Effect of fermentation conditions on antioxidative, antimicrobial and techno-functional properties of Camellia sinensis leaves

Introduction and Aim: Fermented food is an excellent source of nutrition as it contains promising bioactive compounds. Black tea is a fermented product and the most commonly used beverage. In the present study, the effects of various fermentation conditions and drying techniques on the proximate nutritional analysis, phytochemicals, tannin, antioxidative, and antimicrobial activity of Camellia sinensis leaves were examined. Materials and Methods: The FTIR analysis was performed to detect the presence of different functional groups. Using standard methods, total phenol, flavonoid, and tannin content were determined. The antioxidant activity was determined by using DPPH, ABTS, and FRAPS assays. Results: The results showed a decrease in protein content and an increase in ash, moisture, and crude fiber content, indicating that the samples’ nutritional profiles were improved after fermentation processing. The FTIR results showed the presence of hydroxy, carboxyl, amine, alkane, alkene, and alkyne bonding. The antioxidant activity of fermented samples was more than that of unfermented ones. Fermented samples had strong antimicrobial activity against Escherichia coli . An increase in TPC and TFC contributed to the rise in the antioxidant activity of the fermented sample. An increase in TSS suggested improvement in flavour and sweetness. Antimicrobial activity against Escherichia coli , demonstrated their potential as a natural antimicrobial agent. Conclusion: The enhanced antimicrobial activity of fermented tea leaves highlights the importance of the fermentation process in producing tea with improved microbial control and possible health benefits.


INTRODUCTION
ea, a well-known and widely consumed beverage worldwide, is obtained from the plant Camellia sinensis.It belongs to the Theaceae family and is an herbaceous deciduous plant of two varieties Camellia sinensis sinensis from China, also known as the China variety, and Camellia sinensis assamica from Assam, known as the Assam variety (1).This plant is cultivated in parts of southeast Asia, some parts of Africa, and South America.China is the largest tea producer, while India occupies second place in the largest tea-producing countries.In India, Assam and Darjeeling contribute more to producing good-quality tea.The Indian tea variety requires 2000 to 4000 mm of average annual rainfall with a temperature of 28°C to 32°C.It grows well in soil with pH ranges from 4.5 to 5.5 on highland, welldrained soils.In northern India, tea production accounts for around 83% of annual tea production, while the rest,17%, is from states like Kerala, Karnataka, and Tamil Nadu (Tea Research Association).Tea leaves are processed through different methods to produce various types of tea.These are yellow tea, black tea (fully fermented), green tea (unfermented), oolong tea (semi-fermented), dark tea (post-fermented), and white tea (moderately fermented) (2).Tea production involves several processing stages, such as harvesting tea leaves, withering to remove some moisture, rolling using the CTC (Crush Tear Curl) method, hand rolling or using machine rollers, fermentation/ oxidation (exposing to oxygen spontaneously), and drying to increase shelf life (3).Withering can be done for 18-20 hours; spontaneous fermentation takes about 2-3 hours, though this time can be modified to produce different types of fermented tea.The fermentation of tea is an enzymatic oxidation.It begins during one of the processing steps of tea leaves: maceration and rolling.During rolling, the cells of the leaves rupture and expose their chemical constituent over the surface of the leaves, which gets oxidized by indigenous enzymes such as PPO (Polyphenol Oxidase) and PO (Peroxidase) in the presence of oxygen present in the atmosphere (4).The composition of chemical constituents depends upon environmental factors, cultivation, and different processing techniques.Flavonoids, phenolic acids, tannins, phytosterols, and terpenoids are the major bioactive components in tea leaves.Flavonoids have different subclasses: flavonols, flavanones, isoflavonoids, flavones, T anthocyanins, and flavanols (flavan-3-ol) (5).Quercetin, kaempferol, and myricetin are the majorly occurring flavonols.Catechins in tea belong to flavanols, the major bioactive component of tea leaves.Catechins are also of different types-EGCG (epigallocatechin-3-gallate), ECG (epicatechin-3gallate), EC (epicatechin), EGC (epigallocatechin), etc.These catechins are water-soluble, colourless components that provide aroma, bitterness, and astringency to tea (6).
Tea has beneficial effects in preventing and protecting several perilous diseases like diabetes, cancer, cardiovascular diseases (CVDs), and neurodegenerative diseases.The polyphenols of tea have been used to prevent these diseases.They are also effective in weight loss, anti-allergic, as prebiotics, and to avoid osteoarthritis (7).Theanine found in tea has a role in maintaining the integrity of T-cell mediated immune response.Theobromine enhances heart activity, plays a role in vasodilation, and is a more effective diuretic.Catechins are potent antioxidants; they scavenge free radicals and thus help reduce cancer.ECG interacts with lymphocytes, thus reinforcing the immune system.Catechins also play a role in lowering vascular oedema as they are used to treat capillaropathy.They also have antibacterial properties.Flavanols help in the regulation of thyroid gland activity.Anthocyanidins enhance vision, prevent hypertension and atherosclerosis, and strengthen capillaries and joints.Phenolic acids in tea have anti-inflammatory activity.They also improve the anti-toxic action of the liver.As tea is taken worldwide more frequently and highly, it occupies second place after water in beverages.Its fermentation enhances the properties due to the change in its bioactive chemical components, which directly or indirectly has several beneficial effects on human health (8).

