Microorganisms and culture conditions
This study was conducted in a tertiary care hospital in Chennai over a period of 1 year. The present study enrolled 160 patients attending the Obstetrics and Gynaecology Out patients department and Inpatients admitted in the wards with symptoms suggestive of vaginitis. The study has been approved by institutional ethical committee and Informed consent was obtained from all the subjects.

Sterile vaginal swabs were used for collection of vaginal samples from the patients.  Two high vaginal swab samples were collected aseptically from the posterior vaginal fornix using speculum and posterior vaginal wall retractor. The swabs were transferred to the microbiology laboratory and processed immediately.

One of the swabs was used for Gram stain and direct wet mount microscopy using 10% Potassium hydroxide solution to determine the presence of yeast cells in the sample. The second swab was streaked on Sabouraud Dextrose agar (SDA) ( HiMedia, India )  plates containing chloramphenicol and incubated at 37^oC for 24-48 hours. The colonies from SDA plate were subjected to Gram stain to confirm the growth of candida and were further subtyped by streaking on Hichrome Candida differential agar ( HiMedia, India ) and incubated at 37^oC for 24-48 hours. The pigmented colonies were further examined for assimilation of various sugars and candida species were identified using standard microbiological methods. The colonies from the SDA plate were again processed in automated VITEK2 compact (bioMérieux) and the results were documented.

The colonies from SDA plate were also subjected to germ tube test to check for the production of germ tubes. Germ tube test is done by mixing 3-4 colonies in 0.5 ml human serum and incubated at 37^oC for 2-4 hours and examined under microscope for the formation of germ tube.  Colonies suggestive of C.albicans were confirmed by this germ tube test.
Preparation of Cyperus rotundus
C. Rotundus were bought from Alkadhemia city, Baghdad. The tubers were washed with water, air dried and cut into strips. A quantity of 300 g Cyperus rotundus and 1000 mL water were mixed and the mixture blended using home blander. The mixture was then filtered using a cotton cloth filter, autoclaved at 121°C for 30 min.

Fermentation process
The initial pH of the mixture from Section 100 mL was inoculated with 10% (v/v) of L. plantarum and incubated at 37°C for 48 h. The cells counts and pH values were determined after 48 h of fermentation. Non-fermented samples were subjected to same conditions but without starter culture.

Antioxidant Activity of Fermented Cyperus rotundus
DPPH Assay
The Radical Scavenging Activity (DPPH) determines how potent the antioxidant capacity of the fermented mixture⁸. The assay was performed based on the method from previous study using 2, 2-diphenyl-1-picrylhydrazyl radicals⁹. Briefly, in 96 wells plate, 0. 25 mL of C. rotundus extract was mixed with 1. 75 mL DPPH solution that was prepared by diluting DPPH in methanol. The plate was incubated in dark place at room temperature for 30 min and absorption was taken at 515 nm using microtiter plate reader (Biotek, EL 800) and DPPH percentage was calculated as the following:

% DPPH = [(control 515 – sample 515) / control 515] *100

FRAP Assay
Ferric reducing antioxidant power (FRAP) assay is a method used to estimate the antioxidant capacity of the fermented mixture that count on reducing the ferric-tripyridyltriazine¹⁰. FRAP reagent was prepared by mixing 2, 4, 6-tripyridyl-s-triazine (TPTZ) and FeCl3 solutions in (pH 3. 6) buffer acetic acid at the ratio of 1: 1: 10 (v / v / v). In 96 microtiter plate, 20µL sample was mixed with FRAP reagent (200µL) and incubated for 30 min at room temperature. The absorbance was measured at 593 nm and the calculations were determined using different concentrations from 0. 1 to 1 mM of ferrous sulfate as standard, and expressed as mmol Fe (II) / g DW.

Antibacterial Activity of Fermented Cyperus rotundus
The antibacterial activity of the fermented mixture was determined in following the method described by Aween et al.,¹¹. The fermented mixture was tested against Pseudomonas aeruginosa ATCC10415, Bacillus subtilis ATCC11778 and Escherichia coli ATCC11229. Fermented mixture 100 µL was added to the wells and 100 µL nutrient broth containing 10⁶ of each selected pathogens. The control was nutrient broth with the pathogenic bacteria only. The plates were incubated at 37°C for 24 h and the growth inhibition was measured at 600 nm by using microtiter plate reader. The antibacterial activity was calculated by subtracting the 0 h readings from the 24 h readings, then using the following mathematical equation.

¹H-NMR metabolomics analysis
The method used for the extraction was adopted from Abas et al., (2013)¹². Ten mg of the fermented and samples were mixed with CH3OH-d4 (0. 375 mL) and KH2PO4 buffer (0. 375 mL) in D2O (pH 6) containing 0. 1 % TSP as standard. The mixed samples were subjected to sonication for 15 min at 30°C. The samples were centrifuged at 13,000 rpm for 5 min and 600µL transferred into NMR tubes¹³. The NMR works on the INOVA 500 MHz variable spectrometer and at 499. 887 MHz at room temperature 26°C with 64 scans were done on every sample and verified, the acquisition time was 193 s, 3. 75µL pulse width and 1. 0 sec relaxation delay. Each sample was measurement was repeated six times. The residual water region (d 4. 70 to 4. 96) and methanol region (d 3. 28 to 3. 33) were excepted from the analysis.

Statistical analysis
MINITAB version 16 was used to analysis the Data. Statistical measures were used to evaluate the results of these samples and their controls, where one-way analysis of variance (ANOVA). The multivariate analysis was done using SIMCA-P software (Umetrics, Sweden).