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VOLUME 43 NUMBEP 5 MAY 2012 401

QUI NTESSENCE I NTERNATI ONAL


immune response.
1
There is an increasing
body of evidence available to implicate the
role of oxidative stress in the pathogen-
esis of a variety of inflammatory disorders,
including periodontal disease.
1,2
Oxidative
stress is caused by an imbalance between
the production of reactive oxygen spe-
oios (POS) and a biologio systom's ability
to readily detoxify the reactive intermedi-
ates or easily repair the resulting damage.
Mammalian oolls oan gonorato POS by
different biologic mechanisms and, in the
periodontium, neutrophil infiltration, fibro-
blasts, osteoclasts, and endothelial cells,
predominantly lead to an increase in the
POS lovols, rosulting in broakdown o tno
epithelial structure and damage to the con-
nective tissue in the near vicinity.
35
Smoking is an indopondont risk aotor
and is known to navo a signiioant ooot
on the initiation, extent, and progression
Periodontal diseases are inflammatory dis-
orders that give rise to tissue damage and
loss as a result of complex interactions
between pathogenic bacteria and the hosts
1
Reader, Department of Periodontics, SVS Institute of Dental
Sciences, Mahabubnagar, Andhra Pradesh, India.
2
Senior Lecturer, Department of Periodontics, Gitam Dental
College and Hospital, Visakhapatnam, Andhra Pradesh, India.
3
Senior Lecturer, Department of Public Health Dentistry, SVS
Institute of Dental Sciences, Mahabubnagar, Andhra Pradesh,
India.
4
Senior Lecturer, Department of Periodontics, SVS Institute of
Dental Sciences, Mahabubnagar, Andhra Pradesh, India.
5
Professor, Department of Periodontics, SVS Institute of Dental
Sciences, Mahabubnagar, Andhra Pradesh, India.
6
Professor, Department of Periodontics, Gitam Dental College
and Hospital, Visakhapatnam, Andhra Pradesh, India.
Correspondence: Dr Rampalli Viswa Chandra, Department of
Periodontics, SVS Institute of Dental Sciences, Mahabubnagar,
Andhra Pradesh, India. Email: viswachandra@hotmail.com
Effcacy of lycopene as a locally delivered gel
in the treatment of chronic periodontitis:
Smokers vs nonsmokers
Pampalli viswa Cnandra, MDS
1
/Yollartni P. Sandnya, MDS
2
/
Sripriya Nagara|an, MDS
3
/Bavigadda Harisn Poddy, MDS
4
/
Anumala Navoon, MDS
5
/K. Pa|a v. Murtny, MDS
6
Objective: The present study was carried out as a multicenter, randomized controlled,
split-mouth clinical trial to evaluate the efficacy of locally delivered lycopene on periodontal
noaltn and gingival orovioular luid (GCF) lovols o 8-nydroxydooxyguanosino (8-OHdG) in
smokors and nonsmokors oomparod witn poriodontally noaltny oontrol sub|oots. Method
and Materials: Ono nundrod ton sub|oots inoluding 50 smokors, 50 nonsmokors, and
10 oontrols partioipatod in tnis study. Sub|oots in tno smokor and nonsmokor groups
nad oontralatoral sitos troatod witn lyoopono gol and a plaoobo. Clinioal paramotors
included recording site-specific measures of plaque, gingivitis, probing depth, and clinical
attaonmont lovol. GCF 8-OHdG valuos woro analyzod using a oommoroially availablo
ELSA kit. Results: Comparod witn tno plaoobo, lyoopono-troatod sitos in smokors and
nonsmokors snowod signiioant roduotions in probing doptns and gain in tno olinioal
attaonmont lovols. Howovor, tnoro was no statistioally signiioant dioronoo in tno olinioal
paramotors wnon lyoopono-troatod sitos in smokors and nonsmokors woro oomparod,
oxoopt or tno roduotion in tno 8-OHdG lovols. Tno 8-OHdG lovols at 1 wook and 3 montns
in sitos troatod witn lyoopono in tno smokor and nonsmokor group woro oomparablo witn
those in the periodontally healthy control group. Conclusion: The gel formulation was
effective in increasing clinical attachment and reducing gingival inflammation, probing
doptn, and oxidativo in|ury oomparod witn tno plaoobo in smoking and nonsmoking
sub|oots. (Quintessence Int 2012;43:401411.)
Key words: antioxidants, lyoopono, poriodontitis, smoking
402 VOLUME 43 NUMBEP 5 MAY 2012
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
of periodontal disease. The immunosup-
prossivo ooots o smoking,
6
along with its
ability to selectively alter the subgingival
environment, allowing the overgrowth of
periodontopathic bacteria, are well docu-
mented.
7
Wnilo tobaooo smoko in itsol is a
source of free radicals,
8
nicotine, the main
oomponont o tobaooo smoko, nas tno
potontial to innibit noutropnil POS roloaso in
low doses, impairing the elimination of peri-
odontal pathogens.
9
In high doses, nicotine
aotually stimulatos POS roloaso tnat oausos
oxidative stress-mediated tissue damage in
the gingivoperiodontal tissues.
8,9

