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 Gangwar.pdf

Aishwarya

July 19, 2023
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  1. International Journal of Food Studies IJFS April 2018 Volume 7 pages 100–110
    Fermentation of Tender Coconut Water by Probiotic Bacteria
    Bacillus coagulans
    Aishwarya Singh Gangwara, Aastha Bhardwaja, and Vasudha Sharmaa*
    a Department of Food Technology, Jamia Hamdard (Deemed to be University), New Delhi – 110062, India
    *Corresponding author
    [email protected]
    Tel: +91-9810288903
    Received: 20 July 2017; Published online: 18 April 2018
    Abstract
    Coconut water is currently being considered as an elixir for patients suffering from diseases like
    dengue and malaria as well as chikungunia to provide hydration properties to the body. It has become
    a popular beverage for many people owing to its palatability and high mineral content. In this study,
    the growth, survival and fermentation performance of the probiotic bacterium Bacillus coagulans in
    coconut water was assessed in order to produce a novel non-dairy, probiotic beverage. The species
    was characterized on the basis of morphology, physiology and biochemical parameters and its probiotic
    attributes were assessed. Batch fermentations were carried out for 2 days at a constant 37°C, thereafter
    the samples were subjected to microbiological and chemical analysis. The results suggested that the
    specie produced lactic acid and was acid and bile tolerant. The pH and titratable acidity of probiotic
    fermented coconut water were found to be 4.4 and 0.53 % lactic acid, respectively. The viscosity
    of fermented coconut water increased significantly from an initial 5.13 mPa.s to 5.35 mPa.s because
    of the increase in soluble solids content due to exopolysaccharide production by B. coagulans during
    fermentation. Also, the overall acceptability score of probiotic coconut water was higher than tender
    coconut water, suggesting its feasibility for use as a probiotic beverage.
    Keywords: Probiotic non dairy beverage; Fermented coconut water; Sensory evaluation; Physico-
    chemical characteristics
    1 Introduction
    According to the World Health Organization and
    the Food and Agriculture Organization of the
    United Nations (FAO / WHO, 2001), probi-
    otics are defined as “live microorganisms which,
    when administered in adequate amounts, con-
    fer a health benefit on the host”. Numerous
    studies have highlighted the health benefits as-
    sociated with consumption of probiotic bacteria.
    In the past decade, there has been an increase
    in consumer demand for functional foods such
    as yoghurt and other fermented dairy products
    supplemented with probiotic organisms (Penna,
    Rao-Gurram, & Barbosa-Canovas, 2007). How-
    ever, dairy substrates may contain potential al-
    lergens, such as casein and they require cold stor-
    age to enhance their shelf life. Also, the choles-
    terol content of dairy products is high. Owing
    to such facts and the increasing trend of vege-
    tarianism , the demand for novel products with
    non-dairy matrices has expanded (Ranadheera,
    Baines, & Adams, 2010). Also producing probi-
    otic products with foods and beverages which are
    part of day-to-day life is encouraged. This has
    led to an increased demand for non-dairy probi-
    otic foods, such as coconut aqueous extract, fruit
    drinks, nutrition bars, soy products and cereal-
    Copyright
    ©2018 ISEKI-Food Association (IFA) 10.7455/ijfs/7.1.2018.a9

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  2. Probiotic tender coconut water 101
    based products. The nutritive values and wide
    distribution of these raw materials are important
    when they are used as functional food items (An-
    gelov, Gotcheva, Kuncheva, & Hristozova, 2006).
    Tender coconut water (TCW), the liquid en-
    dosperm obtained from immature green co-
    conuts, in its natural form is a refreshing and
    nutritious beverage, widely consumed around
    the world due to its beneficial health proper-
    ties (Pummer, Heil, Maleck, & Petroianu, 2001).
    Moreover, coconut water plays an important al-
    ternative role for oral rehydration and even for
    intravenous hydration of patients in remote re-
    gions (Campbell-Falck, Thomas, Falck, Tutuo,
    & Clem, 2000) in addition to providing protec-
    tion against induction of myocardial infarction
    (Anurag & Rajamohan, 2003). It was identi-
    fied in the late 1930s as a nutrient helping to
    reduce anemia in pregnancy (Jackson, Gordon,
    Wizzard, McCook, & Rolle, 2004) and which also
    helped to prevent mitochondrial toxicity induced
    by methanol metabolites. The major chemi-
    cal constituents of coconut water are sugars and
    minerals and minor ones are fat and nitrogenous
    substances.