Sample collection and processing
The tea plant Camellia sinensis leaves were collected from West Bengal and Assam during March and April.The collected sample was washed with tap water, and further processing was done (Fig. 1 and Fig. 2).
The tea leaves were left for natural withering for 18 hours.After the withering process, leaves were handrolled.Then, for fermentation, different batches of leaves were prepared.From the West Bengal (WB) variety, some tea leaves were fermented at a controlled condition of 30˚C for 2 hours in an incubator.The other batch was left at room temperature for spontaneous fermentation/-oxidation.For the Assam variety, tea leaves were spontaneously fermented at room temperature.The fermented tea leaves were then dried under different conditions.Out of WB incubator fermented (IF) and spontaneously fermented (SF) samples, some were dried at a temperature of 105˚C in an oven, and others were sundried.The dried tea leaves were then ground into fine powder for further analysis.The Assam variety was sundried only.The fresh tea leaves were not fermented; they were only washed and dried under natural sunlight and ground into fine powder.By this, the following batch of samples was prepared: • WB Fresh (West Bengal Fresh-unfermented)

Liquid extract preparation
The aqueous extract was prepared by dissolving 1g of powdered sample in 10 mL of distilled water, and the mixture was shaken for 30 minutes.The extract was filtered using Whatman no.42 filter paper, and the filtrate was used as an aqueous extract for further analysis.

Alcoholic extract preparation
1g of powdered tea sample was infused with 10 mL of 80% methanol.This mixture was kept in a shaker for 3 hours and centrifuged at 9000 rpm at 4°C for 10 minutes.The supernatant was collected, labelled methanolic extract, and used for further analysis.

Moisture content
Weighed the crucible precisely, added 1 g of sample, and weighed it again.After that, the crucible was dried for 3 hours at 105 °C in a hot air oven (Fig. 3).Then, it was taken out of the oven, placed in a desiccator, and weighed.Finally, repeat drying procedures until the constant is reached (9).The following formula was used to calculate moisture percentage:

Ash content
A crucible was filled with 1g of sample, which was weighed, and the sample was burned inside the crucible until no smoke was produced (Fig. 3).After cooling the sample, it was put in a muffle furnace and kept there for 6 to 8 hours at 550-600°C until white ash was produced.The sample was then cooled into a desiccator and weighed.The following formula was used to calculate the ash percentage:

Crude fiber content
The acid-base digestion method was used for the determination of crude fiber.1.25% H2SO4 solution was prepared for acidic digestion, and 1.25% NaOH solution was used for alkaline digestion.2g of powdered sample was added to a beaker containing 200 mL of sulfuric acid solution and heated on a hot plate until it started boiling.After that, the sample was chilled and filtered using muslin cloth, and the material was washed three times using distilled water.The remaining sample content was then transferred to another beaker, and 200 mL of sodium hydroxide solution was added for basic digestion.This was boiled for 30 minutes, kept for cooling, and then strained using a muslin cloth.The remaining solid was kept in an empty, dried crucible and weighed.The sample's moisture was removed in a hot air oven at 105°C for 2 hours until constant weight was achieved.This sample contained organic and inorganic matter.
Then the sample was put in a muffle furnace for 4 hours for ashing.The ash formed was cooled down in a desiccator, and weight was measured.