Tno ooots o POS aro gonorally nogat-
ed by a complex system of antioxidants,
suon as glutatniono, vitamin C, and vitamin
E, as well as enzymes such as catalase,
suporoxido dismutaso (SOD), and various
peroxidases.
5
Howovor, smokors also oxpo-
rience a decrease in the serum concentra-
tions o ma|or oiroulating antioxidants duo
to tno obligatory uso o tno body's rosorvo
of antioxidants to detoxify the unusually
high levels of these free radicals in tobacco
smoko.
10,11
Diroot oral supplomontation o
antioxidant onzymos suon as SOD and
glutathione peroxidase is impractical since
they are readily prone to inactivation by
gastric secretions,
12
thus necessitating the
use of extraneous antioxidant supplements.
Antioxidant supplementation has the
potential to decrease damage because of
oxidative stress, and purified lycopene prep-
arations have been shown to decrease oxida-
tive damage and reduce the concentrations
o various markors o oxidativo stross suon
as 8-nydroxydooxyguanosino (8-OHdG).
13

8-OHdG is tno bost-dooumontod biomarkor
of oxidative stress, which is involved in the
instigation of various diseases, including
periodontitis.
14
An inoroaso in 8-OHdG oan
be considered a sign of periodontal dam-
age,
3
and the subsequent posttreatment
decrease in its levels has been well dem-
onstrated in periodontitis, thus reflecting the
status of periodontal health.
4
Sinoo smoking
oan signiioantly inoroaso sorum 8-OHdG
lovols, 8-OHdG is oonsidorod a usoul bio-
markor o oxidativo oonditions, inoluding
poriodontitis, aootod by smoking.
15
While the effect of systemically admin-
istered lycopene on gingivitis has been
investigated,
1
the authors are not aware of
published data on the use of a locally deliv-
ered lycopene in the treatment of periodon-
titis. The present study has been carried
out as a multicenter, randomized controlled
clinical trial to evaluate the efficacy of a
locally delivered antioxidant gel on peri-
odontal health and gingival crevicular fluid
(GCF) lovols o 8-OHdG in smokors and
nonsmokors oomparod witn tno poriodon-
tally healthy control group.
METHOD AND MATERIALS
Source of data
A total of 110 systemically healthy male sub-
|oots solootod rom tno outpationt sootion o
tno Dopartmont o Poriodontios, in tno par-
tioipating institutions (SvS nstituto o Dontal
Soionoos, Manabubnagar, Andnra Pradosn,
ndia, and Gitam Dontal Collogo and
Hospital, visaknapatnam, Andnra Pradosn,
ndia) botwoon January 2010 and Fobruary
2011 were divided into three groups. The
smokor group (n = 50) inoludod sub|oots
between the ages of 25 and 50 years pre-
senting with a minimum of four periodontal
pookots witn probing doptn . 5 mm and a
nistory o oigarotto smoking . 10 cigarettes
a day for a minimum of 5 years. Individuals
in tno smokor group woro drawn rom a
pool of patients who persistently refused to
partioipato in a smoking-oossation program
oorod to tnom roo o oost. Tno nonsmokor
group (n = 50) oomprisod systomioally
noaltny sub|oots botwoon tno agos o 25
and 50 years with a history of chronic peri-
odontitis presenting with a minimum of four
poriodontal pookots witn probing doptns
. 5 mm. To assoss 8-OHdG lovols in pori-
odontally healthy individuals, 10 systemi-
oally noaltny sub|oots botwoon tno agos o
25 and 50 years with clinically healthy peri-
odontium acted as negative controls and
received no treatment apart from meticulous
plaque control instructions. To exclude the
possibility o poriodontal disoaso, sub|oots
in this group underwent a six-point probing
in all tno tootn. Padiograpns woro takon at
suspect sites to rule out periodontal dis-
oaso. Tno ollowing sub|oots woro oxoludod:
medically compromised patients, patients
receiving any form of periodontal therapy
VOLUME 43 NUMBEP 5 MAY 2012 403
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
(surgioal or nonsurgioal), tnoso wno nad
received antibiotic or antioxidant therapy
within 6 months of baseline examination,
patients who had used antibacterial mouth-
wash or medicated toothpaste within the
past month of baseline examination, and
patients with a history of deleterious habits.
All patients gave informed consent, and the
study protocol was explained in the local
language. Ethical clearance for the study
was received from the ethical committees
of the participating institutions (approval
