Food Chemistry Advances 4 (2024) 100593

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Food Chemistry Advances

journal homepage: www.elsevier.com/locate/focha

Effect of pineapple core powder on white finger millet vegan probiotic

beverage: Nutrition, sensory and storage
B Malini a, C K Sunil a, b, *, Ashish Rawson c, R Vidyalakshmi c, N Venkatachalapathy a, b
a
Department of Food Engineering, National Institute of Food Technology Entrepreneurship and Management-Thanjavur (NIFTEM-T), Tamil Nadu, India
b
Centre of Excellence for Grain Sciences, National Institute of Food Technology Entrepreneurship and Management-Thanjavur (NIFTEM-T), Tamil Nadu, India
c
Department of Food Safety and Quality Testing, National Institute of Food Technology Entrepreneurship and Management-Thanjavur (NIFTEM-T), Tamil Nadu, India

A R T I C L E I N F O A B S T R A C T

Keywords: The present study aims to develop, optimize, and evaluate the effective incorporation of pineapple core powder
Pineapple core (PCP) in plant-based probiotic beverages using Lactocaseibacillus rhamnosus (LGG). The optimization was carried
White finger millet out using pH, viscosity, and LGG viable load as response variables in the D-optimal mixer design. In optimized
Millet probiotic beverage
products, the changes in pH, titratable acidity (TA), and LGG viable load are observed during fermentation and
Lactocaseibacillus rhamnosus
Functional food
storage conditions. The optimum condition for developing PCP-added white finger millet probiotic beverage
(PAMPB) was 14:5:2:2 for white finger millet, sugar, PCP, and LGG inoculums, respectively. A PAMPB provided a
higher LGG viable load of 11.34 log (CFU/mL) within 6 h of fermentation time. The PCP effectively increases the
physiochemical properties like total phenolic count (17.64 mg GAE/g), total antioxidant activity, total flavonoid
count (8.59 mg QE/g), and water holding capacity (41.52 %) of the PAMPB. In addition, the different sugar
profiles and new pineapple aroma compounds like ethyl hexanoate and methyl hexanoate are identified in
PAMPB. During the storage of PAMPB at 4⁰C for 21 days, the reduction in LGG viable count was observed from
11.54 to 9.39 log (CFU/mL). Incorporating PCP in millet beverages provides a positive result by increasing
nutritional quality.

Introduction minerals, proteins, dietary fibers, unsaturated fatty acids, and phyto­
chemicals (Arya & Shakya, 2021; Chavan et al., 2018). Moreover,
Millets are known as the poor man crop for a long period and have millet-based probiotic beverages have higher bioavailability and can be
potential nutritional factors but they are underutilized until now. used for disease management like diabetes and obesity. Several studies
Recently, many researchers have worked on converting these millets reported the beneficial health effects associated with millet probiotic
into marketable products. Also, consumer demands are changed from beverages, especially white finger millets (WFM) are highly nutritious
energy to a balanced nutritional diet. In that line, probiotic, nutraceu­ and have higher dietary fiber content than other millets (Meena et al.,
tical, and functional foods gain progressive growth in research and food 2022; Ananthu et al., 2023; Amarnath et al., 2023). Devi et al. (2014)
market lines (Panghal et al., 2017). Probiotics are live microorganisms noted that cereal-based probiotic beverages could reduce low density
used for health benefits, mainly for restoring gut flora (Tu et al., 2019). lipoprotein (LDL) up to 20–30 %, which helps to reduce cardiovascular
The demand for probiotic beverages has tremendous growth due to their risk. KMR-340 is a hybrid variety obtained from WRT-14 (seed parent)
functional property of the probiotic beverage. Till now, most of the and GE-2924 (pollen parent) in WFM (Balakumaran et al., 2023). The
world’s probiotic market is occupied with milk-based probiotic bever­ germinated KMR-340 powder has the potential nutritional value to
ages even though they have drawbacks like lactose intolerance, a prepare a probiotic beverage (Srinivasreddy et al., 2022). However, the
growing preference for a vegan diet, high cholesterol levels, and major limitation of millet-based probiotic beverages is their millet flavor
milk-related allergies. To rectify those demerits, many researchers work and the selection of microorganisms that have generally been regarded
to find alternatives for milk-based probiotic beverages via food grains as safe (GRAS) and have a positive effect on human health conditions
(cereals, millet, legumes), fruits, and vegetables (Meena et al., 2023). (Devi et al., 2014).
Germinated millets are rich in nutritional values like vitamins, Lactobacillus species are the generally used probiotic microorganism

