic h w as lo wer t ha n fo r other const ituents such as nero lido l, lina lylacetae, α -pinene and pulegone (IC 50 values ranging fro m 0.9 to 1.4 μ m)
Singh et a l(200 7)2 6 inve st igat ed t he a nt ibact er ia l e ffe ct o f euge no l o n t he gro wt h o f Gra m-po s it ive (B.cere us; B.subtili s; S.aur eus) a nd Gra m- negat ive (E . coli; S. typhi; P. ae rugino sa) bact er ia us ing t he agar we ll d iffu s io n met ho d. At 1,000 pp m, eug e no l inh ib it ed t he gro wt h o f t he bact er ia a nd co mp let e inh ib it io n w as o bt a ined ag a inst P.aer ugino sa at a hig h co nc e nt rat io n o f 2,000 pp m. This in h ib it io n wa s hig h in co mpar iso n t o a mp ic illin (1 mg/ mL) w he n used a s a po s it ive co nt ro l.
T rago o lpua et a l (2007)2 7 t est ed t he in vit ro a nt ivira l act ivit y o f euge no l ag a inst t he herpe s simp le x - 1: H SV-1 a nd HSV-2 viru se s. T he rep lic at io n o f t hes e viru se s w as inh ib it e d w it h I C 50 va lu e s o f 2 5.6 μg/mL and 16.2 μg/mL against HSV-1 and HSV-2, respect ively.
Ad d it io na l inve st ig at io ns re ve a led s yn erg ist ic int era ct io ns w it h a co mbinat io n o f eug e no l a nd ac yc lo vir, a kno w n a nt ivira l drug. St ud ie s ha ve s ho w n t hat app lic at io n o f eug e no l de la yed t he de ve lo p me nt o f herpes viru s- indu ced ker at it is in a mo u se mo d e l. Euge no l wa s e va luat ed fo r it s a nt i- HSV pro pert ie s o n st a ndard HSV-1( F), st a nd ard HSV-2(G) a nd t e n HSV iso lat es using t he p laque redu ct io n a ssa y. O n l y HSV-1 iso lat es 1 a nd 30 were inh ib it e d by euge no l a nd t he inh ib it io n
aga inst t hes e iso lat es w as great er t ha n fo r t he e xt ract o bt a ined fro m t he flo w er bu ds o f E. caryop hyllata.
B arc e lou x et a l (2008)2 8 repo rt ed t hat C lo ve has bee n u sed in a nc ie nt C hina a s a s p ice a nd fra gra nc e fo r mo re tha n 2,000 year s. Med ic ina lly, t he w e l l-k no wn t rad it io na l r e med y o f a pp lying c lo ve o il t o t reat a toothache was documented for the first time in 1640 in „Pract ice o f Physic‟ in France. In Chinese tradit io nal medicine clo ve o il has been used a s c ar minat ive, a nt isp asmo d ic, a nt ibact er ia l a nd a nt para s it ic age nt , while t he bud s w ere used t o t reat d yspep s ia, a cut e/c hro nic gast r it is a nd d iarr ho ea.
Danie l et a l (2009)2 9 inve st igat ed a bo ut Euge nia car yo p hyllat a, w hic h co nt a ins eug e no l a nd ac et yleu ge no l as ma jo r co nst it uent s fo r po t ent ia l a nt i- infla mmat o r y a ct io n o n CO X-2 (c yc lo o xyg e na se) a nd LOX e nz yme s. T he o il e xhib it ed st ro ng inh ib it o r y act iv it y aga inst CO X -2 (58.15%) and 1 5 -LO X (86.15%) e nz yme s at 10μg/ mL a nd 25μg/ mL, respe ct ive ly. T he a nt i- in fla mmat o r y act ivit y o f euge no l w as e va luat ed by in fla mmat o r y e xudat e vo lu me a nd leuko c yt e migrat io n in carrage e na n- ind uced p le ur is y a nd c arra gee na n- induc ed paw o ede ma t est s in r at s. Euge no l (20 0 a nd 400 mg/ kg) wa s fo u nd t o reduce t he vo lu me o f p le ura l e xudat es w it ho ut cha ng ing t he t o t al blo o d leu ko c yt e co unt ind ic at ing t he a nt i- in fla mmat o r y po t ent ia l o f euge no l.
Oka moto et a l (2009)3 0 e va luat ed t he a naest het ic pro pert y o f euge no l in ju ve nile Lepo r inusma cro cep ha lu s, a fish spec ie s a lso k no w n as
“piavuçu”. Eugeno l had strong anaesthet ic effect on L. ma crocep hal u s
a nd a do se o f 37.5 mg/L o f eu ge no l w as reco mme nd ed fo r t he fast a nd sa fe a na est hes ia o f p ia vuçu ju ve n ile s (n = 72). I n a s imilar inve st igat io n, t he e ffic ac y o f be nzo ca ine a nd e uge no l a s a nae st het ic s in Trachino t us marg inat us ju ve nile fish w as st ud ied. Be nzo ca ine a nd euge no l, at 50 pp m, induc ed fa st a na est he sia a nd r eco ver y (3 min a nd 5 min, re spe ct ive ly). I n Euro pea n sea bas s (D ice nt rarc hu s la br a x) ( n = 240), euge no l indu ced dee p a nae st he sia w it ho ut subst ant ia lly a ffe ct ing t he ir b lo o d pro file a s mea sured b y seru m co rt iso l, g luco se a nd hae mat o cr it va lue s. I nduct io n wa s ra p id but reco ver y w as ver y s lo w e ve n at lo w co nce nt rat io ns.