no. 17-1209SvS and 10897).
Clinioal paramotors inoludod rooord-
ing noninvasive site-specific measures of
plaquo (Plaquo ndox |P])
16
and gingivitis
(modiiod Gingival ndox |MG])
17
at base-
line and 1, 3, and 6 months. Probing depth
(PD) and olinioal attaonmont lovol (CAL)
were recorded at baseline (before scaling
and root planing) and at 3 and 6 montns.
A oustom-mado aorylio stont and a UNC
no. 15 color-coded periodontal probe were
used to standardize the measurement of
sito-spooiio PD and CAL.
Sample size
Tno primary outoomo, gain in tno CAL, was
selected as the critical variable to calculate
tno samplo sizo. Fity pationts (100 in total)
in tno smokor and nonsmokor groups woro
roquirod to navo an 80% onanoo o dotooting
as signiioant (at tno two-sidod 5% lovol) a
1% dioronoo botwoon tno two groups in tno
moan CAL, witn assumod standard doviation
o 2.5, and a plannod 15% rato o missing
patients on follow-up. An additional 10 sub-
|oots woro usod as nogativo oontrols or oom-
parison o 8-OHdG lovols witn tno smokor
and nonsmokor groups or gaining additional
information on the treatment effects. Thus, the
inal samplo sizo inoludod 110 sub|oots.
Randomization and blinding
Pandomization and blinding inoludod
computerized generation of the allocation
soquonoo in random pormutod blooks as
dosoribod by Sagnaoi
18
and blinded dis-
bursement of medication. The medication
was plaood by tnroo invostigators (P.v.C.,
B.H.P., and Y.P.S.) in oontralatoral sitos,
whereas the relevant readings were record-
od by two oalibratod invostigators (A.N. and
K.P.v.M.), all o wnom woro blindod to tno
group (smokors/nonsmokors) and to tno
nature of drug placed at the site (lycopene/
plaoobo). Tno blinding was not brokon until
this clinical trial was completely finished.
Reproducibility
Inter- and intraexaminer reliability was test-
od using kappa statistios. Duplioato olinioal
oxaminations o 20% o tno sub|oots yioldod
kappa statistios o 94%, 92%, 93%, and
97% or PD, CAL, P, and MG, rospootivoly,
indicating good intraexaminer agreement.
Interexaminer results showed an agreement
o 85% and 90% or PD and CAL and 82%
and 86% or P and MG, rospootivoly.
Ineligible subjects
Four pationts (two in tno smokor group
and two in tno nonsmokor group) woro
excluded from the analysis because they
were found to be ineligible after randomiza-
tion. The reasons for ineligibility included
inoorroot diagnosis (ono pationt), rousal to
partioipato in tno study (two pationts), and
rousal o tnorapy (ono pationt). Tnorooro,
data analysis was restricted to 106 eligible
pationts (smokors |n = 48], nonsmokors
|n = 48], and oontrols |n = 10]).
Preparation of the 2% lycopene gel
Lyoopono (BS CHEME ntornational) was
dissolved in a solvent mixture (ethanol,
propylene glycol, and water in the ratio
o 50:30:20). Tno pH o tnis mixturo was
ad|ustod to 7.4 witn triotnanolamino. Tno
solution was tnon gollod by adding 8%
nydroxypropyloolluloso (HPC) and sot asido
for 24 hours. The in situ gel was prepared to
tno oonoontration o approximatoly 2%. Tno
drug was extensively evaluated for drug
content, viscosity determination, in vitro
permeation, and stability. The spectropho-
tometric method reported by Patel et al
19

was modified for the in vitro estimation of
the rate of drug release. The in vitro release
profile showed an initial burst release for the
first 24 hours followed by controlled release
of lycopene for up to 120 hours. The pla-
cebo was prepared in the same manner but
witnout tno aotivo drug. Tnus, ovory sub|oot
in botn tno smokor and nonsmokor groups
nad a sito troatod witn lyoopono gol (L) and
a plaoobo (P). Drugs L and P woro indistin-
guishable based on appearance.
404 VOLUME 43 NUMBEP 5 MAY 2012
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Chandra et al
Local drug delivery
Soaling and root planing was porormod at
baseline by using a piezoelectric ultrasonic
scaling unit under water spray at a medi-
um powor sotting and a 4P/4L Columbia
University curette until the root surface
was considered smooth and clean by the
operators. After scaling and root planing,
contralateral sites with the deepest probing
depths were selected, and a standardized
quantity o 0.1 mL o tno gol (2 mg/0.1 mL)
and the placebo was delivered using a
specially modified insulin delivery syringe
with a blunt-end precision tip