* Corresponding author.
E-mail address: [email protected] (C.K. Sunil).

https://doi.org/10.1016/j.focha.2023.100593
Received 12 September 2022; Received in revised form 13 December 2023; Accepted 21 December 2023
Available online 27 December 2023
2772-753X/© 2023 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-
nc-nd/4.0/).
B. Malini et al. Food Chemistry Advances 4 (2024) 100593

in most applications, in that Lactocaseibacillus rhamnosus are well known Preparation of millet flour
for their beneficial activity in intestinal mucosal, gut microbiota mod­
ulation, and lowering the glycemic load (Terpou et al., 2019). Also, The better quality of 1 kg of WFMs was soaked in distilled water for
probiotics improve the antioxidant property of food materials by 24 h, the excess water drained out, and the grains were placed in the
down-modulating the reactive oxygen species production; in particular, incubator for germination (MAX, model MWS231, New Delhi) for 48 h
the L. rhamnosus strain modulates circulatory oxidative stress, which at 30⁰C. The germinated grains are air dried in a hot air oven at 40⁰C for
helps the cells to protect against carcinogen-induced damage (Syngai 8 hrs with a fixed air velocity of 1 m/s (Everflow scientific instrument,
et al., 2016). Several studies denoted the prebiotic nature of fruits and Chennai). The dried grains are pulverized and sieved using a 250 µm
vegetable waste due to their dietary fiber content, nutrient availability, sieve; the uniform-size particles are taken for analysis.
and fructooligosaccharides (Pop et al., 2021). In pineapple fruit
co-products like peel, pomace, core, and peel exhibit the prebiotic PCP
property, hence the availability of direct carbohydrate sources to helps
microbial growth and leads to the production of several organic acids in The PCP was prepared as described by Sengar et al. (2022). The
yogurt (Sah et al., 2016). Within pineapple co-products, nearly 9–10 % pineapple was peeled and the inner core was sliced into thin slices and
of pineapple core was discarded as waste due to their high poly­ freeze-dried (Penguin Classic, Lark Innovative, Chennai, India) for 36 h
saccharide and fiber nature, which can have higher nutritional avail­ to get the dried pineapple core. And then, the dried sample is pulverized
ability during steaming under pressure and can be used as a better and sieved using a 500 µm size sieve and stored at − 20⁰C for further
prebiotic source for L. rhamnosus growth (Buvaneshwaran et al., 2022; analysis.
Hadidi et al., 2020; Sengar et al., 2022). One important rationale for
using pineapple core waste is that during fermentation, various flavor Development of PCP added millet based probiotic beverage (PAMPB)
active and bioactive compounds are produced; in the case of millet
beverage, the millet flavor has a negative impact on consumer prefer­ The millet-based probiotic beverage developed as described by
ence, so pineapple flavor can be added to mask the millet flavor of the Navyashree et al. (2022a) using 14 % (w/v) germinated millet flour
beverage. Adding pineapple co-products to beverages affects their (GMF), 5 % (w/v) of sugar, and 1–3 % of PCP (v/v) were mixed with
physicochemical property (viscosity and acidity) and fermentation time 500 mL of distilled water and sterilized. It was cooled for 15 min, and
due to their hygroscopic nature and prebiotic effect, which can have an 1–3 % (v/v) of LGG was inoculated and incubated at 37⁰C for 6 h in a
impact on the overall quality of the beverage (Buvaneswaran et al., shaking incubator at 150 rpm as described by Srinivasreddy et al.
2023; Srinivasreddy et al., 2022). Therefore, the addition of pineapple (2022). The samples were drawn out every hour and stored at 2⁰C for
co-products to millet-based beverages needs to be optimized. The effect further analysis. For analysis, the incorporation of PCP and LGG inoc­
of pineapple co-products on probiotic growth in beetroot juice and ulation concentration is taken as an independent variable. The probiotic
yogurt was studied by researchers, and it has a positive effect on pro­ beverage without PCP and 1 % LGG inoculum addition were taken as a
biotic growth (Panghal et al., 2017; Sah et al., 2016). control.
Only a few studies used pineapple co-products as a prebiotic source
to improve the bio-functionality of probiotic millet beverages (Srini­ Determination of response variables
vasreddy et al., 2022). Therefore, this study aimed to explore the pos­
sibility of utilizing pineapple core waste as a prebiotic source to produce To study the effect of different LGG inoculums and PCP % on the
millet-based probiotic beverages and also analyze the effect of pineapple developed probiotic beverage, the titratable acidity, pH, viscosity, and
core powder (PCP) on millet-based probiotic beverage physicochemical, viable bacterial count (LGG) were taken as response variables.
and functional properties.
Titratable acidity (TA) and pH
Materials and methods A calibrated portable pH meter is used to measure the pH of the
probiotic beverage (Hanna Instruments, USA). Titratable acidity (TA)
Raw materials procurement was determined according to the procedure described by Tu et al.
(2019), by titrating 10 mL of a sample against 0.1 N NaOH until ach­
The KMR 340 variety WFM was procured on March 2023 from the ieved the pH of 7.
College of Agriculture, V.C. Farm, Karnataka, India. Pineapples were
purchased from the local market in Thanjavur, Tamil Nadu. All other Viscosity
chemicals used in the analysis are analytical grade and purchased from The viscosity of the samples was measured by a brook field
Sigma Aldrich and Hi-media, India. Lactobacillus rhamnosus ATCC53103 viscometer as described by Srinivasreddy et al. (2022), using the SC4–18
(LGG) NCDC 347 was procured from the National Dairy Research spindle, in the viscosity range of 1.5–30 KmPa.s at 25 ± 1⁰C at 50 rpm.
Institute (NDRI), Karnal, India. Initially, the viscometer with SC4–18 spindle assembly was calibrated
using Brookfield’s Silicon Viscosity Standard fluids (viscosity values of
Revival of lactobacillus culture 9.7 and 97 cP at 25±1⁰C). The equipment accuracy was found to be ±2
% of these standard viscosity values. 5–10 mL of probiotic beverage was
The lactobacillus culture was revived as per the procedure in the taken to analyze viscosity (measured in centipoises (cP) using SC4-18
protocol. A 0.1 mg of freeze-dried LGG culture was added with 50 mL of spindle, at the temperature of 25±1⁰C, at 50 rpm.
De Man, Rogosa, and Sharpe (MRS) broth and incubated at 37⁰C for 24
to 48 h. A 10 mL was taken from this, added with 100 mL of MRS broth, Viable cell count
and incubated under the same conditions as the prepared culture used To enumerate the LGG viable counts the spread plate method as
for the analysis. The purity of the colonies was cross-checked by the described by Yahyaoui et al. (2017) was used. The probiotic beverage
gram staining technique. The LGG culture has 7 log (colony forming unit was diluted 109 times, and the 0.1 mL of final dilution was spread on
(CFU) /mL) used for the inoculation. The density of the colony was MRS agar plates using a sterilized glass rod. Sterile peptone water is used
observed using an Ultraviolet-visible (UV) spectrophotometer (UNICO for the dilution. The plates were incubated at 37⁰C for 48 h for bacterial
spectrophotometer, Kyoto, Japan) at 660 nm. (LGG) enumeration. After incubation, the colonies in the plates were
counted manually and expressed as CFU per mL of a sample using Eq.
(1).

