Original Article

Comparative evaluation of locally

delivered probiotic paste and
chlorhexidine gel as an adjunct to
scaling and root planing in treating
chronic periodontitis: A split‑mouth
randomized clinical trial
Jeevanandam Vishnusripriya, Anil Melath, Mohammed Feroz, Kayakool Subair,
Nanditha Chandran

Department of Abstract:
Periodontics, Mahe Introduction: The etiological factors of periodontal diseases are the habitation of dysbiotic bacteria, absence of
Institute of Dental beneficial bacteria, and susceptibility of the host. Irresolute pattern in the periodontal diseases pathogenesis leads
Sciences and Hospital, to the evolution of novel antimicrobial therapy. Objective: The objective of the study is to assess and compare
Mahe, Kerala, India the competency of locally delivered probiotic paste with chlorhexidine gel as a supplement to scaling and root
planing (SRP) in chronic periodontitis patients. Materials and Methods: A split‑mouth randomized controlled
trial was designed on 10 systemically healthy participants having chronic periodontitis at three distinct quadrants
with 5–6‑mm pocket depth. The sites in each participant were randomly selected for Group A (negative control),
Group B (positive control), and Group C (test). In Group A, only SRP, Group B – SRP + chlorhexidine local
drug delivery (LDD), Group C – SRP + probiotic LDD were done, respectively. Gingival index (GI) and bleeding
index (BI) were determined at baseline, 3rd, 6th, and 9th weeks, whereas probing pocket depth (PPD), Russell’s
periodontal index, and clinical attachment level were checked at baseline and after 9 weeks. Results: It shows
a significant reduction in GI, BI, PPD, and gain of CAL in probiotic LDD group. Conclusion: Nowadays, since
Access this article online microbes are rapidly developing resistance to antibiotics, the development of probiotics is a boon for the treatment
of periodontal diseases. Diseases of the periodontium are not restricted to the oral cavity alone but also have
Website:
www.jisponline.com
strong systemic effects. Hence, probiotics give a natural and promising choice of therapy to establish both good
oral and systemic health.
DOI:
10.4103/jisp.jisp_704_20
Key words:
Antibiotics, formulation, local drug delivery, periodontitis, probiotics
Quick Response Code:

INTRODUCTION patient compliance, avoidance of  gastrointestinal

tract (GIT)‑related issues, bypass of first‑pass

P eriodontal disease is a chronic multifactorial

microbial infection. Conventional treatment
modalities include mechanical debridement and
metabolism by the liver, enhanced therapeutic
efficacy of the drug, and reduced treatment cost.

Address for adjunctive antibiotic administration for entire This is an open access journal, and articles are
correspondence: elimination of the oral microflora. distributed under the terms of the Creative Commons
Dr. Jeevanandam Attribution‑NonCommercial‑ShareAlike 4.0 License, which
allows others to remix, tweak, and build upon the work
Vishnusripriya, Oral microbiota include two variants, one non‑commercially, as long as appropriate credit is given and
Department of being symbiotic organisms seen in healthy the new creations are licensed under the identical terms.
Periodontics, Mahe
periodontium and the other is dysbiotic organisms
Institute of Dental Sciences For reprints contact: WKHLRPMedknow_reprints@
and Hospital, Chalakkara, seen in diseased periodontium. Antimicrobial wolterskluwer.com
Palloor, Mahe ‑ 673 310, supplements in recent years include probiotics,
Kerala, India. photodynamic therapy, onestage fullmouth How to cite this article: Vishnusripriya J, Melath A,
E‑mail: vishnusripriya.j@ disinfection, and local drug delivery (LDD). Feroz M, Subair K, Chandran N. Comparative
gmail.com evaluation of locally delivered probiotic paste
Goodson et al. in 1970[8] stated that local delivery and chlorhexidine gel as an adjunct to scaling
Submitted: 04‑Oct‑2020 and root planing in treating chronic periodontitis:
Revised: 02-Apr‑2021 system is more beneficial than systemic drug
A split‑mouth randomized clinical trial. J Indian Soc
Accepted: 24‑Jun‑2021 administration because of the following reasons:
Periodontol 2022;26:262-8.
Published: 02-May-2022 direct access to target disease, improvement of

262 © 2022 Indian Society of Periodontology | Published by Wolters Kluwer - Medknow

 Vishnusripriya, et al.: Locally delivered probiotic as an adjunct to scaling and root planning

Therefore, LDD systems have been advanced as they maintain

effective intrapocket levels of antibacterial agents for extended
period of time that can alter subgingival flora and influence
the healing of attachment apparatus.