Hussain et al (2011)3 1 t est ed euge no l a lo ne a nd in co mbinat io n w it h a c he mo t herape ut ic dru g (ge mc it a bine ) t o e va luat e it s inh ib it o r y e ffe ct aga inst ca ncer c e lls. Euge no l sho we d do se -depe nd e nt se le ct ive c yt o to xic it y t o wards He La ce lls in c o mp ar iso n to no rma l c e lls.
Furt her mo re, euge no l a nd ge mc it a bine in co mbinat io n ind uced gro wt h inh ib it io n a nd apo pt o sis (pro gra mmed ce ll d eat h) at lo wer co nce nt rat io ns in co mp ar iso n t o t he ind ivid ua l co mpo unds ind ic at ing synergist ic int eract io ns. Eugeno l (150 μm) and gemc itabine (15 μm) resu lt e d in a de crea se in ce ll via bilit y fr o m 84% (euge no l a lo ne) a nd 51% (ge mc it a bine a lo ne) t o 47% in co mbinat io n. Tre at me nt o f co lo n ca nc er ce lls w it h euge no l resu lt ed in t he r educt io n o f int ra ce llu lar no n - prot e in t hio ls a nd a n incr ea se in t he ear lier lip id la ye r brea k. In add it io n, d iss ip at io n o f mit o c ho ndr ia l me mbra ne po t ent ia l a nd ge nerat io n o f re act ive o xyge n spe c ie s acco mp a nied t he euge no l - ind uced a po pto sis.
M arco s-A ri as et a l (2011)3 2 e va lu at ed t he e ffect o f euge no l ag a inst 38 Ca nd id a sp ec ie s iso lat ed fro m de nt ure -wearers a nd 10 co lle ct io n Ca nd id a st ra ins u s ing t he C LSI M27 - A3 bro t h micro d ilut io n met ho d.
It was fo u nd t hat euge no l e xhib it ed a nt ifu ng a l act ivit y w it h t he MIC va lue ra ng ing fro m 0.06 t o 0.25% (v/ v), w hile t he min imu m co nce nt rat io n o f drug t hat inhib it ed 50% o f t he iso lat es t est ed (de fine d as MI C 50 ) ra nged fro m 0.06 t o 0.12% (v/ v) . MIC a nd sk in le s io n sco r ing t est s were p er fo r med in vivo t o e va luat e t he a nt ifu nga l e ffe ct o f euge no l in a gu ine a p ig mo de l in fect ed by M icro spo ru m g yps eu m.
Euge no l e xhib it ed pro mis ing act ivit y w it h t he MIC va lu e ra ng ing fro m 0.01 t o 0.03%. T he sk in le sio n s co r ing t est s ho wed t hat eug e no l wa s c lin ica lly e ffect ive at impro ving t he les io n d ur ing t he fir st week o f app licat io n. Ho we ver, euge no l d id no t impro ve t he sk in le s io ns in fe ct ed by M. gyps eum a s det er mined in t he ha ir cu lt ure t est .
The wo rd o zone (O3) is d er ived fro m t he Gre ek wo rd o ze in (o do rant ). It is o ne o f t he mo st po wer fu l a nt imicro bia l ag e nt s a va ila ble fo r u se in me d ic ine a nd d e nt ist r y. I n 19 20, D r.Edwin Pa rr, a Sw is s de nt ist st art ed t he us e o f O3 a s a part o f his d is in fe ct io n s yst e m3 3.
M udd JB et al (1969)3 4 co nduct ed a st ud y o n t he re act io n o f o zo ne w it h a mino a c ids a nd pro t eins a nd st at ed t hat o zo ne act s o n ba ct er ia l ce ll me mbra nes by o xid at io n o f t he ir lip id and lipo pro t ein co mpo ne nt .
Roy D et a l (1981)3 5 co nduct ed a st ud y o n t he mec ha nis m o f e nt ero vira l inact ivat io n w it h o zo ne by co mpar ing t he sed ime nt at io n
pro file s o f nu c le ic ac id s o f o zo ne -e xpo sed viru se s w it h t ho s e o f co nt ro l viru se s, t he y o bser ved t hat vira l nuc le ic ac id wa s d a mag ed as a res u lt o f o zo nat io n. It wo u ld app ear t hat t he ma jo r cause o f vira l ina ct ivat io n, as det er mine d w it h po lio viru s 1, by o zo ne u s ing a re s idua l co nce nt rat io n le s s t ha n 0.3 mg/ lit er a nd a co nt act t ime up t o 2 min is da mage o f t he R N A. The y co nc lud ed t hat a ll virus es ar e su sc ept ib le t o o zo ne; yet d iffer w id e ly in t he ir suscept ibilit y a nd a lso st at ed t hat t he a na lys is o f vira l co mpo ne nt s s ho wed da ma ge t o t he po lypept ide a nd e nve lo p e pro t eins impa ir ing vira l at t ach me nt capa bilit y.
