Evaluation of Polyphenolic Profile and Antibacterial Activity of Pomegranate Juice in Combination with Rifampin (R) against MDR-TB Clinical Isolates
Authors: AlMatar Manaf , Var Işıl , Kayar Begüm , Eker Emel , Kafkas Ebru , Zarifikhosroshahi Mozhgan , Köksal Fatih 
Published: March 2019

Background:
The global rise of multi-drug resistant M. tuberculosis demands unconventional
treatment to enhance the efficiency of current drugs. Punica granatum, which is known as pomegranate,
is considered as a member of the Punicaceae family. Pomegranate, which is broadly documented for its
activity against a wide spectrum of bacterial pathogens, deserves further scrutiny in this respect.

Methods:
Within this scope, this study investigated the effect of fresh pomegranate juice (FPJ) on the
antibacterial activity of anti-tuberculosis drugs (Rifampin (R) and Isoniazid (INH)) against MDR-TB
clinical isolates. The drug resistance profiles in M. tuberculosis clinical isolates were determined by
susceptibility test using BACTEC MGIT 960 system. Four concentrations of fresh pomegranate juice
(FPJ) (5%, 10%, 15%, and 20%) were evaluated in combination with R and INH at a dose range of
(1.0 µg/ml) and (0.1 µg/ml), respectively against the MDR-TB isolates by the BACTEC MGIT 960
system. Moreover, this study scrutinized individual phenolic compounds of FPJ by using highperformance
liquid chromatography (HPLC). The total polyphenols (TP), total flavonoid (TF), total
anthocyanins content (TAC), and the antioxidant capacity were also assessed in FPJ.

Results:
Synergistic effects were observed between R and INH with FPJ against all tested strains.
However, combination therapy of rifampin was more effective than isoniazid one. Therefore, the combination
of R and FPJ has been used against (27) MDR-TB clinical isolates. 5% of FPJ plus R (1.0
µg/ml) were found to suppress the growth of one isolates for first group (INH and R resistant). However,
5% of FPJ demonstrated no synergistic impact with R for second (SM, R and INH resistant) and
third group (INH, EMB, R and SM resistant). Moreover, 10% of FPJ and R (1.0 μg/ml) inhibited the
bacterial growth of three isolates of first group and two isolates and one isolate for second and third
group, respectively. Remarkably, 15% of FPJ plus R (1.0 µg/ml) appeared to inhibit the growth of
MDR-TB isolates for all tested groups indicating a strong synergistic effect. Regarding H37RV, the
complete inhibition of the bacterial growth was found to occur at 15% and 20% concentrations of FPJ
only. Minimum inhibitory concentration (MIC) of FPJ ranged from (4% to13%) for first group and
from (10% to15%) for second and third group. Thus, FPJ at 15% inhibited 100% of bacteria for all
tested isolates (MIC100% =15%). Phenolic compounds identified in FPJ were gallic acid, benzoic
acid, syringic, folic acid, pelargonidin, naringin+ellagic acid, naringenin, chlorogenic acid, caffeic
acid, catechin, myricetin, kaempferol, quercetin, cyanidin-3-glycoside, p-cummaric acid, ferulic acid,
and rutin. Total phenolic (TP), total flavonoid (TF), and total anthocyanin (TA) content were 841.5 mg/L,
638.73 mg RE/L, and 47.43 mg/L, accordingly.

Conclusion:
Overall, FPJ displayed synergistic effect with R against MDR-TB clinical isolates due to
its high content of polyphenol and antioxidant capability.