Antibiotic resistance and recolonization by pathogenic

bacteria led to the evolution of probiotics in LDD system. Elie
Metchnikoff in 1907[15] introduced the concept of probiotics
and the term probiotics were coined by Lilley - Stillwell in
1965.[15] The current definition for probiotics is given by the
World Health Organization, which defines probiotics as
live microorganisms, most often bacteria (sometimes fungi),
which, when consumed, confer beneficial effects to the host
(World Health Organization, 2002).[15] The most commonly
used species of probiotics belong to the Lactobacillus,
Bifidobacterium, Escherichia, Enterococcus, and Bacillus genera.
These probiotic benefits oral health by defending the growth
of harmful bacteria and by modulating mucosal immunity Figure 1: Formulated paste (F1-F5) with different concentrations of glycerine
in the oral cavity.[16] Therefore, this study was designed to
formulate a probiotic‑based LDD material evaluating its in vitro
pharmaceutical properties and to compare its clinical efficiency
with chlorhexidine gel as a supplement to scaling and root
planing in the treatment of chronic periodontitis patients.

MATERIALS AND METHODS

The in vitro pharmaceutical maneuvering of the novel probiotic

LDD material was done under standardized protocol in Manipal
College of Pharmaceutical Science, Manipal. Pharmaceutical
armamentarium includes probiotic powder (Sporlac*),
edible glycerol, glass slab, mixing spatula, measuring scale.
Formulation of Lactobacillus sporogenes  (250  mg) in five
different proportions of glycerol (0.1 ml, 0.2 ml, 0.3 ml, 0.4 ml,
and 0.5 ml) was prepared by taking Lactobacillus sporogenes
powder and glycerol on a glass slab, and the powder was
mixed with glycerol in geometric proportions until smooth
Figure 2: pH determination
consistency paste was obtained [Figure 1]. The formulated
pastes were then evaluated for various parameters.

The pH of the drug solution in 3 different solvents,

i.e., water, glycerol, and methanol was determined using pH
meter [Figure 2]. The pH values were found ranging from
3 to 6.5. The viscosity of all 5 formulations was determined
using Brookfield viscometer at 0.01  rpm and temperature
of 27°C. The viscosity of all 5 formulations was identified
to be >30,650 centipoises [Figure 3]. Spreadability of the
formulations was evaluated by measuring the spreading
diameter of 1 g of sample placed between two horizontal
glass plates after 1 min. The standard weight applied to the
upper plate was 25 g. It was found that with increasing the
amount of glycerol, the spreadability of the formulation
increased [Figure 4]. Homogeneity and texture were tested
by pressing a small quantity of the formulation between the
thumb and index finger. The consistency of the formulations
a b
and the presence of coarse particles were used to determine
the texture and homogeneity of the formulations. All the five Figure 3: Brookfield viscometer (a) and (b)
formulations (F1‑F5) showed that the formulated paste had
appealing appearance and smooth texture, and they were all shows maximum absorption. λmax of LS was determined in
homogenous as indicated by the absence of gritty particles. different solvents such as glycerol and phosphate buffer (pH‑6.8)
and it was found to be 208 nm and 205 nm, respectively [Figure 5].
Lambda max (λmax) value is the wavelength at which a substance The stock solution (1000 mg/ml) of Lactobacillus sporogenes was
has its strongest photon absorption. At this wavelength, the drug

Journal of Indian Society of Periodontology - Volume 26, Issue 3, May-June 2022 263
 Vishnusripriya, et al.: Locally delivered probiotic as an adjunct to scaling and root planning

a b
Figure 4: Spreadability test indicating (a) original diameter before addition of
weights and (b) increased diameter after addition of 25 g weights Figure 5: Ultraviolet-visible spectrum of LS in glycerol. LS – Lactobacillus
sporogenes; Abs – Absorbance