Alv a re z et a l (19 82)3 6 st ud ied t he g er mic ida l p ro pert ies o f t he o zo niz ed su nflo wer o il (O LEO ZON), t he y dec id ed t o make a n inve st igat io n in 8 0 c hildr e n bet wee n 1 a nd 14 ye ar o ld, w it h t he ma in o bject ive o f e va luat ing t he e ffe ct ive ness o f t he OLEO ZO N in pr imar y p yo der ma, co mp ar ing t his w it h G e nt a myc in cr ea m. The inve st igat io n was carr ied o ut in ar ea s o f hea lt h o f t he ge nera l t ea c hing ho s p it a l
"Co ma nda nt e P inare s ", Sa n Cr ist o ba l, P inar de l R io , last ing 6 mo nt hs.
Pat ie nt s wer e ra ndo mly a s sig ne d t o bo t h gro up o f t reat me nt ; e xper ime nt a l (u s ing OLEO ZO N) a nd c o nt ro l (u s ing Ge nt a myc in) .To a ll p at ie nt s bact er io lo g ic a l st ud ie s were do ne, at t he beg inn ing a nd a ft er 15 da ys o f t reat me nt . The re su lt s de mo nst rat ed t hat t he mo st frequ e nt iso lat ed ger m w as t he St repto coccus hae mo lyt ic o f gro up A.
Hea ling e ffe ct ive ne ss a ft er 15 da ys o f treat me nt , was 68 % fo r t he OLEO ZON a nd 2 8 % in t ho se pat ie nt s w here Ge nt a myc in w as use d. No side e ffe ct s were o bser ved.
M orgo li s HL et a l (1985)3 7 repo rt ed t hat o zo ne had a s e vere ly d isrupt ive e ffe ct o n car io g e nic ba ct er ia, resu lt ing in e liminat io n o f ac ido ge nic bact er ia. The st ro nge st nat ura lly o c curr ing ac id, pro duced by a c ido ge nic ba ct er ia, p yru vic ac id is d ecar bo x ylat ed to acet ic ac id, whic h in t urn e nco urage s re miner a lisat io n.
A rsa lane K et a l (1995)3 8 co nduct ed a s t ud y o n o zo ne st i mu lat io n o f in fla mmat o r y med iat o rs and repo rt ed t hat o zo ne be ha ves as a w eak c yt o k ine suc h as t u mo ur necro sis fa ct o r (TNF -α), int er leu k in (IL-2, IL- 6) and t rans fo r ming gro wt h fa ct o r -bet a (TGF-β). In t he pre se nt st ud y, t he in vit ro e ffe ct o f lo w co nc e nt rat io ns o f O3 (0.1 to 1 ppm) wa s e va luat ed o n ce ll via b ilit y a nd c yt o k ine se cret io n by a lveo lar ma cro pha ge s ( AM) fro m gu inea p ig s a nd hea lt h y su bje ct s. Ce ll in jur y was e st imat ed imme d iat e ly a ft er O3 e xpo s ure by e va lu at io n o f ATP ce ll co nt ent as me asur ed by bio lu mine sce nce a nd la ct ic de h ydro ge na se (LDH) re lea se in t he cu lt ure med iu m.
M udd JB et al (1997)3 9 invest igat ed t he e ffe ct o f o zo ne o n blo o d ce lls a nd co nc luded t hat o zo ne reduce s o r e liminat e s c lu mp ing o f re d blo o d ce lls a nd it s fle x ib ilit y is re st o red a lo ng w it h o xyg e n c arr ying a b ilit y.
Ozo ne wa s app lie d t o sea led r ed c e ll g ho st me mbra ne s at t he rat e o f 95 mmo l/ min fo r per io d s up t o 20 min. Acet ylc ho line e st erase, o n t he o ut er fa ce o f t he me mbr a ne, wa s inh ib it e d up t o 20%. G lyc era lde h yde - 3-pho sp hat e de hydro ge nas e, o n t he inner sur face o f t he me mbra ne, wa s inh ib it ed up t o 87%. These d iffer e nces re fle ct ed t he inhere nt suscept ib ilit ie s o f t he t wo e nz yme s a nd t he pre se nce o r a bse nce o f t he
me mbr a ne barr ier. T he y a lso st at ed t hat t here was no dec line in t he a mo u nt o f c ho lest ero l in t he lip ids der ive d fro m o zo ne -t reat ed red ce ll me mbr a ne s.
Liu Z et a l (1998 )4 0 st at ed t hat t he ap p lic at io n o f o zo niz ed o ils o f veget a l so urce co nst it ut es a n appro pr iat e met ho d fo r t he lo ca l t herap y o f se ver a l d is ea se s. The int era ct io n o f o z o ne w it h t hes e o ils pro duces a mixt ur e o f c he mic a l co mpo u nd s na me ly o zo no id es a nd pero xid e s w it h bet t er ger mic id a l po wer. The resu lt s o f t op ica l app lic at io n o f o zo nize d o il ( OLEO ZO N) in d er mat o lo g ica l d isea se s o f vir a l, fu ng i a nd bact er ia l o rig in, d ur ing 9 year s o f st ud y, w ere pre s e nt ed. Amo ng t he d is ea se s o f bact er ia l o r ig in (p yo d er ma), 600 pat ie nt s wer e t reat ed, o bt a ining hea ling cr it er io n in 87%. I n cas e o fmyco t ic d is e as es (ep ider mo p hyt ho s is a nd o nyc ho myco s is ), 1000 pat ie nt s rece ive d t he t reat me nt , w it h 91% o f hea ling. I n v ir a l d is ea se s ( herpe s s imp le x), 300 pat ie nt s w ere t reat ed and in 74% t h e r e lap se s d is appe ared. No s id e e ffe ct s were o bs er ved in a n y o f t he c ases a ft er t reat me nt .