Protein content
Protein content was determined by using Lowry's method as described by Mahesha (10).For this, four reagents were made.Reagent A was made using 2 % sodium carbonate in 0.1 N NaOH.Reagent B was freshly prepared using 0.5% copper sulphate in 1 % potassium sodium tartrate.Reagent C was an alkaline copper solution made by mixing 1 mL of B reagent and 50 ml of A reagent.Reagent D was diluted with Folin's reagent formed with an equal volume of 0.1 NaOH.Standard for this was made by dissolving 50 mg of BSA in 50 mL of water.Out of this stock solution, 10 mL was taken in another flask and diluted to 50 mL.200 µg protein was present in 1 mL of this solution.In test tubes, 0.2, 0.4, 0.6, 0.8, and 1 ml of working standard was taken in a series. 1 mL of each sample was taken in other test tubes, and the volume was made up to 1 ml with distilled water and 1 mL of distilled water in a test tube as a control.In all test tubes, including blank, 5 mL of reagent C was added, and then, using a vortex, all the contents were mixed and waited for 10 minutes.Immediately, 0.5 mL of D reagent was added, appropriately mixed, and incubated for 30 minutes at room temperature in the dark and at 660 nm absorbance against blank.

Qualitative phytochemical analysis
The preliminary phytochemical analysis was done by following Tariq and Rayiz method (11,12).Tannin test -2mL of aqueous extract was added into a test tube, and 3mL of distilled water was added, and this was shaken well, 2 drops of FeCl3 were added to the mixture.Very dark-coloured precipitation shows the tannin's presence.
Saponin test -This test is also called the "Froth Test".
In a test tube, 2 mL of aqueous extract was added to 6 mL of distilled water.The mixture was shaken well, and the froth formation indicates saponin's presence.
Flavonoid test -Flavonoids were determined by an acid-alkaline test in which 2mL of aqueous extract in a test tube was added with a few drops of concentrated ammonia.Yellow colour formation depicts the presence of flavonoids.Gallic tannin test/ Catecholic tannin test -1 mL of distilled water was added to 0.5 mL of aqueous extract, mixed well, and 2 drops of FeCl3 were added.Blue colour formation shows the presence of gallic tannin, while green and black indicate the presence of catecholic tannin.

Fourier transform infrared spectroscopy (FTIR) analysis
An FTIR spectrophotometer (Agilent, model no.Carry 630) was used to analyze all the samples, with a wavenumber region lying within the range from 800 to 3800 cm -1 with a 2 cm -1 resolution.10 mg of fine powder samples were put in the sample holder of the instrument, and transmittance was measured (13)(14)(15).

Total phenol content (TPC)
Folin-Ciocalteu's phenol reagent method was used for the quantitative estimation of the total phenols of the tea extract according to the technique employed by Kc et al., (16).For this, 100 µl of methanolic tea extract was taken in a test tube, and then 0.8 mL of 7.5% w/v sodium carbonate was added to it and incubated for 5 minutes.After 5 minutes, 1 mL of 10 times diluted Folin-Ciocalteu's reagent was added to it and left at room temperature in the dark for 40 minutes.The blue colour complex was formed, and the sample absorbance was taken at 765 nm.The results of TPC were expressed as GAE (Gallic acid equivalent).

Total flavonoid content (TFC)
The TFC was determined by the aluminium chloride method, as described by Dwiputri and Feroniasanti (17).500 µl of the methanolic tea extract sample was dissolved in 1.5mL methanol (80%).100 µl AlCl3(10% w/v) was added to this reaction mixture, followed by 100 µl 1M potassium acetate solution.After this, 2.8mL distilled water was added, the mixture was incubated for 30 minutes at room temperature, and optical density was taken at 415 nm.Quercetin was used to make the standard curve, and the equation obtained was used for the calculations.The result was expressed as milligram quercetin equivalent per gram of the sample.The same was repeated for all the samples.

Tannin content
Total tannin was determined using Folin-Ciocalteu's method Kc et al., (16), with few modifications.0.2 mL of methanolic extract from the sample was added to a test tube, and the volume was made up to 1 mL by adding 0.8 mL of distilled water.After that, 0.5mL of 1N Folin-Ciocalteu's reagent and 2 mL of 20% saturated NaCo3 were added.The mixture was kept at room temperature (RT) in the dark for 40 minutes.The absorbance was taken at 725 nm of wavelength using a UV-Vis Spectrophotometer.All samples were analyzed in triplicates.The results of all samples were correlated with the tannic acid standard curve.The total tannin of every sample was expressed in milligram (mg) of tannic acid equivalents per gram of sample.