    Interestingly, the perception and utilization of
    coconut water has evolved over the years ow-
    ing to its unique chemical composition of sug-
    ars, vitamins, minerals, amino acids, enzymes
    and phytohormones that play different functional
    roles in the human system (Yong, Ge, Ng, &
    Tan, 2009). One example is the consumption
    of coconut water as a refreshing and hydrat-
    ing beverage due to its rich mineral content
    of sodium, potassium, magnesium and calcium,
    which can replenish the electrolytes of the hu-
    man body excreted through perspiration (Saat,
    Singh, Sirisinghe, & Nawawi, 2002). Studies
    have shown that coconut water has hydrating
    and exercise performance effects that are compa-
    rable to those of carbohydrate electrolyte sports
    drinks (Kalman, Feldman, Krieger, & Bloomer,
    2012). Chauhan, Archana, Singh, Raju, and
    Bawa (2014) blended coconut water with lemon
    juice to develop a refreshing beverage by optimiz-
    ing the pH, colour and sensory attributes (ap-
    pearance, aroma, taste, consistency and overall
    acceptability).
    Current knowledge on the fermentation of co-
    conut water is rather limited (Kuswardani,
    Kusumawati, Srianta, & Sabrina, 2011), espe-
    cially fermentation with probiotic bacteria. How-
    ever, Dharmasena (2012) recently developed a
    novel non-dairy probiotic beverage with a mix-
    ture of oat meal and coconut water using probi-
    otic Lactobacillus plantarum Lp 115- 400B. Al-
    though lactic acid bacteria (LAB) are the most
    commonly used probiotics, some spore-forming
    bacteria have also been exploited as probiotics
    due to their unique properties. Lee, Boo, and Liu
    (2013) studied the fermentation performance,
    growth patterns and survival of Lactobacillus aci-
    dophilus and Lactobacillus casei in coconut wa-
    ter. Prado et al. (2015) developed a non dairy
    fermented functional beverage using coconut wa-
    ter for its hydrating properties, functional health
    properties and nutritional benefits.
    The genus Bacillus is the most extensively stud-
    ied group of spore-forming probiotics. Other
    spore-formers being used as probiotic bacteria
    are Paeni Bacillus polymyxa and Brevi Bacil-
    lus laterosporus that were initially classified as
    Bacillus species (Cutting, 2011). There are sev-
    eral advantages of using spores over other non-
    spore forming bacteria. Spores are heat resistant
    and can survive harsh conditions during produc-
    tion and storage processes. They are also able to
    withstand the extreme physiological conditions
    such as the low pH of the gastrointestinal tract,
    bile salts and enzymes (Cutting, 2011). Bacil-
    lus coagulans, a widely used probiotic, has been
    shown to induce antibody production in humans.
    This probiotic bacterium is the most commer-
    cially available and investigated probiotic bac-
    terium, with proven beneficial impacts on health
    in animal and human trials (Hawrelak, 2003).
    In order to be able to exert its beneficial effects,
    a successful potential probiotic strain is expected
    to have a number of desirable properties. Bacte-
    rial characterization with good probiotic proper-
    ties is of great importance in probiotic functional
    foods. In addition to production of lactic acid,
    the acid and bile tolerance are two fundamental
    properties that indicate the ability of probiotic
    microorganism to survive the passage through
    the upper gastrointestinal tract, particularly the
    acidic conditions in the stomach and the pres-
    ence of bile in the small intestine (Hyronimus,
    Le Marrec, Sassi, & Deschamps, 2000).
    The objective of the present investigation was to
    IJFS April 2018 Volume 7 pages 100–110

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  3. 102 Gangwar et al.
    assess the growth, survival and fermentation per-
    formance of probiotic bacterium B. coagulans in
    coconut water to produce a novel non-dairy pro-
    biotic beverage, which could provide both hydra-
    tion as well as probiotic benefits to all individu-
    als, especially athletes and recreationally active
    fitness enthusiasts.