Oizu mi M et a l (1998)4 1 repo rt ed t hat o ra l micro o rganis ms a nd no via b le C.a lb ica ns w ere det ect ed a ft er e xpo sure t o flo w ing o zo nat ed wat er (2 o r 4 mg/ L) fo r 1 min, sugg est ing t he app lic at io n o f o zo nat ed wat er mig ht be u se fu l in redu c ing t he nu mber o f C.a lb ic a ns o n de nt ure base s.
Ve lano HE et a l (2001)4 2 co nduct ed inv it ro st ud ie s a nd re po rt ed t hat o zo ne was e ffe ct ive o ver mo st o f t he ba ct er ia fo u nd in ca se s o f p u lp
necro s is. Ozo ne is e ffect ive w he n it is us ed in su ffic ie nt co nce nt rat io n, fo r a n ad equat e t ime a nd de livered ap pro pr iat e ly.
Ebensb e rg e r U et a l (2002)4 3e xp er ime nt ed t he u se o f o zo nat ed wat er in deco nt a minat io n o f a vu ls ed t o ot h be fo re rep la nt at io n a nd co nc lude d t hat 2 minut e irr ig at io n o f a vu lsed t eet h w it h no n - iso t o nic o zo nat ed wat er no t o nly pro vide s me c ha nic a l c le a ns ing but a lso deco nt a minat ed t he root sur fac e w it h no negat ive e ffect on per io do nt a l c e lls.
Reth et a l (2002)4 4 repo rt ed t hat o zo ne react s w it h u nsat urat ed fat t y ac id s o f t he lip id la ye r in ce llu lar me mbr a ne s, fo r ming h ydro ge n pero xide, o ne o f t he mo st s ig nific a nt c yt o k ine induc ers. It also p la ys a n impo rt ant ro le a s a se co nd ar y me s se ng er in t he in it iat io n a nd a mp lific at io n o f s ig na ling at t he a nt ig e n recept o r. The a nt ige n recept o rs t he mse lve s are H2O2-ge nerat ing e nz yme s a nd t hat t he o xidat ive burst in ma cro phag es see ms to p la y a ro le no t o nly in pat ho ge n k illing but a lso in t he act ivat io n o f t he se c e lls a s we ll a s ne ig hbo r ing c e lls.
Holme s J (200 3)4 5 st ud ie d t he c lin ic a l re versa l o f ro ot car ie s by o zo ne using do uble b linded ra ndo mize d co nt ro l st ud y a nd repo rt ed t hat o zo ne is cap a ble o f c lin ic a lly re ver sing t he lea t her y ro o t cario us le s io n. H e as se ss ed t he e ffect o f a n o zo ne d e liver y s yst e m, co mb ine d w it h t he da ily us e o f a re - miner a liz ing p at ie nt k it , o n t he c lin ic a l s e ver it y o f no n- ca vit at ed leat her y pr imar y ro o t cario us lesio ns (PRCL’s), in a n o ld er po pu lat io n gro up. He a lso repo rt ed t hat le at her y no n -ca vit at ed pr imar y ro ot car ie s ca n be arrest ed no n -o perat ive ly w it h o zo ne a nd re -
miner a liz ing pro duct s a nd t his t reat me nt reg ime is a n e ffe ct ive alternat ive to convent iona l “drilling and filling”.
Walke r JT et a l (2003)4 6 repo rt ed t hat in mo de l de nt a l u nit wat er line s, o zo ne a c hie ved a 57% redu ct io n in bio film a nd a 6 5% redu ct io n in via b le bact er ia in sp it e o f be ing u se d in a ver y lo w do se a nd w it h a s ho rt t ime o f ap p licat io n. The y a lso co nc lude d t hat de nt a l ma nu fact urers s ho u ld be inv it ed t o des ig n de nt a l u nit s t hat inco rpo rat e aut o mat ed de vic es t o dis in fe ct DUWLs.
Nagayoshi M et al (2004)4 7 reported that ozonated water had strong bacteric idal act ivit y against bacteria in pla que bio film. It was fo und t hat ozonated water (0.5–4mg/L) was highly effect ive in killing o f bot h gram - posit ive and gram-negat ive microorganis ms . Gram-negat ive bacteria, such as P. gingivalis P.endodontalis were more sensit ive to ozonated water than gram-posit ive o ra l st rept o co cc i a nd C . a lbic a ns in p ure cu lt ure.
B aysan A et a l (2004)4 8 co nd uct ed a st ud y o n t he e ffic ac y o f o zo ne o n pr imar y ro ot car ie s le s io n (PR C L) a nd fo und t hat o zo ne app lic at io n fo r e it her fo r 10 o r 20 s dra mat ica lly redu ce d mo st o f t he micro o rganis ms in PR C Ls w it ho ut any s ide e ffe ct s reco rded at reca ll int er va ls bet wee n 3 and 5.5 mo nt hs.