Total soluble solids (TSS)
Total Soluble Solids were determined using a digital Refractometer at about 28-30°C.The instrument was calibrated using distilled water before measuring the TSS of the sample.A drop of the extract was placed on the prism of the refractometer to measure the TSS.The result was expressed as °Brix.

Antioxidant activity DPPH radical scavenging activity
DPPH radical scavenging activity was measured by following the method of Zhao et al., (18), with few changes.The DPPH solution (0.1M) was created by dissolving 0.39mg of DPPH in 80% methanol, and the volume was made up to 100 mL.This purple-coloured solution of DPPH was stored at -20°C for later usage.
For calibration, known concentrations of Trolox were employed.200 µL different concentrations of Trolox were added to 800 µL of methanolic DPPH.200 µL of samples were combined with 800 µL of methanolic DPPH, which was kept dark for 30 minutes.Absorbance was taken at 517nm using a UV-Vis Spectrophotometer, and each sample's results were expressed as TE per g of DW (Trolox equivalents per g of dry weight).

ABTS assay
The ABTS radical scavenging assay was determined by following the Aboagye et al., (19) and Ma et al., (20) method.The ABTS radical was generated by mixing 2.4mM of potassium persulfate and 7mM of ABTS stock solution in ratio 1: 1.This was kept for 12 hours to complete the reaction in the dark at RT until constant absorbance was observed.50 mL of methanol was added to 1 mL of prepared ABTS radical solution to dilute the ABTS solution.Then, 800µL of diluted ABTS solution was added to 200µL of sample, and this was mixed and kept in the dark at RT for 30 minutes.Using a UV-Vis spectrophotometer, absorbance was measured at 734 nm, and the results of each sample were correlated with the Trolox standard curve and expressed as TE per g of DW.

Ferric reducing antioxidant potential (FRAP) assay
Benzie and Strain's method was slightly modified to assess the ferric-reducing capacity of tea extracts (21).0.5 mL of methanolic tea extract was taken in a test tube, 7 mL of FRAP reagent (acetate buffer, TPTZ and ferric chloride) was added to it, and distilled water (blank sample) was also taken.Then, the sample was incubated at room temperature in the dark for 30 minutes.The absorbance was measured at 593 nm, and the results were expressed as Trolox equivalents.

Antimicrobial activity
The antimicrobial activity of each extract was evaluated using two bacterial strains, one of which was Gram-negative (Escherichia coli) and the other was Gram-positive (Staphylococcus aureus).The Agar well diffusion method was used to check the antimicrobial activity of the extracts of the fermented tea samples using MH (Mueller-Hinton) agar.All the materials were first autoclaved, and the media was autoclaved at 121°C for about 15 minutes.The media was then poured into sterile Petri plates and allowed to solidify at room temperature.After that, 50 µl of the inoculum suspension was placed over the agar using a micropipette and was spread uniformly using a sterile glass spreader.This was then allowed to dry for 10 minutes.Then, 3 wells were made on the alreadyflooded agar.The wells were filled with: 1. 50µL methanolic extract of sample 2. 50µL aqueous extract of sample 3. 50µL methanol 4. Antibiotic disc The plates were then incubated in an incubator for 12 hours, and the zones of inhibitions were observed.The same procedure was followed for all the five samples.

Proximate analysis
The highest moisture and ash content values were found in the West Bengal incubator fermented sundried sample with values of 1.80+ 0.001% and 5.93 + 0.06%, respectively.For crude fiber and protein, the highest values were observed in West Bengal spontaneous fermented sundried and West Bengal incubator fermented sundried, respectively, with values being 17+ 0.08% and 2.89+ 0.03 mg/g, respectively.Table 1 shows the results of the proximate nutrition analysis.The results of preliminary tests are shown in Table 2.
It indicates the presence of phytochemicals, i.e., tannin, saponin, flavonoid, and gallic tannin, and this is indicated by the change of colours of the reaction mixtures to black, frothy, yellow, and green-black, respectively.These bioactive compounds benefit plant defence mechanisms and influence other plant activities like antioxidant and antimicrobial activity.