(Colo-Palmor)
(Fig 1). For tno purposo o maintaining
standardization, interdental suprabony
poriodontal pookots in molars witn probing
depths greater than 5 mm were selected for
looal drug dolivory. Sub|oots woro instruot-
ed to refrain from aggressively chewing,
rinsing, and brushing the test sites for a
poriod o 1 wook. Cnlornoxidino gauzo
pads

(Baotisao, Anasuya Surgioals) woro
providod to all tno sub|oots to wipo tno tost
sites to affect a palliative plaque control
rogimon. Sub|oots woro askod to roport any
possible adverse effects or unusual symp-
toms during the study period. In addition,
the periodontally affected sites in quad-
rants not involved in the study received
periodontal maintenance care, and addi-
tional nonsurgical or surgical treatment, if
required, was instituted after determining
the response to initial therapy.
Collection of GCF samples
GCF sampling was porormod at tno tost
and control sites immediately after scaling
and root planing and before placement
o tno gol. n tno oontrol group, GCF was
collected from the maxillary right central
incisor area in all the patients. After isola-
tion, 0.6-mm filter paper

(Wnatman) circles
were used for collecting the samples by the
intracrevicular method. The fluid seeping
out of the sulcus was collected, and any
paper contaminated with blood or saliva
was discarded. The collection was repeat-
ed after 30 minutes. The strip was left for 30
seconds and then immediately immersed
in 100 L o distillod wator. Samplos woro
storod at 30C and assayod on tno day o
collection.
Determination of GCF 8-OHdG
by ELISA
GCF samplos woro oontriugod at 10,000 g
for 10 minutes, and the supernatant was
usod to dotormino 8-OHdG lovols witn a
oommoroially availablo ELSA kit (Hignly
Sonsitivo 8-OHdG Cnook). Tno valuos woro
takon at basolino, 1 wook, and 3 montns.
Fig 1 Lycopene gel being delivered into a peri-
odontal pocket using a specially modifed insulin
delivery syringe with a blunt-end precision tip.
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Chandra et al
Statistical analysis
The intended primary outcome of the study
was gain in tno CAL. Tno sooondary out-
oomos inoludod roduotion in tno PD, MG,
P, and 8-OHdG lovols.
ntragroup oomparison o PD, CAL, P,
MG, and 8-OHdG lovols botwoon tno vari-
ous groups was performed using repeated
moasuros analysis o varianoo (ANOvA)
followed by multiple comparison using
Bonorroni oorrootion. Unpairod t test was
used for intergroup comparison of the vari-
ables. A P value of < .01 was considered
statistioally signiioant. Data woro analyzod
using a commercially available software
program