Young SB et al (2004)4 9 e xa mine d t he mec ha nis m o f k illing B. subti lus spo res by o zo ne a nd r epo rt ed t hat it see ms t o render t he spo res de fect ive in ger minat io n p erhap s bec au se o f da ma ge t o t he s po re ‟s inne r me mbra ne.
Huth K C et al (2005)5 0 co nduct ed in vit ro st ud ie s w it h s ho rt t erm fo llo w-up a nd a s se ss ed t he e ffe ct o f o zo ne o n p it a nd fis sur e c ar ie s a nd pr imar y ro ot caries w it h re su lt s sho w ing s ig n ific a nt reduct io ns in t he nu mber o f micro o rganis ms in t he car io us le s io ns. In t his st ud y ga seo us o zo ne (Hea lOzo ne) wa s app lie d o nce fo r 40 seco nd s t o t he ra ndo mly as sig ned t est mo lar o f eac h pa ir w it ho ut t he u se o f re- miner a liz ing so lut io ns. Les io n pro gress io n o r rever sa l was mo n it o red by t he la ser fluo r esc e nc e s yst e m DI AGNOd e nt fo r up to 3 mo nt hs a nd t he det er io rat io n o r impro ve me nt co mpare d bet wee n t he o zo ne -t reat ed le s io ns a nd t he u nt reat ed co nt ro l le sio ns
Ce libe rti P et a l (2 006)5 1 st at ed t hat o zo ne ha d no influ e nce o n t he phys ic a l pro pert ies o f t he e na me l t o enha nce o r hind er t he sea ling a bilit y. Thu s, o zo ne ca n be app lie d o ve r int act a nd prepared e na me l dur ing t he re st o rat io n pro ces s. T he y a lso st at ed t hat t he app lic at io n o f gas ifo r m o zo ne do e s no t affe ct t he mo du lus o f e la st ic it y a nd t he Vicker‟s hardness o f dent in.
Po lydo rou OPK et al (2006)5 2 e xp er ime nt ed t he a nt imicro bia l e ffica c y o f o zo ne a nd repo rt ed t hat lo ng e xpo sur e to o zo ne ga s ha s a st ro ng a nt i- bact er ic id a l e ffe ct o n m icro o rganisms w it hin t he de nt ina l t u bu le s o f de ep ca v it ie s, w hic h co u ld resu lt in increa s ing t he c lin ic a l su cce s s o f t he re st o rat io n. I n t his st ud y t hirt y- fiv e hu ma n mo lar s w ere d iv ided int o 5 gro ups. Ca v it ie s were t he n cut int o t he t eet h ( n=2 8 ca vit ie s pe r gro up). Aft er st er iliz at io n, t he t eet h were le ft in bro t h cu lt ures o f 1 06 co lo ny- fo r ming u nit s (C FU) ml) o f S. mut ans at 36ºC fo r 48 hr s. T he
appro pr iat e t reat me nt fo llo w ed (gro up A, co nt ro l; gro up B, C le ar fil SE Bo nd ; gro up C, C lear fil Pro t ect Bo nd; gr o u p D, 40 s o f t reat me nt w it h o zo ne; a nd gro up E, 80 s o f t reat me nt w it h o zo ne), a nd t he ca v it ie s were t he n fille d w it h co mpo s it e resin. Aft er 72 hr s, t he re st o rat io ns were re mo ved, de nt in c hip s wer e co lle ct ed w it h a n e xca vat o r, and t he tot a l nu mber o f mic ro o rganis ms wa s det ermined. All t reat me nt s s ig nific a nt ly re duc ed t he nu mber o f S. mut ans pre se nt co mpar ed w it h t he co nt ro l gro up. The a nt imicro bia l e ffect o f bo t h bo nd ing s yst e ms a nd t reat me nt w it h 80 s o f o zo ne wa s sig nific a nt ly h ig her t ha n t he 40 s o zo ne t reat me nt . The y co nc lude d t hat Hea l O zo ne a nd t he bo nd ing s yst e ms s ho wed st r ik ing a nt imicro bia l e ffect s aga inst S. mut a ns.
B aysan A et a l (2007)5 3 a sses sed t he e ffe ct o f t he o zo ne - me d iat ed k illing o f ba ct er ia in in fect ed de nt ine a sso c iat ed w it h no n -ca v it at ed o cc lu sa l car io us le s io ns a nd st at ed t hat t he lo ng er t he co nt act t ime, t he bet t er t he micro bio lo g ic a l k ill rat e. It was repo rt ed t hat by incr ea s ing t he co nt act t ime fro m 1 0 t o 20 s, t he ba ct er ia l k ill rat e c ha nged fro m o zo ne be ing a d is in fe ct a nt t o acqu ir ing st er iliz ing e ffe ct .
B aysanA et a l (2007)5 4 sugge st ed t hat t he re ver sa l o f car io us le s io ns a ft er t reat me nt w it h o zo ne dep e nd o n t he s ize a nd lo ca liz at io n o f t he car io u s les io n. No n- ca vit at ed le sio ns wer e mo re lik e ly t o revers e t ha n ca vit at ed le s io ns. T he hard nes s va lue s impro ved s ig nific a nt ly in t he o zo ne-t reat ed t est les io ns a ft er 4, 6, and 8 mo nt hs (P <0.05) co mpare d w it h ba se line w hile t he co nt ro l le sio ns had no sig nific a nt cha nge in hard ne ss at a ny r eca ll int er va l.