FTIR analysis
The results obtained as a graph (Fig. 4) were interpreted using the infrared spectrum (24), as presented in Table 3.Total phenol, flavonoid, and tannin content TPC in the samples was found to be between the range 1.26 + 0.07 to 2.55 + 0.015mg GAE/g of dry sample (Table 4), the highest concentration being found in the West Bengal incubator fermented sun dried sample.Among WB fermented samples, the sun-dried sample had more phenolic content than oven-dried ones.TFC in the samples was found to be between the range 0.9 + 0.01 to 1.51 + 0.01 mg QE/g of dry sample (Table 4), the highest concentration being found in the West Bengal incubator fermented sun dried sample.The total tannin content in the samples was found to be between the range 0.29 + 0.06 to1.01 + 0.01 mg tannic acid equivalents/g of dry sample; the lowest concentration is found in West Bengal spontaneous fermented oven-dried sample (Table 4).Values are mean ± standard deviation of duplicate determination.

Total soluble solids
The total soluble solids were performed on WB samples only.It was observed that the amount of TSS increased in fermented samples.The WB fresh showed 2.10 ± 0.14 °Brix while fermented samples had a range from 4.15 ± 0.21 to 5.55 ± 0.49 °Brix (Table 5).Values are means ± standard deviation of duplicate determination

Total antioxidant activity
The method was performed on WB samples only.The antioxidant activity determined by the DPPH method showed that fermented samples had more antioxidant capacity to scavenge DPPH free radicals than fresh samples.The antioxidant activity of WB-fermented samples ranges from 0.895 ± 0.015 mg TE/g to 1.37 ± 0.01 mg TE/g.The ABTS assay also showed similar trends as that of DPPH.The fermented sample showed more antioxidant activity than the fresh one.The ABTS scavenging activity of fermented samples ranges from 38.825 ± 0.365 mg TE/g to 45.315 ± 1.125 mg TE/g.The FRAP is minimum by fresh leaves, i.e., 0.98 mg / g DW; maximum antioxidant activity is shown by Spontaneous fermented sun-dried tea leaves, i.e., 3.21 mg / g DW.Table 6 shows the results of all three antioxidant assays.Values are means ± standard deviation of duplicate determination

Antimicrobial activity
The highest antibacterial activity against E. coli was observed in WB IF OD aqueous extract with a zone of inhibition of 1.65 + 0.21 mm (Table 7).The methanolic extracts showed almost no zone of inhibitions, as the methanol present might interfere with the activity (Table 8).6, represented the plates of antibacterial activity of samples against E. coli and S. aureus, respectively.Also, no antibacterial activity was observed against the Gram-positive strain S. aureus.The fermented tea samples exhibit antimicrobial activity due to the presence of polyphenols.