    2 Materials and Methods
    2.1 Procurement and preparation
    of raw material
    Tender coconuts of the Cocus nucifera type, age
    5-7 weeks, were chosen for this study. These were
    purchased from a local market in Delhi. Tender
    coconut water was collected in a sterile beaker
    (500 ml capacity) under aseptic precautions as
    per method given by Acharya, Gupta, Golwala,
    Store, and Sheth (1965). The flask was plugged
    with cotton and autoclaved at 121
    °C at 15 psi
    for 15 minutes. The flask with sterile coconut
    water was cooled and stored at 4
    °C prior to the
    fermentation stage.
    2.2 Chemicals
    Sodium hydroxide, sodium chloride, hydrochlo-
    ric acid, bile salts, L-cysteine, dextrose, peptone,
    yeast extract, beef extract, MRS agar, MRS
    broth, GYE broth and agar were obtained from
    Sigma-Aldrich (New Delhi, India). , Gallic acid,
    phenol, 3, 5-dinitro salicylic acid (DNS) reagent,
    sulphuric acid, methanol, ethanol, hexane, ether,
    crystal violet, Rochell’s salt, bovine serum albu-
    min (BSA) and sodium sulphite were procured
    from HiMedia (Mumbai, India). All chemicals
    employed were of reagent grade.
    2.3 Procurement of Probiotic
    culture and Preparation of
    Bacterial Suspension Culture
    B. coagulans MTCC 5856 strain used in the
    study was procured from Microbial Type Cul-
    ture Centre and Gene Bank (MTCC) at the
    Institute of Microbial Technology (IMTECH),
    Chandigarh, India. The spores of B. coagulans
    were propagated separately in sterile MRS broth
    in a sterile Erlenmeyer flask for up to 48 h at 37
    °C
    aerobically and then stored at 4
    °C until use.
    2.4 Analysis of Probiotic
    attributes
    The probiotic attributes of the species such as
    the ability to produce lactic acid, high acid tol-
    erance and their ability to deconjugate bile salts
    were investigated (Aly, Abd-El-Rahman, John,
    & Mohamed, 2008).
    Analysis of Probiotic attributes
    To determine the tolerance of the specie to low
    pH, the method of Pennacchia et al. (2004) was
    used with slight modifications. For this purpose,
    active cultures were used (incubated for 16-18 h).
    A 0.5 ml aliquot of the bacterial culture was in-
    oculated in 10 ml of phosphate buffered saline
    adjusted to pH 2.5 with 4 N HCl. Cultures were
    incubated at 37
    °C. During 0, 1, 2 and 3 h of
    incubation, viable microorganisms were enumer-
    ated using the pour plate technique on MRS agar
    plate at 37
    °C.
    Bile salt tolerance
    The tolerance capacity of B. coagulans for high
    bile concentration was checked using the method
    suggested by Chung, Kim, Chun, and Ji (1999).
    A 1 % concentration of bile salts in sterile dis-
    tilled water was inoculated with 1 % active bac-
    terial suspension and incubated at 37
    °C. After
    incubation for 4 h, viable colonies were enumer-
    ated each hour using pour plate technique.
    Production of lactic acid
    The qualitative test for lactic acid production by
    B. coagulans was carried out using the method
    as described by Demirci, Pometto, and Johnson
    (1993). Glucose yeast agar plates were prepared
    and dilutions were made from the main culture
    suspension. 1 ml of the bacterial suspension was
    pour plated from the final dilution tube. Af-
    ter solidification, the plates were incubated at
    37
    °C for 48 hours. The colonies thus obtained
    IJFS April 2018 Volume 7 pages 100–110

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  4. Probiotic tender coconut water 103
    were transferred aseptically to 15 ml of previ-
    ously sterilized and cooled glucose yeast extract
    liquid broth. This was incubated at 37
    °C for 48
    hours and was then centrifuged at 2500-3000 rpm
    for 10 minutes. The clear supernatant was trans-
    ferred to a separating funnel and extracted by
    using 5 ml of dilute sulphuric acid (10%) and
    50 ml of ether. The ether layer was then col-
    lected, evaporated in water bath and the residue
    thus obtained was dissolved in 5 ml of water. To
    this, Uffelman’s reagent (prepared by adding two
    drops of 1N ferric chloride to 10 ml of 1% phe-
    nol solution) was added dropwise and the colour
    change was observed.