Huth K C et al (2007)5 5 st at ed t hat Ozonat ed wat er has a n e xce lle nt a nt i- in fla mmat o r y cap ac it y o n N F ( nu c lear fact o r) -kappa s yst e m, a parad ig m fo r in fla mmat io n - as so c iat ed sig na lling/t ra ns cr ipt io n. T he ir resu lt s s ho wed t hat NF -ka ppa B a ct ivit y in o ra l ce lls in per io do nt a l lig a me nt t issu e fro m ro ot surface s o f per io do nt a ly d a mage d t eet h was inh ib it ed fo llo w ing incu bat io n w it h o zo nized med iu m.
M ulle r et a l(2007)5 6co mpared t he in flu e nce o f o zo ne g as w it h pho t o dyna mic t herap y (PDT) a nd k no wn a nt isept ic ag e nt s (2%
C hlo r he xid ine, 0,5 and 5% hypo c hlo r it e so lut io ns) o n a mu lt is pe c ie s o ral b io film in vit ro . The fo llo w ing bact er ia were st ud ied – A . naeslundii, F. nucl eatum, S .sob rinu s, S . orali s a nd C. albican s. Gas ifo r m o zo ne wa s pro duced by vacuu m o zo ne de liver y s yst e m Ka vo Hea l o zo ne. The y co nc lud ed t hat t he mat r ix -e mbe dde d micro bia l po pu lat io ns in bio film ar e we ll pro t ect ed to wards a nt imicro bia l ag e nt s.
Only 5 % hypo c hlo r it e so lut io n w as a ble t o e liminat e a ll bact er ia e ffe ct ive ly. U sag e o f ga s ifo r m o zo ne o r PDT was no t able t o reduce s ig nific a nt ly o r co mp let e ly e liminat e ba ct er ia in t he bio film.
Pet ru cci M T et a l (200 7)5 7 co nd uct ed a s t ud y in w hic h t he y co mpar ed t he use o f o zo nat ed o il in a n e xper ime nt a l gro up to a co nt ro l gro up in whic h a nt ibio t ic t herap y w as used in t he t reat me nt o f a lveo lit is , it wa s fo u nd t hat pat ie nt s t reat ed w it h o zo nat ed o il he a led mo re qu ick ly.
There wa s co mp let e he a ling o f t he le sio ns w it h t he d is appe ara nc e o f sympt o ms.
Noga le s CG et a l (2 008)5 8 co ndu ct ed a n e xp er ime nt a l st ud y o n Ozo nat ed o il a nd co nc lud ed t hat whe n a pp lie d o n herp es la bia lis a nd ma nd ibu lar o st eo mye lit is d e mo nst rat ed fa st er hea ling t ime s t ha n co nve nt io na l pro to co ls. Ozo ne in t hese ca se s neut ra lis e s herp es vir io ns by d ire ct act io n, t hus inh ib it ing bact e r ic id a l supra - infe ct io ns a nd st imu lat ing t he he a ling o f t is sue s t hro ugh c ir cu lat o ry pro mpt ing.
Noguchi et a l (2009)5 9 repo rt ed t hat irr igat io n o f t he ro ot sur fac e o f a vu ls ed t eet h w it h o zo nat ed wat er d id n ot revea l a neg at ive e ffe ct o n per io do nt a l liga me nt ce ll pro liferat io n. In t his st ud y a ft er t he pre - t reat me nt o f LPS ( Lipo po lysacc har ide) w it h aqueo us o zo ne, e ffe ct s o f LPS a nd aqu eo us o zo ne -t reat ed LPS on c e ll v ia bilit y; ca lc ific at io n a bilit y; e xpre ss io n o f c yc lo o xyg e nase 2 ( COX -2), int er le uk in 6 (IL -6), and tumour necrosis factor α (TNF-α); and act ivat io n of p38 o f KN -3 ce lls were e xa mine d. The ir resu lt s sug gest t hat o do nt o bla st ic ce lls e xhib it in fla mmat o ry re spo nses aga inst LPS a nd t hat o zo nat ed wa t er has t he a bilit y t o impro ve LPS - ind uced infla mmat o r y re spo nse s a nd suppre s s io n o f o do nt o blast ic pro pert ies o f KN -3 ce lls t hro ug h d ire ct inh ib it io n o f LPS.
Huth K C et a l (2 009)6 0de mo nst rat eda nt imicro bia l act io n ag a inst bact er ia l st ra ins, s uc h a s My cobact er ia, E .f aecalis, and C. albica ns.
Not ably, w he n t he s pec ime n wa s irr igat ed w it h so nicat io n, o zo nat ed wat er had ne ar ly t he s a me a nt imicro bia l act ivit y as 2.5% NaO C l. T his st ud y in ve st igat ed w het her g aseo us o zo ne (4 · 106 l g m.) a nd aqueo us o zo ne (1.25 –20 l g m.) e xert any c yt o to xic e ffect s o n hu ma n o ra l
ep it he lia l ce lls a nd g ing iva l fibro bla st (HG F -1) ce lls co mpar ed w it h est a blis hed a nt is ept ic s [c hlo r he xid ine d ig lu co nat e (CH X) 2%, 0.2%;
so d iu m h ypo c hlo r it e (Na OC l) 5.25%, 2.25 %; h ydro gen pero xide (H2O2) 3%), o ver a t ime o f 1 min, a nd co mp ared w it h t he a nt ibio t ic, met ro nida zo le, o ver 24 hrs. Ce ll co u nt s, met abo lic a ct ivit y, Sp -1 bind ing, a ct in le ve ls , a nd apo pt o sis wer e e va luat ed. Ozo ne ga s wa s fo u nd t o ha ve t o xic e ffe ct s o n bo t h ce ll t yp es. Es se nt ia lly no c yt o to xic s ig ns w ere o bser ved fo r aqueo us o zo ne s uggest ing it s bio co mp at ibilit y whe n co mp ared t o ot her che mica ls.