Figure 6: Calibration curve. y – linear regression calculated;

r2 – Correlation coefficient

prepared, and further dilutions were made in the concentration Figure 7: Vertical diffusion cell
range of 50–500 mg/ml using phosphate buffer pH‑6.8 as diluent.
All samples were analyzed by ultraviolet (UV) spectrophotometer participants from the department of periodontics who were
by measuring the absorbance at 205 nm with phosphate buffer diagnosed as systemically healthy having chronic periodontitis
pH 6.8 used as a reference standard. The absorbance readings in at least three different quadrants with 5–6 mm pocket depth
obtained were plotted against concentration to get standard were selected for the study.
calibration curve of Lactobacillus sporogenes. Slope of graph
was calculated for estimating the drug released in receptor Inclusion criteria for the study were patients with chronic
compartment during diffusion studies using UV/visible generalized periodontitis (AAP 1999 classification), probing pocket
spectroscopic method [Figure 6]. In vitro drug release studies depth (PPD) of 5–6 mm (all the surface of the tooth was evaluated
were carried out by using vertical diffusion cell with a nominal and the highest measure of the pocket depth was considered for
volume of the acceptor compartment of 20 ml of phosphate categorizing the tooth for respective group), and the exclusion
buffer (pH 6.8). Cellophane membrane was fixed between donor criteria include patients undergone any periodontal therapy
and receptor compartment. In the donor compartment, 250 mg previously, systemic disease or medication, patients with any
of formulation was added. The experiments were conducted bone disorders, vitamin D deficiency, postmenopausal women.
at temperature of 37°C and 900 rpm for 12 h. Samples were
withdrawn from the receptor compartment at different time Clinical armamentarium includes chlorhexidine
points, and the amount of drug released in receptor solution gel (Hexigel*‑1.0%w/w), probiotic formulation
was estimated by using UV spectrophotometer at 205 nm. The (Sporlac * Powder + Glycerol), glass slab, mixing spatula,
image of the diffusion cell used in the present study is shown plastic filling instrument, coie‑pack. Three sites in each
in Figure 7. The drug release from the formulated paste was by participant were randomly selected as Group A (control),
diffusion mechanism and was found to be increased, and almost Group B (positive control), and Group C (test).
100% release was observed at the end of 12 h.
Parameters analyzed at baseline, 3 weeks, 6 weeks, and
The present study was a split‑mouth randomized clinical trial 9 weeks are gingival index (GI) (Loe and Silness) and bleeding
conducted under standardized protocol after obtaining ethical index (BI) (mSBI Mombelli 1987) and at baseline and after
clearance by the college ethical clearance committee, Mahe 9 weeks are PPD, Russell’s Periodontal Index (RPI), and clinical
Institute of Dental Sciences and Hospital, Mahe. A total of 10 attachment level.

264 Journal of Indian Society of Periodontology - Volume 26, Issue 3, May-June 2022
 Vishnusripriya, et al.: Locally delivered probiotic as an adjunct to scaling and root planning