    2.5 Preparation of the inoculum
    For preparation of the inoculum, 25 ml of sterile
    tender coconut water was inoculated aseptically
    with 1% v/v ml of bacterial suspension culture
    and incubated at 37
    °C for 12–14 h. This was
    then serially diluted to obtain a working culture
    containing 108 CFU/ml. 1 ml from the respec-
    tive tube was pour plated onto the MRS agar
    media plate and the plate was incubated at 37
    °C
    for 48 h. The number of colonies between 30 -
    300 were considered ideal during counting. The
    viable spore count was obtained by the following
    formula:
    The viable spore count = Number of colonies
    per plate × Final dilution factor
    2.6 Fermentation of the Tender
    Coconut Water
    Fermentations with B. coagulans were carried
    out in 150 ml of sterile coconut water in sterile
    250 ml Erlenmeyer flasks. These flasks contain-
    ing sterile coconut water were inoculated with 1
    % (v/v) pre-culture of the probiotic strain from
    the respective broth. The batch fermentations
    were carried out for 2 days at a constant 37
    °C
    in triplicate. After two days, the samples were
    taken aseptically after swirling the conical flasks
    gently for homogenization and these were sub-
    jected to microbiological and chemical analysis.
    2.7 Analytical determination
    Samples (20 ml) were taken after 2 days of fer-
    mentation, and the viability of the probiotic cul-
    ture, pH, total soluble solids (
    °Brix), acidity
    and cell biomass of the probiotic coconut water
    were determined. Similar tests were also carried
    out for the tender coconut water sample. The
    pH of tender coconut water and fermented co-
    conut water was measured using a digital pH
    meter (TOSHCON, India) at 25
    °C. The total
    soluble solids were determined using an Abbe
    refractometer (AC0012, MRC Scientific Instru-
    ments, India) and the total soluble solid content
    was expressed as
    °Brix and Refractive Index at
    25
    °C. The rheological measurements were carried
    out at 25
    °C using a controlled stress viscometer
    (Brookfield VIS-S2, MRC Scientific Instruments,
    India) equipped with a coaxial cylinder (cylinder
    no. 4); the radii ratio of coaxial cylinder was
    1.08477. The acidity was determined by titra-
    tion with standard 0.01M NaOH solution, using
    phenolphthalein as indicator and acidity was ex-
    pressed as % citric acid (Ranganna, 1986). The
    biomass/cell density was determined spectropho-
    tometrically at 540 nm using the MacFarland
    scale (Kandler & Weiss, 1986), both pre and post
    the prebiotic fermentation.
    Estimation of total sugars and
    reducing sugars
    Total sugars of tender coconut water and fer-
    mented coconut water were determined col-
    orimetrically using the phenol-sulphuric acid
    method and expressed as percentage sugar
    (Miller, 1959). The absorbance was measured
    at 490 nm and expressed as glucose concentra-
    tion (mg/ml). Similarly, the reducing sugars
    of tender coconut water and fermented coconut
    water were determined colorimetrically using 3,
    5-dinitro salicylic acid (DNS) reagent and ex-
    pressed as % (Miller, 1959). The absorbance was
    measured at 540 nm and expressed as glucose
    concentration (mg/ml).
    2.8 Viable cell determination
    Appropriate dilutions from coconut water sam-
    ples were made using sterile peptone water (1
    IJFS April 2018 Volume 7 pages 100–110

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  5. 104 Gangwar et al.
    gl−1) and pour plated onto MRS agar. Plates
    were then incubated at 37
    °C for 48 h. The ex-
    periment was performed in triplicate and the av-
    erage number of colony-forming units per millil-
    itre (CFU/ml) were determined using a Darkfield
    Quebec Colony Counter.
    2.9 Sensory evaluation of
    Fermented Coconut water
    Sensory quality of the fermented coconut water
    was measured after 7 days of fermentation us-
    ing a 9 point hedonic scale, with respect to the
    appearance/colour, smell/odour, aroma/flavour,
    taste and texture/mouthfeel, and for assessment
    of overall acceptability of the product. Sensory
    evaluation was carried out by a semi trained
    panel consisting of 30 food scientists and technol-
    ogists (between 20-45 years of age) chosen from
    faculty members and post graduate food tech-
    nology students of the department. The samples
    were presented at 20
    °C in the sensory evalua-
    tion laboratory. The samples were coded and
    presented individually to each panellist to avoid
    bias. Potable water to rinse between the two
    samples was also supplied. The panellists were
    asked to record their observations on the sensory
    sheet using the scales described above.