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

( )
CFU CFU prepared by mixing an equal volume of a 7 mmol/L ABTS stock solution
Log = Log10 (1)
mL Dilution factor ∗ Aliquot with a 2.45 mmol/L potassium persulfate solution, the mixture was then
diluted with 10 mmol/L phosphate-buffered saline (pH 7.4). Then, 30 µL
of the sample was added to 3 mL of diluted ABTS+ solution. radical and
Physiochemical analysis incubating for 6 min in a dark room. The absorbance of sample and
control is measured at 734 nm (A1). The radical scavenging activity is
Proximate analysis calculated by Eq. (3).
Proximate composition, namely moisture, ash, fat, protein, and
crude fiber was calculated using method numbers 931.04, 923.03, ABTSradicalscavengingcapacity(%) = [(A0 − A1) / A0] × 100% (3)
945.38, 2001.11, and 985.29 in AOAC (2019), respectively. Total car­
where A0 is the absorbance of the control and A1 is the absorbance of
bohydrate was determined by the difference method, while protein
the sample
content was obtained after multiplying the nitrogen content by 6.25 (the
conversion factor).
Water holding capacity
The water holding capacity (WHC) of the probiotic beverages was
Total solid content (TSS)
calculated as described by Dimitrellou et al. (2021) using Eq. (4).
TSS (⁰B) was measured using a hand-held refractometer (0–32⁰Brix)
to analyse the % Brix of sugar in 250 µL of the sample (Navyashree et al., 100 ∗ (B − WE)
WHC (%) = (4)
2022a). B
Where WE = water expelled in g and B = initial probiotic beverage in
Reducing sugar
g. Water expelled (WE) quantity was obtained and weighed after
A 3,5-dinitro salicylic acid (DNS) colorimetric method was used for
centrifugation of 10 g of probiotic beverage (B) at 438 × g for 10 min at
the reducing sugar analysis as described by Ujiroghene et al. (2019). A 1
20⁰C.
mL of sample was added with the same volume of DNS and boiled for 5
min. The resulting mixture was cooled at 25⁰C and added with 10 mL of
Suspension stability
distilled water. Absorbance was recorded at 540 nm. A known concen­
The suspension stability of the millet-based probiotic beverage was
tration of glucose standards is used to calculate the reducing sugar
calculated based on Arya and Shakya (2021). Eq. (5) and Eq. (6) are used
concentration.
for calculating the separation index and percentage of separation,