The baseline parameters were recorded after obtaining consent three groups with significant improvement for GI in Group B
from the patient for participating in the study, and periodontal from score 3.1 to 2.2 and in Group C from score 3.4 to 1.9 and
therapy was done based on the group allotment. in case of BI in Group B from score 2.9 to 2.2 and in Group C
from score 3 to 1.9, respectively.
Group A was advised SRP alone, Group B was advised
SRP + LDD (chlorhexidine gel), and Group C was advised In case of PPD [Tables 5 and 6], RPI [Tables 7 and 8] and clinical
SRP + LDD (probiotic LDD material). All the data were attachment level [Tables 9 and 10] measured at baseline and
statistically analyzed using SPSS is Statistical Package after 9 weeks showed gradual gain in the clinical attachment
for the Social Sciences. It is a software used for statistical level with significant improvement for PPD in Group B from
analysis of the present study. Descriptive analysis was done 5.6 mm to 4.2 mm and in Group C from 5.6 mm to 3.9 mm, for
and independent sample t‑test was employed between the RPI in Group B from score 3.3 to 2.2 and in Group C from score
groups. Two‑way analysis of variance tests was applied for 3.5 to score 1.9, finally in case of CAL in Group B from 1.4 mm
intergroup comparison. For all tests, a P < 0.05 was considered to 1.2 mm and in Group C from 1.5 mm to 1.2 mm, respectively.
as statistically significant.
DISCUSSION
RESULTS
The prevailing concept for etiology of periodontal disease
The results for the present investigation were estimated based includes three main factors – a susceptible host, the presence
on the parameters analyzed. The GI score [Tables 1 and 2] of pathogenic species, and the absence of socalled “beneficial
and BI score [Tables 3 and 4] were assessed at base, 3 weeks, bacteria which determine whether an active periodontal disease
6 weeks, and 9 weeks showed gradual improvement in all will occur.”[1]

Table 1: Mean gingival index score (Loe and Silness)

Site (tooth Baseline 3rd week 6th week 9th week Guidelines
number)
Group A (SRP) 3 2.8 2.6 2.6 0. Normal
Group B (CHX) 3.1 2.6 2.4 2.2 1. Mild inflammation, no bleeding
Group C (probiotic) 3.4 2.4 2.4 1.9 2. Moderate inflammation, bleeding on probing
3. Severe inflammation, spontaneous bleeding
SRP – Scaling and root planing; CHX – Chlorhexidine

Table 2: Intergroup comparisons of gingival index

Groups Mean SD Comparison group Mean difference P 95% CI
Lower bound Upper bound
A 2.75 0.157 B (CHX) 0.175 0.439 −0.282 0.632
C (probiotic) 0.225 0.321 −0.232 0.682
B 2.58 0.157 A (control) −0.175 0.439 −0.632 0.282
C (probiotic) 0.050 0.824 −0.407 0.507
C 2.53 0.157 A (control) −0.225 0.321 −0.682 0.232
B (CHX) −0.050 0.824 −0.507 0.407
SD – Standard deviation; CI – Confidence interval; CHX – Chlorhexidine; P – Probability value; P < 0.05 was considered as statistically significant

Table 3: Mean bleeding index score (mSBI Mombelli 1987)

Site (tooth Baseline 3rd weeks 6th week 9th week Guidelines
number)
Group A (SRP) 2.8 2.7 2.6 2.6 0. No bleeding when probe is passed along marginal gingiva
Group B (CHX) 2.9 2.6 2.4 2.2 1. Isolated bleeding spots visible
Group C (probiotic) 3 2.4 2.4 1.9 2. Blood from a confluent red line on margin
3. Heavy or profuse bleeding
SRP – Scaling and root planing; CHX – Chlorhexidine

Table 4: Intergroup comparisons of bleeding index score

Groups Mean SD Comparison group Mean difference P 95% CI
Lower bound Upper bound
A 2.68 0.132 B (CHX) 0.150 1.000 −0.326 0.626
C (probiotic) 0.250 0.574 −0.226 0.726
B 2.53 0.132 A (control) −0.150 1.000 −0.626 0.326
C (probiotic 0.100 1.000 −0.376 0.576
C 2.43 0.132 A (control) −0.250 0.574 −0.726 0.226
B (CHX) −0.100 1.000 −0.576 0.376
CI – Confidence interval; SD – Standard deviation; P – Chlorhexidine; CHX – Probability value; P < 0.05 was considered as statistically significant

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Table 5: Mean probing pocket depth

Site (tooth number) Baseline 9th week Guidelines
Group A (SRP) 5.5 4.9 At baseline ‑ 5-6 mm
Group B (CHX) 5.6 4.2 Six points probing (mesiobuccal, midbuccal, distobuccal,
Group C (probiotic) 5.6 3.9 mesiolingual, midlingual, distolingual)
SRP – Scaling and root planing; CHX – Chlorhexidine