    The research was approved by the institutional
    human experimentation committee or equivalent,
    and informed consent was obtained from the par-
    ticipants.
    2.10 Statistical analysis
    Results were expressed as mean values
    ± stan-
    dard deviation of at least three replications.
    Results were statistically evaluated by ANOVA
    (Minitab 14) at a confidence level of 0.95.
    3 Results and Discussion
    3.1 Analysis of probiotic
    attributes
    Resistance to low pH
    Strains need to be resistant to the stressful con-
    ditions of the stomach (pH 1.5-3.0) so resistance
    to pH 3 is often used in in vitro assays to deter-
    mine the resistance of probiotic species to stom-
    ach pH. Food usually stays in the stomach for
    about 3 h so this time limit was taken into ac-
    count in the research (Prasad, Gill, Smart, &
    Gopal, 1998) Since a significant decrease in the
    viability of strains is often observed at pH 2.0
    and below, phosphate buffered saline (PBS) was
    used with the pH adjusted to 3.0 to select strains
    resistant to low pH. Effects of low pH (at 2.5) and
    survivability of B. coagulans at 0, 1, 2 and 3 h
    intervals are shown in Table 1. No effect of low
    pH (at 2.5) on B. coagulans was observed, sug-
    gesting that the colonies were able to survive the
    low pH conditions and were tolerant to high acid.
    This agreed with results reported by Argyri et al.
    (2013) where nine strains of Lactobacillus showed
    very high resistance to low pH (L. plantarum, L.
    pentosus, L. casei subsp paracasei). Acid tol-
    erance can be mediated by membrane ATPases
    as described for L. acidophilus by Lorca and de
    Valdez (2001).
    Bile salts tolerance
    As the mean intestinal bile concentration is be-
    lieved to be 0.3% (w/v) and the residence time
    of food in small intestine is estimated to be 4 h
    (Prasad et al., 1998), this parameter was con-
    sidered. Bile salts tolerance of B. coagulans at
    various time intervals are shown in Table 2. The
    results showed that the specie retained viability
    with no reduction in the cell count at 1% bile
    salt concentration. B. coagulans showed a good
    tolerance towards bile salts. Similar results have
    been reported by Jensen, Grimmer, Naterstad,
    and Axelsson (2012) where Lactobacillus species
    were found to tolerate gastric juices with negli-
    gible reduction in the viability.
    In high bile salts concentration, most bacteria
    show an inability to survive, but spore formers
    show a better tolerance. Bacillus sp. as pro-
    biotics, survive the transit very well since they
    are in the form of spores (Duc, Hong, Barbosa,
    Henriques, & Cutting, 2004). Bile secreted in
    the small intestine reduces the survival of bac-
    teria by destroying their cell membranes, whose
    major components are lipids and fatty acids and
    these modifications may affect not only the cell
    permeability and viability, but also the interac-
    IJFS April 2018 Volume 7 pages 100–110

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  6. Probiotic tender coconut water 105
    Table 1: Effect of low pH and bile salts on survivability of B. coagulans
    pH Time duration (h) No. of viable colonies (log CFU/ml)
    2.5 0 9.74
    ±0.44c
    2.5 1 9.71
    ±0.31c
    2.5 2 9.55
    ±0.05b
    2.5 3 9.08
    ±0.57a
    Bile salt concentration (%) Time duration (h) No. of viable colonies (log CFU/ml)
    1 0 9.83
    ±0.43d
    1 1 9.81
    ±0.35c
    1 2 9.73
    ±0.21b
    1 3 9.69
    ±0.18a
    1 4 9.65
    ±0.36a
    Means and standard deviation for n=3; Values within columns with different superscripts were significantly different (p<0.05) according to
    Duncan’s multiple test range
    tions between the membranes and the environ-
    ment (Gilliland, Staley, & Bush, 1984).
    Production of lactic acid
    B. coagulans showed positive results for lactic
    acid production capability. The solution turned
    bluish, violet to yellow which suggested the pres-
    ence of lactic acid in the medium. This meant
    that the medium contained sugars that could
    be fermented by the bacterium to produce lac-
    tic acid.