K im HS et al (2009)6 1 has u ndert ak e n a st ud y t o eva lu at e t he t herapeut ic e ffe ct s o f t o pica l o zo nat ed o live o il o n a cut e cut a neo us wo und hea ling in a gu ine a p ig mo de l a nd a lso t o elu c idat e it s t herapeut ic me c ha n is m. Aft er cre at ing fu ll -t hic k ne s s s k in wo u nd s o n t he bac ks o f gu in ea p ig s b y using a 6 mm pu nc h bio p s y, w e e xa mined t he wo und hea ling e ffe ct o f to pica lly ap p lie d o zo nat ed o live o il (o zo ne gro up), as co mpared t o t he pure o live o il (o il gro up) a nd no n -t reat me nt (co nt ro l gro up). Immu no hist o c he mic a l st a ining de mo nst rat ed up regu lat io n o f p lat e let der ived gro wt h fa ct o r (PDGF), t rans fo r ming gro wt h fact o r -bet a (TGF bet a) a nd vascu lar e ndo t he lia l gro wt h fact o r (VEGF) e xpre ss io ns, but no t fibro bla st gr o wt h fa ct o r expre ss io n in t he o zo ne gro up o n da y 7, as co mpared w it h t he o il gro up. He co nc lu ded t hat t hese re su lt s de mo nst rat e t hat to p ica l app lic at io n o f o zo nat ed o liv e o il ca n a cce lerat e a cut e cut a neo us wo und repa ir in a g u ine a p ig in asso c iat io n w it h t he incr ea sed e xpressio n o f PDG F, TGF - bet a, and VEGF.
Atabe k et a l (2011)6 2 co nduct ed a st ud y to eva luat e t he e ffic ie nc y o f o zo ne a lo ne a nd w it h a re - miner a liz ing so lut io n fo llo w ing app lic at io n o n init ia l p it a nd fis sure c ar ies le s io ns in per ma ne nt ma nd ibu lar mo lar s. I n t he ir st ud y fo rt y c hildre n (9 -1 2 year s) ha v ing no n-ca vit at ed fis sure c ar ies le s io ns o n bilat era l 40 fir st per ma ne nt ma nd ibu la r mo lar t eet h were part ic ipat ed in t he st ud y. Pat ie nt s were ra ndo mly a llo cat ed to 2 exper ime nt a l gro ups co nsist ing o f 2 0 su bje ct s. In t he fir st gro up, o zo ne wa s a pp lied o nc e fo r 40 seco nd s to t he as s ig ned t est t eet h o f eac h pa ir. In t he se co nd gro up, o zo ne wa s app lie d o nce fo r 40 seco nds to t he as s ig ned t est t eet h o f ea c h p a ir w it h t he us e o f r e - minera liz ing so lut io n. Pro gres s io n o r impro ve me nt o f t he c ar ies wa s as se ss ed at base line, imme d iat e ly a ft er t reat me nt a nd at 1 -, 2-, 3-, and 6 mo nt h fo llo w-up by co mp ar ing t he DI AGNO d e nt va lue s, C lin ic a l Se ver it y Inde xe s, Ora l H yg ie ne Sco res. The y co nc luded t hat o zo ne t reat me nt e it her a lo ne o r co mb ined w it h a re - mine r a liz ing so lut io n wa s fo u nd t o be e ffe ct ive fo r re miner a lis at io n o f in it ia l fis sure car ie s le s io ns.
Ge rsp ach et a l (20 12)6 3 inve st ig at ed t he a nt imicro bia l e ffic ac y o f gaseo us o zo ne o n bact er ia ad her ed t o var io us t it a niu m a nd z ir co nia sur face s a nd t o eva luat e a d he s io n o f o st eo bla st lik e M G-63 c e lls to o zo ne-t reat ed sur fa ce s. Sa liva - co at ed t it a niu m (S LA a nd po lis hed) a nd z irco nia (ac id et che d a nd po lis he d) d isk s ser ved a s su bst rat es fo r t he ad here nc e o f S. sangu inis a nd P.gin giv alis. The t est spec ime ns were t reat ed w it h g aseo us o zo ne (140 pp m; 33 mL/s) fo r 6 a nd 24 s. Ba ct er ia were re- su spe nd ed using u lt r aso n icat io n, ser ia lly d ilut ed a nd cu lt ur ed.