DISCUSSION
An increase in moisture content is observed after fermentation of the tea leaves.When the sun dried and the oven-dried samples are compared, the sun-dried sample has more moisture than the oven-dried one in both incubator and spontaneous fermentation.A similar trend is observed in ash and crude fiber content.This nutritional biotransformation is observed due to the processing and fermentation of the leaves.The increase in nutritive values observed when fresh and fermented leaves are compared is due to the activity of microorganisms resulting in the conversion of various chemical compounds.Oven-dried samples have less moisture, ash, and crude fiber content than the sun-dried samples as they were subjected to a very high heat of 100°C in a hot air oven after fermentation, and heat might have caused these changes.Also, the microorganisms could have digested the fibers into sugars, which might have caused a decrease in fiber content.It was observed that the protein content of the tea leaves decreased during fermentation and processing.This would be because the microorganisms would have used amino acids during the fermentation process.Another reason for this decrease could be the heat treatment given to the tea leaves at 100°C, which would have degraded the proteins in the leaves (22).
The FTIR analysis of the samples showed the presence of functional groups in tea leaves.Various peaks were observed in each sample.The peaks ranging from 3280 cm -1 to 3300 cm -1 corresponded to O-H/N-H functional groups, showing the presence of hydroxyl, carboxylic, and primary aliphatic amine (23).It can demonstrate the existence of particular polyphenols, proteins, and polysaccharides.WB fresh samples showed the presence of an alkyne group at 2115 cm-1, while fermented samples showed a peak ranging from 2105 cm -1 to 2123 cm -1 .Also, fermented samples showed the presence of another functional group common in fresh/unfermented samples.The presence of bonding such as C-H, C=O, C≡C, O-H, and N-H indicates the presence of functional groups, which shows that there are several compounds present in tea leaves, which can be the polyphenols and flavonoids as their structures also contain the observed functional groups.Alcohol from WB fresh is converted to carboxylic acid and amine in fermented samples, showing that the components have undergone some alterations during fermentation.These functional groups were observed in fermented samples but not in fresh/unfermented ones.The FTIR data can be correlated with antioxidant potential, TPC, and TFC.Such groups indicate the presence of phenolic and flavonoid compounds, which can be related to antioxidant activity (24).
The high number of polyphenols indicates a high quality of tea (25).This is possible as the plant cell walls might have weakened at high temperatures during drying, and the phenolic contents would have leached out.Also, it is known that phenolic compounds are present in bound form in the food matrix; therefore, due to high temperatures, the interactions between the food matrix and phenolic components were broken, resulting in their leaching and high phenolic content.In previous works, phenolic content decreased due to fermentation, but in the current study, the phenols increased due to the activity of enzymes PPO and PO (2).Also, compared with previous studies, it can be concluded that ovendried samples showed lower values than sundried due to the instability of PPO and PO at higher temperatures.Flavonoid content was higher in fermented tea samples than in fresh tea leaves.The increase in flavonoids due to fermentation increases the acidic value, which results in the liberation of bound flavonoid components and makes flavonoids more available (22).Another reason could be the change in the metabolic activity of the microorganisms.Flavonoid content increases in tea after fermentation due to enzymatic reactions that transform flavan-3-ols into complex polymers (26).Tannin content was high in fresh leaves as compared to fermented tea leaves.Tannic acid content may have decreased due to the actions of the enzyme polyphenol oxidase or microflora.The decline can be attributable to the Lactobacillus enzyme tannase, which breaks down tannin-protein complexes during fermentation (27).The tannin content of fermented tea leaves decreases because of the fermentation temperature, which was 30°C (28).The oxidation of tea leaves due to the presence of enzymes PPO and PO converts polyphenols into their oxidized form in the presence of oxygen.
According to the literature, the total soluble solids should have decreased after the fermentation process due to conversions by the microbes.Still, according to our findings, TSS increased, which can be studied on a molecular basis, and till now, no reason is to be found.
Flavonoids, a class of phenolic compounds, play a significant role in antioxidant activity.The biological protection flavonoid provides because of its capacity to act as an electron donor to prevent free radicals and active oxygen species from being oxidized (17).The antioxidant activity of both methods showed a positive correlation with polyphenols and flavonoids.They significantly impact the free radical scavenging activity of the samples.The antioxidant activity of tea leaves is highly increased due to fermentation.In comparison to fresh leaves, fermented tea leaves have high antioxidant activity.The increase in antioxidant activity is due to the oxidation of phenolic components of tea leaves.However, antioxidant activity can vary by different processing conditions.These components help scavenge free radicals from the body (29).
Tea polyphenols undergo oxidation during fermentation, which produces complex polymeric compounds like theaflavins and thearubigins.The tea's catechins are significant contributors to the inhibition of bacterial growth.In the following experiment, the activity differs according to the type of extract.It has been discovered that these substances have strong antimicrobial effects on various bacteria, viruses, and fungi.They can prevent their growth and proliferation by interfering with microorganisms' cellular architecture and metabolic functions.Tea may not exhibit antimicrobial activity against Staphylococcus aureus due to the presence of proteins that can bind and inactivate tea catechins.The difference in results is due to the different conditions provided for the preparation of samples.It can also be concluded that tea extracts do not show microbial inhibition against Staphylococcus aureus due to the thick peptidoglycan cell wall; therefore, the compounds cannot penetrate the cells and inhibit their growth.The antimicrobial properties of fermented tea leaves emphasize the importance of the fermentation process in producing tea with enhanced microbial control and potential health advantages.

CONCLUSION
The present investigation examined the proximate nutritional composition, bioactive compounds, and antioxidant activity of Camellia sinensis before and after different fermentation conditions.Results suggested a significant difference in nutritional and phytochemical components among all tea samples with different conditions.It was concluded that fermentation improved the composition of the physiochemical properties and bioactive compounds and affected the tea leaves' antimicrobial activity.The FTIR analysis of the samples showed the presence of

Table 1 :
Nutritional Composition of different tea samples Values are means ± standard deviation of triplicate determination.

Table 3 :
FTIR spectral peaks obtained and their interpretation of respective groups

Table 4 :
Total phenolic, Total Flavonoid, and Total Tannin Content

Table 5 :
Total soluble solids

Table 6 :
Antioxidant activity of tea samples by different methods

Table 7 :
Zone of inhibitions against E. coli

Table 8 :
Zone of inhibitions against S. aureus