Table 6: Intergroup comparisons of probing pocket depth

Group Mean SD Comparison group Mean difference P 95% CI
Lower bound Upper bound
A 5.20 0.179 B (CHX) 0.300 0.737 −0.345 0.945
C (probiotic) 0 0.259 −0.195 1.095
B 4.90 0.179 A (control) −0.300 0.737 −0.945 0.345
C (probiotic) 0.150 1.000 −0.495 0.795
C 4.75 0.179 A (control) −0.450 0.259 −1.095 0.195
B (CHX) −0.150 1.000 −0.795 0.495
CI – Confidence interval; SD – Standard deviation; P – Probability value; CHX – Chlorhexidine; P < 0.05 was considered as statistically significant

Table 7: Mean Russell’s periodontal index score

Site (tooth number) Baseline 9th week Guidelines
Group A (SRP) 3.1 2.7 Negative. There is neither overt inflammation in the investing tissues nor loss of function due
Group B (CHX) 3.3 2.2 to destruction of supporting tissue
Group C (probiotic) 3.5 1.9 Mild gingivitis. There is an overt area of inflammation in the free gingivae which does not
circumscribe the tooth
Gingivitis. Inflammation completely circumscribes the tooth, but there is no apparent break in
the epithelial attachment
Gingivitis with pocket formation. The epithelial attachment has been broken and there is a
pocket (not merely a deepened gingival crevice due to swelling in the free gingivae). There
is no interference with normal masticatory function, the tooth is firm in its socket, and has not
drifted
Advanced destruction with loss of masticatory function. The tooth may sound dull on
percussion with a metallic instrument; may be depressible in its socket.
SRP – Scaling and root planing; CHX – Chlorhexidine

Table 8: Intergroup comparisons of Russell’s periodontal index

Group Mean SD Comparison group Mean difference P 95% CI
Lower bound Upper bound
A 2.90 0.173 B (CHX) 0.150 1.000 −0.476 0.776
C (probiotic) 0.200 1.000 −0.426 0.826
B 2.75 0.173 A (control) −0.150 1.000 −0.776 0.476
C (probiotic) 0.050 1.000 −0.576 0.676
C 2. 0.173 A (control) −0.200 1.000 −0.826 0.426
B (CHX) −0.050 1.000 −0.676 0.576
CI – Confidence interval; SD – Standard deviation; P – Probability value; CHX – Chlorhexidine; P < 0.05 was considered as statistically significant

Table 9: Mean clinical attachment level

Site (tooth number) Baseline 9th week Guidelines
Group A (SRP) 1.5 1.5 The CAL is the measurement of the position of the soft
Group B (CHX) 1.4 1.2 tissue in relation to the CEJ that is a fixed point
Group C (probiotic) 1.5 1.2
CAL – Clinical attachment level; CEJ – Cementoenamel junction; SRP – Scaling and root planing; CHX – Chlorhexidine

Table 10: Intergroup comparisons of clinical attachment level

Groups Mean SD Comparison group Mean difference P 95% CI
Lower bound Upper bound
A 1.50 0.144 B (CHX) 0.200 1.000 −0.321 0.721
C (probiotic) 0.150 1.000 −0.371 0.671
B 1.30 0.144 A (control) −0.200 1.000 −0.721 0.321
C (probiotic −0.050 1.000 −0.571 0.471
C 1.35 0.144 A (control) −0.150 1.000 −0.671 0.371
B (CHX) 0.050 1.000 −0.471 0.571
CI – Confidence interval; SD – Standard deviation; P – Probability value; CHX – Chlorhexidine; P < 0.05 was considered as statistically significant