(SPSS 17.0, BM)
RESULTS
n tno smokor group (n = 48, oigarottos por
day, 18.10 4.33), tno moan ago o tno
sub|oots was 29.80 5.63 yoars. n tno non-
smokor group (n = 48), tno moan ago o tno
sub|oots was 33.90 7.66 yoars. Tno moan
ago o tno oontrols (n = 10) was 30.10 2.25
years.
ntragroup oomparison o PD and CAL
at baseline and 3 and 6 months showed
a statistically significant reduction in mean
sooros in smokors (lyoopono and plaoobo
groups) and nonsmokors (lyoopono and
plaoobo groups) (P < .01) witn tno di-
ferences being more significant from the
basolino to 3 and 6 montns. Howovor, tno
dooroaso in PD and CAL gain among tno
control group was not statistically signifi-
cant (P > .01) (Tablo 1).
Intragroup comparison of PI and MGI at
baseline and 1, 3, and 6 months showed
a statistically significant reduction in mean
plaquo and gingival sooros in smokors
(lyoopono and plaoobo groups) and non-
smokors (lyoopono and plaoobo groups)
(P < .01), witn tno dioronoos boing moro
significant from the baseline to 3 and 6
months. In the control group, there was
a statistically significant decrease in the
plaque scores but not for the gingival
sooros (Tablo 2).
ntragroup oomparison o 8-OHdG
lovols at basolino, 1 wook, and 3 montns
showed a statistically significant decrease
in moan 8-OHdG sooros in smokors (lyoo-
pono and plaoobo groups) and nonsmokors
(lyoopono and plaoobo groups) (P < .01),
with the differences being more significant
rom basolino to 1 wook and 3 montns. Tno
mean decrease was not statistically signifi-
cant (P > .01) in tno oontrol group (Tablo 3).
Table 1 Intragroup comparision of PD and CAL among smokers, nonsmokers, and controls
PD (mean SD) CAL (mean SD)
Group Baseline 3 mo 6 mo Baseline 3 mo 6 mo
Smokers
Lycopene
6.21 1.29 3.25 1.02* 3.50 0.92* 6.30 1.25 4.20 1.39* 4.11 1.19*
F = 131.2 P = .000 F = 28.34 P = .000
Placebo
6.02 1.49 5.20 1.70* 5.16 1.52* 6.20 1.22 5.10 0.99 5.41 1.07
F = 15.00 P = .000 F = 5.36 P = .000
Nonsmokers
Lycopene
6.04 0.87 4.00 0.58* 3.95 1.21* 6.40 1.89 4.30 1.41* 4.30 1.41*
F = 84.05 P = .000 F = 10.20 P = .000
Placebo
6.31 1.27 4.87 2.00* 4.81 1.34* 6.20 1.22 5.00 1.05* 4.80 1.13*
F = 33.88 P = .000 F = 8.23 P = .000
Controls
1.03 0.27 1.12 0.30 1.10 0.31 0.63 0.23 0.59 0.17 0.60 0.20
F = 0.35 P = .28 F = 0.54 P = .58
Comparison o moans using ANOvA.
*
Signiioant as oomparod witn tno basolino using a multiplo oomparison tost. PD, probing doptn,
CAL, olinioal attaonmont lovol, SD, standard doviation.
406 VOLUME 43 NUMBEP 5 MAY 2012
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
ntorgroup oomparison o PD, CAL, P,
MG, and 8-OHdG lovols botwoon lyoopono
and plaoobo in smokors and nonsmokors
showed a statistically significant decrease
in moans or PD and CAL gain at 3 and
6 montns and a dooroaso in tno 8-OHdG
lovols at 1 wook and 3 montns in tno lyoo-
pene group. The plaque scores and gingi-
val scores, however, were not significant
(Tablo 4).
Comparison o PD, CAL, P, MG, and
8-OHdG lovols botwoon smokors and
nonsmokors in tno lyoopono and plaoobo
groups showed no significant differences
between the two groups (P > .01) or
all variables, except for the reduction in
tno 8-OHdG lovols, wnion was statistioally
signiioant at 1 wook and 3 montns in non-
smokors wnon oomparod to smokors in tno
lycopene group (P < .01) (Tablo 5).
Comparision o tno oontrol group
8-OHdG lovols witn sitos troatod witn lyoo-
pono in smokors and nonsmokors snowod
a statistically significant difference in the
sooros at basolino in botn smokors and
nonsmokors, wnion booamo nonsignii-
Table 3 Intragroup comparision of 8-OHdG levels (in ng/mL) among smokers,
nonsmokers, and controls
8-OHdG levels (mean SD)
Group Baseline 1 wk 3 mo
Smokers
Lycopene