MG-63 ce ll a d he sio n wa s a na lyse d w it h re fer e nce t o ce ll at t achme nt ,
mo rp ho lo g y, spread ing, a nd pro liferat io n. Sur fac e t o po graphy a s we ll as ce ll mo rp ho lo g y o f t he t est spec ime ns was ins pect ed by SEM. The y co nc lud ed t hat gas eo us o zo ne sho wed se lect ive e ffica c y t o reduce ad here nt ba ct er ia o n t it aniu m a nd z irco nia w it ho ut a ffe ct ing ad he s io n a nd pro liferat io n o f o st eo bla st ic ce lls. T his in vit ro s t ud y ma y pro vide a so lid ba s is fo r c lin ic a l st ud ie s o n ga seo us o zo ne t reat me nt o f p er i- imp la nt it is a nd re ve a led a n e ss e nt ia l ba se fo r su ffic ie nt t issu e rege nerat io n.
Chand ra SP et a l (2014)6 4 co ndu ct ed a st ud y t o eva luat e t he s ucc es s rat e o f mixt ure o f o zo nat ed o il a nd z inc o xide as a pr imar y t eet h ro ot filling mat er ia l. T he st ud y inc luded 6 0 in fe ct ed pr imar y ma nd ibu lar mo lar s w hic h w ere equa lly d iv id ed int o st ud y gro up (o zo nat ed o il -z inc o xide) a nd co nt ro l gro up (z inc o xid e -eu ge no l). Pu lpect o my pro cedur e was per fo r me d a nd t he c hildr e n were fo llo wed at regu lar int er va ls. All t he c hildr e n wer e a va ila ble fo r e va luat io n at t he e nd o f 12 mo nt hs. The t eet h were e va lu at ed fo r suc ce ss o r fa ilure ba sed o n c lin ica l a nd rad io grap hic cr it er ia by a blinde d inve st ig at o r. Clin ica l a nd rad io grap hic e va lu at io n sugg est ed t hat teet h o bt urat ed w it h o zo nat ed o il-z inc o xid e de mo nst rat ed go o d succe ss rat e (93.3 %) as co mpar ed to z inc o xide eu ge no l (63.3 %). Ho wever , no st at ist ic a lly s ig n ific a nt var iat io n (p = 0.408) w as o bser ve d bet wee n t he gro ups. T he y co nc lud ed t hat Ozo nat ed o il- ZnO de mo nst rat ed a goo d c lin ic a l a nd rad io grap hic s ucc ess at 12 mo nt hs fo llo w -up a nd it ca n be co nsider ed as a n a lt er nat ive o bt urat ing mat er ia l in in fect ed pr imar y t eet h.
Noites et a l (2014)6 5 co ndu ct ed a st ud y t o det ermine w het her irr ig at io n w it h so d iu m h ypo chlo r it e, c hlo r he x id ine, and o zo ne g as, a lo ne o r in co mbinat io n, were e ffe ct ive aga inst E. faecali s and C . albican s a nd co nc lud ed t hat o nly t he co mbine d act io n o f 2% C HX a nd o zo ne ga s fo r s ho rt perio d pro mo t es t he co mp let e e limin at io n o f bo t h micro o rganis ms t est ed in t he t oot h mo de l. T he re su lt s o f t his st ud y co u ld le ad t o red ir ect e ffo rt s t o wards new pro to co ls fo r reduc ing micro bia l lo ad in in fe ct ed ro ot cana ls lo o k ing fo r synerg is ms bet wee n new mo da lit ie s (o zo ne) o r a lread y k no w n a nt imic ro bia l pr o duct s.
Anand et a l (2015)6 6e va luat ed t he a nt ibact er ia l pro pert y o f o zo nat ed wat er, whit e w ine (1 4%) a nd de -a lco ho lis e d w hit e w ine aga inst S.mutans a nd E.f aecalis a nd co nc lud ed t hat o zo nat ed wat er has t he be st a nt ibact er ia l pro pert y a mo ng t he st ud y gro ups and t he a nt ibact er ia l act io n o f C a lc iu m h ydro xid e is e nha nce d w he n it is mixe d w it h de - a lco ho lis e d w hit e w ine.
Ahmedi et a l (2016)6 7a sse s sed t he e ffic ac y o f o zo ne ga s (O3) o n t he reduct io n o f dr y so cket (DS) o ccurrenc e fo llo w ing surg ic a l e xt ract io n o f lo wer ja w t hird mo lar s a nd co nc lud e d t hat O3 gas ha s a po s it ive e ffe ct o n reduc ing t he d e ve lo p me nt o f DS a nd p a in fo llo w ing t hird mo lar surg er y. T he micro bio lo g ic a l a nd met abo lic c apa bilit ie s o f O3 fo r pro mo t ing ha e mo st as is, increa sin g t he supp ly o f o xyg e n, a nd inh ib it ing ba ct er ia l pro lifer at io n incr ea se t he o ppo rt unit ie s fo r it s u se in a ll fie ld s o f surg er y. T hese c ha nge s o n t he ce llu lar le ve l e xp la in e le vat io n o f fu nct io na l act iv it y a nd reg e ne rat io n po t ent ia l o f t issue s
a nd o rgans w hic h ma y a cco unt fo r t he inc rease d perc e nt age o f via b ilit y in mo us e fibro bla st s w he n t reat ed w it h o zo nat ed euge no l. It a lso act ivat es aero bic pro ces se s like g lyco lys is a nd Kr e bs c yc le at ce llu lar le ve l t o st imu lat e b lo o d c ir cu lat io n; he nce o zo ne is a lso us ed fo r t reat me nt o f c ircu lat o r y d iso rders.