    3.2 Analytical determination
    pH and total soluble solids
    The pH of tender coconut water and probiotic
    fermented coconut water was found to be 5 and
    4.4 respectively. Fermentation causes a rapid de-
    crease in pH from 5.02 to 4.44. B. coagulans
    could tolerate acid medium and survive during
    fermentation process. The total soluble solids in
    tender coconut water and probiotic fermented co-
    conut water were found to be 5.0 and 6.0
    °Brix
    respectively, suggesting that the increase in vi-
    able cell count corresponded to the decrease in
    pH and sugars consumed during fermentation.
    Total soluble solids content was 5.0
    °Brix which
    indicated that solids present in tender coconut
    water was mainly soluble solids such as sugars.
    An increase in total soluble solids content of the
    fermented probiotic coconut water was due to the
    increase in viable cell counts after fermentation.
    The refractive index was found to be 1.340 and
    1.342 in tender coconut water and fermented co-
    conut water, respectively. It showed the purity
    of the coconut water.
    Titratable acidity
    Titratable acidity in coconut water samples was
    found to be 0.18 % (citric acid) and 0.53 % (lactic
    acid) respectively. Tender coconut water showed
    a titratable acidity value of 0.18 % (citric acid)
    due to the presence of ascorbic acid. After fer-
    mentation, the titratable acidity value was 0.53
    % lactic acid. Lactic acid is the major end prod-
    uct of the conversion of carbohydrates due to
    utilization of sugars present in coconut water.
    B. coagulans is a typical strain reported for lac-
    tic acid production; the thermophilic character
    of this strain (growth at 52
    °C) indicates that
    it is particularly adapted for industrial produc-
    tion of lactate without sterile conditions (Payot,
    Chemaly, & Fick, 1999).
    Total sugars and reducing sugars
    Total sugars and reducing sugar in tender co-
    conut water was 3.96 % and 2.37 % after fermen-
    tation, which decreased to 3.15 % and 2.27 %
    respectively due to utilization of sugars present
    IJFS April 2018 Volume 7 pages 100–110

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  7. 106 Gangwar et al.
    Table 2: Physico-chemical characteristics of tender coconut water (TWC) and fermented coconut water
    (FWC)
    S.No. Parameter TCW FCW
    1. pH 5.02
    ±0.03a 4.44
    ±0.12b
    2. Total soluble solids (
    °Brix) - Refractive Index 5.0 – 1.340 (RI)a 6.0 – 1.342 (RI)b
    3. Viscosity (mPa.s) at 25
    °C 5.13
    ±0.04a 5.35
    ±0.02b
    4. % Titrable acidity 0.18
    ±0.01a (% Citric acid) 0.53
    ±0.02b (% Lactic acid)
    5. Biomass/Cell density at 540 nm 0.121
    ±0.02a 0.583
    ±0.01b
    6. Total Sugars (%) 3.96
    ±0.10a 2.37
    ±0.07b
    7. Reducing sugar (%) 3.15
    ±0.05a 2.27
    ±0.02b
    Means and standard deviation for n=3; Values within rows different superscripts were significantly different (p<0.05) according to a paired
    t-test
    in tender coconut by the species during fermen-
    tation.
    Flow behaviour
    Rheological parameters are good indicators of
    texture and important for consumer acceptance.
    The viscosity values of tender coconut water
    and fermented coconut water was 5.13 and 5.35
    mPa.s at 25
    °C depending upon the concentra-
    tion. Total soluble solids content had a signif-
    icant effect on viscosity of tender coconut wa-
    ter. The magnitude of viscosity of fermented
    coconut water increased significantly 5.35 mPa.s
    with the increase in soluble solid content due to
    exopolysaccharide production by B. coagulans.
    Several strains of B. coagulans have been studied
    for their exopolysaccharide production. The pro-
    biotic bacterium produces an exopolysaccharide
    (EPS) during exponential and stationary growth
    phases (Kodali & Sen, 2008).
    Microbial exopolysaccharides are getting atten-
    tion as natural thickeners. Most of the econom-
    ically important bacterial EPS are produced by
    LAB, which are manipulated as probiotics to im-
    prove rheology and texture of fermented prod-
    ucts.