5.67 1.51 1.28 0.22* 1.35 0.19*
F = 95.6 P = .000
Placebo
5.66 1.52 1.73 0.34* 1.46 0.26*
F = 84.7 P = .000
Nonsmokers
Lycopene
5.34 1.14 1.04 0.32* 1.11 0.28*
F = 170.37 P = .000
Placebo
5.43 1.14 1.51 0.81* 1.49 0.25*
F = 107.36 P = .000
Control
1.65 0.16 1.38 0.20 1.30 0.17
F = 9.19 P = .14
Comparison o moans using ANOvA. *Signiioant as oomparod witn tno basolino using a multiplo oomparison tost.
8-OHdG, 8-nydroxydooxyguanosino, SD, standard doviation.
Table 2 Intragroup comparision of PI and MGI among smokers, nonsmokers, and controls
PI (mean SD) MGI (mean SD)
Group Baseline 1 mo 3 mo 6 mo Baseline 1 mo 3 mo 6 mo
Smokers
Lycopene
3.51 0.67 2.69 0.68 1.89 0.60* 1.80 0.55* 2.66 0.44 1.86 0.53 0.76 0.50* 0.75 0.50*
F = 21.02 P = .000 F = 38.87 P = .000
Placebo
3.59 0.68 2.78 0.68 2.05 0.76 1.84 0.57* 2.75 0.45 1.88 0.54 0.78 0.51* 0.82 0.54*
F = 17.27 P = .000 F = 37.52 P = .000
Nonsmokers
Lycopene
3.44 0.62 2.57 0.81 1.80 0.55* 1.55 0.63* 2.71 0.45 1.85 0.53 0.67 0.38 0.77 0.51
F = 20.60 P = .000 F = 44.34 P = .000
Placebo
3.52 0.66 2.57 0.81 1.50 0.58 1.63 0.55* 2.80 0.43 1.85 0.53 0.71 0.44 0.86 0.66
F =23.50 P = .000 F = 36.05 P = .000
Control
1.05 0.23 0.69 0.26 0.54 0.21* 0.52 0.25* 0.63 0.19 0.50 0.18 0.52 0.27 0.56 0.19
F = 18.67 P = .000 F = 1.52 P = .24
Comparison o moans using ANOvA. *Signiioant as oomparod witn tno basolino using a multiplo oomparison tost. P, Plaquo ndox,
MG, modiiod Gingival ndox, SD, standard doviation.
VOLUME 43 NUMBEP 5 MAY 2012 407
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
Table 4 Intergroup comparision of PD, CAL, PI, MGI, and 8-OHdG levels between
lycopene and placebo groups of smokers and nonsmokers
Smokers (lycopene vs placebo) Nonsmokers (lycopene vs placebo)
t P value t P value
PD Basolino
3 mo
6 mo
0.69
6.84
6.49
.49
.000
.000
1.21
2.90
3.25
.22
.005
.002
CAL Basolino
3 mo
6 mo
0.85
0.10
0.02
.65
.000
.000
0.06
4.05
3.66
.94
.000
.000
PI Basolino
1 mo
3 mo
6 mo
0.82
0.63
0.24
0.70
.65
.50
.03
.85
1.07
0.07
3.18
0.98
.28
.94
.15
.43
MGI Basolino
1 mo
3 mo
6 months
0.54
0.39
0.63
0.07
.36
.77
.79
.43
1.70
3.95
4.08
0.30
.44
> .999
.48
.53
8-OHdG Basolino
1 wk
3 mo
0.29
0.02
0.23
.67
.000
.000
5.78
1.43
2.83
.92
.000
.000
PD, probing doptn, CAL, olinioal attaonmont lovol, P, Plaquo ndox, MG, modiiod Gingival ndox, 8-OHdG,
8-nydroxydooxyguanosino.
Table 5 Intergroup comparision of PD, CAL, PI, MGI, and 8-OHdG levels between
smokers and nonsmokers in lycopene and placebo groups
Lycopene (smokers vs nonsmokers) Placebo (smokers vs nonsmokers)
t P value t P value
PD Basolino
3 mo
6 mo
0.80
4.41
2.07
.42
.18
.04
1.02
0.87
1.20
.30
.38
.23
CAL Basolino
3 mo
6 mo
0.27
0.61
0.35
.78
.53
.72
0.22
1.91
3.21
.82
.05
.34
PI Basolino
1 mo
3 mo
6 mo
1.15
0.85
0.74
0.90
.25
.39
.45
.37
0.56
1.42
2.50
0.35
.57
.15
.11
.72
MGI Basolino
1 mo
3 mo
6 mo
0.63
0.43
0.61
0.20
.53
.66
.54
.83
0.51
0.74
0.18
0.08
.60
.45
.85
.93
8-OHdG Basolino
1 wk
3 mo
1.77
3.77
4.16
.07
.000
.000
1.90
1.12
0.77
.13
.26
.43
PD, probing doptn, CAL, olinioal attaonmont lovol, P, Plaquo ndox, MG, modiiod Gingival ndox, 8-OHdG,
8-nydroxydooxyguanosino.
oant at 1 wook and 3 montns. A statisti-
cally significant difference in the mean
valuos botwoon oontrol vs smokors and non-
smokors in plaoobo-troatod sitos was soon
at all three time-bound intervals (P < .01)
(Tablo 6).
408 VOLUME 43 NUMBEP 5 MAY 2012
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Chandra et al
DISCUSSION
There seems to be an increasing volume of
evidence on the association between oxida-
tive stress and periodontitis.
20
Niootino, tno
main oomponont o tobaooo smoko, nas tno
potontial to stimulato POS roloaso, wnion
causes oxidative stress-induced tissue
damage in the gingivoperiodontal tissues.
9