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Subgingival debridement, surgical interventions, and BI, which are in accordance with the reported observations
supplemental use of antibiotics and antiseptics are contemporary of Krasse et al.,[2] Noordin and Kamin,[4] Twetman et al.,[5] and
methods to decrease the pathogenic bacteria. According to Vivekananda et al.,[6] where they found a remarkable decrease
Quirynen et al., 2002; Teughels et al.,[7] 2011, these treatments in gingivitis scores with the use of probiotics along with scaling
result in a temporary decline in the bacterial load and associated and root planing (SRP).
inflammation and are often not sufficient to control the disease.
Therefore, the administration of beneficial bacteria, probiotics Further, in the current study, a significant reduction in
with antimicrobial and anti‑inflammatory properties, is one of periodontal pocket depth and improvement in clinical
the novel supplemental approaches for periodontal therapy. attachment level were found, which are in agreement with
The mechanism of probiotics in humans involves direct the reported observations of Vicario et  al.,[9] Ince et  al.,[10]
interaction, competitive exclusion, and modulation of host Tekce et al.,[14] Laleman et al.,[12] suggesting the potent role of
immune response.[3] probiotic‑based LDD paste as an adjunct to SRP in chronic
periodontitis patient.
Various studies have been published investigating the
effective health benefits of probiotics on systemic health, In the present study, using a negative (SRP alone) and a
but investigations regarding their use in oral health are positive control (chlorhexidine gel), we were able to state
limited. These vary in terms of probiotics strains used, its that the probiotic LDD (Sporlac) has shown to be a potential
concentrations, and vehicles for the application including LDD material in comparison to chlorhexidine. Chlorhexidine
cheese, lozenges, milk, kefir, ice cream, gum, drops, powder, seldom caused local side effects including brown staining,
and mouthwash.[7] taste alterations, increased supragingival calculus formation,
and rarely desquamation of the oral mucosa, irrespective of
In periodontal disease, studies on the role of probiotics in which type of vehicle is used.[13] Sporlac probiotic LDD paste
gingivitis reported a significant decrease of plaque and gingival shows a very effective and economical substitute for patients
indices, bleeding on probing, and gingival inflammation in with periodontal disease
the probiotic groups.[11] The results of the animal and clinical
periodontitis studies support the concept of probiotics in the CONCLUSION
management of periodontitis and thereby offer a low‑risk,
inexpensive, easy‑to‑use prevention, or treatment option for Sustained drug release devices are proving to be more effective,
treating periodontal disease. more convenient, and easier to use than regular systemic
administration of medicines. This development definitely
Glycerol is a colorless, odorless, and sweet compound with paves the way for future trademarking of novel, economically
cryoprotectant activity. Based on a study conducted by Hafiz feasible, and physiologically acceptable intrapocket‑targeted
Shehzad Muzammil, Barbara Rasco and Shyam Sablani, drug delivery systems.
Glycerol supplementation with probiotic frozen yogurt,
increased the stickiness from 2.4% to 18.7%, and decreased the The multifactorial and complex microbial nature of periodontal
hardness from 8.0% to 14.5%, respectively.[16] and peri‑implant infections, consisting of both erobic and
anerobic bacteria, renders difficulty in predicting all the
The present investigation focused primarily on the formulation restrictions with the clinical use of drug formulated in the
of a paste incorporating probiotic and glycerol, with sustained present study. However, based on the positive results from
drug release capacity, ideal pH, viscosity, homogeneity, and in vitro pharmaceutical analysis (the pH is suitable for use in
spreadability for ease in delivering the formulation into the oral cavity, viscosity was suitable for a paste material with
pocket. adequate spreadability, homogeneous and smooth texture,
and the drug was completely absorbed in 12 h) and the clinical
After testing various concentrations and formulations, we trial (significant reduction in GI, BI, PPD, and gain of CAL) of
developed a mix of materials that represent satisfactory the drug formulated in the present study suggest a definitive
pharmaceutical property for a material to be used as LDD platform for further welldesigned clinical studies with larger
material. The formulation with 250 mg of probiotic powder sample sizes, administering a booster dose and long‑term
and 0.1 ml of edible glycerol was found to have ideal properties follow‑ups.
for a LDD material. The formulated drug is professionally
applied (in dental office) biodegradable material with sustained Financial support and sponsorship
drug release ability. These are delivery systems whose action Nil.
lasts for <24 h and therefore require multiple applications. It
follows the first‑order kinetics.[8] Various drug delivery systems Conflicts of interest
are available for treating periodontitis, the material prepared There are no conflicts of interest.
in our study is a paste.
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268 Journal of Indian Society of Periodontology - Volume 26, Issue 3, May-June 2022