    The viscosity of tender coconut water is strongly
    depended on inter-molecular forces between
    molecules and water-solute (sugars and acids)
    interactions, which result from the strength of
    hydrogen bonds and inter-molecular spacing as
    both were strongly dependent on concentration
    and temperature. An increase in soluble solid
    content leads to increase in hydrated molecules
    and hydrogen bonding with hydroxyl groups of
    solute, which would enhance the flow resistance
    that leads to increase in viscosity of liquid. In
    case of tender coconut water, soluble solids was
    mainly due to the sugars content and in case
    of fermented coconut water, viable cells and ex-
    opolysaccharide played an important role in the
    viscosity values.
    Biomass / Cell density
    The biomass / cell density was determined spec-
    trophotometrically at 540 nm. The optical den-
    sities of tender coconut water and fermented co-
    conut water were 0.121 and 0.683 respectively.
    The cell density of fermented coconut water de-
    termined at 540 nm was 0.683, which was higher
    than 0.600 that corresponded to 109 CFU/mL,
    using the Mac Farland scale. This is ideal for
    probiotic beverage functionality.
    Viable cell counts
    In order to obtain the potential health benefits,
    the population of probiotics in a product, the
    viability of probiotic microorganisms and their
    ability to activate at the desired site in the al-
    imentary canal are very important. The initial
    inoculum size of probiotics in the selected food
    item is critical. The effective daily dose of probi-
    otics is considered to be 109-1011 CFU (Sanders,
    1999). Hence, consumption of 100 ml or a g of a
    product bearing the therapeutic minimum (106-
    108 CFU/ml or g of the product), would satisfy
    the daily requirement. The viable cell counts for
    IJFS April 2018 Volume 7 pages 100–110

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  8. Probiotic tender coconut water 107
    Table 3: Evaluation of sensory properties of coconut water after 7 days of fermentation. Attribute scales:
    1 - 9
    Attributes Fermented Coconut Water Tender Coconut Water
    (Control)
    Appearance / Colour 7.5
    ±.04a 9
    ±.30b
    Smell / Odour 7.5
    ±.12a 7.5
    ±.10a
    Taste 7.5
    ±.04a 6
    ±.14b
    Mouthfeel 7.5
    ±.08a 6
    ±.21b
    Overall acceptability 7
    ±.12a 6
    ±.20b
    The experimental values within rows with different superscripts were significantly different (p<0.05) according to a paired t-test
    fermented coconut water were found to 9.73 log
    CFU/ml (Table 1), showing that it could be used
    successfully as a vehicle for probiotics.
    The physico-chemical characteristics of tender
    coconut water and fermented coconut water are
    summarized in Table 2.
    3.3 Sensory evaluation of
    Fermented Coconut water
    Sensory properties were chosen as the main crite-
    rion of the quality of fermented products, being
    the most important attribute for consumers.
    According to the consensus of the panellists dur-
    ing sensory evaluation, the overall acceptability
    on a 9 point hedonic scale of fermented coconut
    water was found to be higher than tender coconut
    water. It was determined that the main descrip-
    tors that characterized the product were acidity
    and sweetness, with acidity being the attribute
    responsible for the sensory difference perceived
    by the panellists.
    The parameter of fluid food quality related to
    rheological viscosity is known as mouthfeel and
    is defined as the mingled experience derived from
    the sensation on the skin of the mouth after in-
    gestion of a food or beverage. Nevertheless, the
    fermented coconut water still had high concen-
    trations of residual sugars, which would enable
    retention of sweetness. The evaluation parame-
    ters and their respective scores are shown in Ta-
    ble 3.
    4 Conclusions
    In this study, tender coconut water was used as
    the sole fermentation medium, without any addi-
    tives, to ensure that it was the only raw material
    that regulated the growth and metabolism of the
    probiotic bacteria. The good adaptation of B.
    coagulans in the tender coconut water showed
    that if a potential probiotic strain is used as a
    starter culture then it might produce a fermented
    product with defined and consistent character-
    istics and possibly health-promoting properties.
    Fermented coconut water gives the advantages
    of plant-based products, and the presence of live
    bacteria with probiotic qualities enhances the
    benefits. In conclusion, the present study demon-
    strated good growth of probiotic B. coagulans in
    tender coconut water. These results suggest the
    feasibility of fermenting coconut water into a pro-
    biotic beverage, especially for its nutrition, with
    the health benefits of probiotics.
    Acknowledgements
    The authors gratefully acknowledge Department
    of Food Technology, Jamia Hamdard, for support
    and development towards this project.
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