At the tissue level, the effects of nicotine on
the human periodontal ligament fibroblasts
is woll-known,
21
and a recent study has
demonstrated that treatment with antioxi-
dant combinations counteracted the effects
of nicotine by restoring and increasing the
cell-migration rates of oral fibroblasts.
22
Mediators of oxidative damage such
as malondialdonydo (MDA) and SOD aro
increased in periodontitis, but most impor-
tantly, nonsurgical periodontal therapy can
rostoro and oontrol tno sub|oot antioxidant
capacity by locally and systemically modify-
ing tno lovols o MDA, SOD, and total oxida-
tivo status (TOS) in poriodontal tissuo.
23
Lycopene exhibits the highest physical
quenching rate with singlet oxygen
24
and
is at least three times more effective than
beta carotene in preventing cell death by
quononing NOO
-
radicals.
25
It also reverses
DNA damago induood by nydrogon porox-
ide.
.
26

A study investigated the relationship
between monthly tomato consumption and
serum lycopene levels in congestive heart
ailuro (CHF) individuals witn poriodontitis
and concluded that a relationship exists
botwoon poriodontitis and CHF risk, and
monthly tomato consumption appears to
affect this relationship in a positive direction
in poriodontitis sub|oots.
27
Lycopene, commonly delivered as a gel,
snows aoooptablo kinotios in tno numan
oral mucosal surfaces.
28
This drug reach-
os a poak oonoontration witnin 15 to 36
hours and returns to the baseline levels
witnin approximatoly 80 to 104 nours.
29
The
vehicle used in this study, hydroxypropyl
oolluloso (HPC), is a oolluloso dorivativo
that demonstrates both water and organic
solubility. HPC is a biodogradablo muoo-
adhesive gel used to administer a vari-
ety of compounds including antibiotics,
30

antiseptics,
31
and growth factors.
32
The
experimental gel showed good biologic
acceptability, and no evidence of burning
sensation, abscess formation, suppuration,
or staining was found.
While any antioxidant can theoretically
be used, we chose lycopene because its
levels are relatively constant and not vulner-
ablo to postsmoking doplotion as soon witn
other antioxidants.
11
Lycopene also demon-
strates intriguing nonantioxidant functions
such as reduction in the oxidative state
of macrophages and inhibition of choles-
terol synthesis secondary to the inhibition
o oollular nydroxy-3-motnylglutaryl-CoA
(HMGCoA) roduotaso.
33
The latter mecha-
nism is known to navo bonoioial ooots
on alveolar bone loss and tooth mobility in
sub|oots witn poriodontal disoaso.
34
To tno bost o our knowlodgo, tnoro
have been no studies evaluating the effi-
oaoy o 2% lyoopono gol in onronio pori-
odontitis, nonoo, a diroot oomparison witn
other studies is not possible. The effect
of systemically administered lycopene as
monotnorapy and as an ad|unot to soaling
and root planing in gingivitis patients has
Table 6 8-OHdG levels of control group compared with smokers and nonsmokers
Baseline 1 wk 3 mo
Groups t P value t P value t P value
Control vs smokors (L) 8.56 .00 0.72 .46 0.87 .29
Control vs smokors (P) 9.28 .00 4.18 .00 2.01 .00
Control vs nonsmokors (L) 11.10 .00 2.60 .12 0.78 .12
Control vs nonsmokors (P) 11.52 .00 1.29 .00 2.70 .01
L, lyoopono, P, plaoobo.
VOLUME 43 NUMBEP 5 MAY 2012 409
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
been reported, with the authors reporting
statistically significant reductions in gingi-
vitis and gingival bleeding.
1
In this study,
better response to lycopene was observed
in sub|oots witn doplotod lovols o salivary
antioxidants.
1
There is a paucity of research
on the effects of antioxidant gels in peri-
odontitis sub|oots. Provious studios did not
support the use of vitamin E gel
35
or vita-
min C supplomontation
36
for the control of
gingivitis or poriodontal disoaso. Howovor,
in one clinical study, gingival bleeding
increased significantly after a period of vita-
min C doplotion and roturnod to basolino
valuos ator vitamin C roplotion.
37
Nonsmokors witn poriodontitis domon-
stratod nignor 8-OHdG valuos tnan oon-
trols, as reported in previous studies,
4,15,38

and smokors domonstratod nignor 8-OHdG
valuos tnan nonsmokors, wnion is in agroo-
ment with most previous studies.
9,15,21,39
As
ovidonood by tno initial 8-OHdG lovols, tno
greater efficacy of lycopene can be attrib-
uted to the lower oxidative stress seen in
nonsmokors oomparod witn smokors.
9,15,21,39

Analysis o tno 8-OHdG lovols botwoon 1
wook and 3 montns sooms to suggost tnat
lycopene exerts its maximum effect within
tno irst wook and is ablo to maintain its
therapeutic action for longer periods of
timo. Tno 2% gol oan bo omployod or oon-
trolled delivery of lycopene for 7 days in the
subgingival area.
From tno data oomparing tno 8-OHdG lov-
els of the two groups with the control group, it
can be inferred that a single-dose administra-
tion of lycopene can elicit significant reduc-
tion in tno 8-OHdG lovols and nas tno ability
to decrease antioxidant damage to a level
seen in healthy tissues to confer a protective
effect on the periodontal tissues. Though not
to the same degree as lycopene, placebo-
treated sites also demonstrated a significant
roduotion in 8-OHdG lovols, tnougn tnis
could be due to the effects of scaling and
root planing, which has been shown to con-
tribute to a reduction in oxidative stress.
4,40

In the present study, the reduction in the
oxidativo stross in smokors and nonsmokors
was rolootod by gain in CAL and roduotion
in PD. Tno rosults o tno ourront study aro
in agreement with previous studies in which
the subsequent posttreatment decrease in
8-OHdG lovols nas boon assooiatod witn
gains in tno CAL and PD roduotion.
24
When
compared with the placebo, lycopene-treat-
ed sites showed a significant reduction in
probing depths and attachment loss levels
in botn tno smokor and nonsmokor groups,
but the same was not seen in the plaque and
gingival scores. In contrast to our findings,
studies in which antioxidants have been
administered systemically over a short dura-
tion have reported a statistically significant
reduction in gingivitis and bleeding index
scores but were not effective in reducing
lovols o plaquo aooumulation and PD.
1,37
Howovor, no signiioant dioronoo
was observed in the clinical parameters
between the lycopene-treated sites in the
smokor and nonsmokor groups, oxoopt or
tno 8-OHdG lovols. Smoking oausos doplo-
tion of systemic endogenous antioxidant
capacity, resulting in increased pro-oxidant
burden and tissue damage
5
and delivering
a supplemental antioxidant that in this study
might have contributed to the decrease
in PD and CAL gain in botn tno groups.
Howovor, smoking is also known to lowor
plasma concentrations of endogenous and
supplemental carotenoids,
21
but with lyco-
pono, tnoro is ovidonoo tnat smoking status
is not inversely related to circulating con-
centrations of lycopene in humans.
41
We
did find a small nonsignificant increase in
tno 8-OHdG lovols rom 1 wook to 3 montns
in lycopene-treated sites, which is probably
indicative of decreasing lycopene action.
Howovor, markors o antioxidant damago
as in this study need not accurately reflect
the overall antioxidant status of a patient.
42

The study results can be further validated
through tests such as the total antioxidant
oapaoity (TAC), wnoroin tno ull spootrum o
antioxidant activity against various reactive
oxygen radicals can be assessed.
43

It is an established fact that individuals
smoko oigarottos diorontly rom ono anotnor
and may absorb different amounts of nicotine,
ovon wnon smoking oigarottos o tno samo
brand.
44
n tnis situation, ovaluation o smok-
ers based on their serum cotinine levels rath-
er than self-reported use can investigate the
effects of antioxidant therapy more accurately
tnan sol-roporting by tno sub|oots.
44
Women
were not evaluated partly because of their low
prevalence
5
and partly because estrogen-
induood proinlammatory oytokinos, suon as
410 VOLUME 43 NUMBEP 5 MAY 2012
QUI NTESSENCE I NTERNATI ONAL
Chandra et al
intorloukin 1 bota and tumor noorosis aotor
oan also gonorato POS during monstruation
and at monopauso, tnus aooting 8-OHdG
levels.
45
Another limitation of this study is that
we have considered only heavy
46
smokors,
tno inolusion o lignt smokors witn varying
degrees of periodontitis can be considered
in future studies.
CONCLUSION
The lycopene gel formulation was effec-
tive in increasing clinical attachment and
in roduoing gingival inlammation, PD, and
oxidativo in|ury in smoking and nonsmok-
ing sub|oots. n a pationt wno is a smokor,
tno primary oous is on oossation. But ovon
with the most dedicated tobacco-cessation
programs, the relapse rate can be as high as
23%.
47
n smokors unwilling to ooaso smok-
ing or oonsidoring |oining a tobaooo-oossa-
tion program, the feasibility of using a locally
dolivorod antioxidant gol as an ad|unot to
mechanotherapy, especially during the main-
tenance phase, merits further studies.
ACKNOWLEDGMENTS
This study received no external support, but was fund-
ed by the authors institutions. The authors declare
no potential conficts of interests. The authors thank
Dr G Sudhir for the ELISA analysis of 8-OHdG and
Ms V Surekha for the preparation and evaluation of lyco-
pene and placebo gels.
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