Total phenolic content (TPC) Ht

Separation index (SI) = (5)
The TPC in the beverage was calculated as described by Poveda et al. H0
(2019) with some modification, 100 µL of samples were added with 500
µL of Folin-Ciocalteu reagent and 6 mL of distilled water. After 3 min of Percent separation = SI × 100 (6)
incubation 1.5 mL of 20 % Na2CO3 and 10 mL of water. The mixture was Where Ht is the height of the lower phase after the storage time ‘t’ (10
kept in the darkroom for 2 h; after that, absorbance was measured at days) and H0 is the initial height of the beverage.
760 nm using a UV spectrophotometer. Gallic acid was used for the
standard curve preparation; the TPC was expressed as milligrams of Fourier transform infrared spectroscopy (FTIR)
gallic acid equivalent per gram (GAE/g) of dry weight.
The FTIR instrument was used to characterize the probiotic beverage
Total flavonoid content (TFC) using the method described by Buvaneswaran et al. (2021) with slight
TFC was determined as described by Sengar et al. (2022) with simple modification. The transmittance in IR spectra was recorded using an
modifications, 1 mL of ethanol, 100 µL of AlCl3, and 100 µL of potassium FT-IR spectrophotometer, PerkinElmer, USA (Spectra Two). The results
acetate are added with 1 mL of sample, and the final volume of mixture were obtained at a resolution of 4 cm− 1 and 32 scans in a wavelength
made up of 1.5 mL and incubated at room temperature for 30 min. range of 450 to 4000 cm− 1. The data were assessed using Origin Pro8
Absorbance was measured at 415 nm, and the values were compared software (Origin-Lab, Northampton, MA, U.S.A.).
with the quercetin standard curve and expressed as quercetin equivalent
per gram (QE/g) of dry weight. Sugar analysis

DPPH radical scavenging activity The sugar analysis in a probiotic beverage was analyzed by the High-
The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging ac­ Performance Liquid Chromatography (HPLC) method described in
tivity of probiotic beverages was examined according to the procedure Freire et al. (2017) with some modifications. A Shimadzu liquid chro­
described by Srinivasreddy et al. (2022). Control for the analysis was matography system coupled with a refractive index detector 1260 In­
prepared using methanol (0.1 mL) and DPPH (3.9 mL). The absorbance finity II (for alcohol and carbohydrates) was used for the sugar analysis.
for samples and control were recorded at 517 nm (OD) to determine the Zorbax NH2 Analytical (4.6 × 250 mm x 5 µm) column, sample injection
free radical scavenging activity. The DPPH radical scavenging activity is volume 5 µL, an isocratic solvent system consisting of (75:25 v/v) of
calculated using Eq. (2). acetonitrile and HPLC water at 0.9 mL/min flow rate at a column oven
(OD control − OD Sample) temperature of 35 ◦ C was used for the analysis.
DPPH radical scavenging activity (%) = × 100
OD control
(2) Gas chromatography-mass spectrometry (GC–MS) analysis

where OD control is the absorbance of the control and OD sample is the The phytochemical compounds were analyzed using gas chroma­
absorbance of the sample. tography coupled with a mass spectrometer (8890GC/5977B GC/MSD-
Agilent). The probiotic beverage (1 g) was extracted using methanol
ABTS radical scavenging activity (75 mL) at 60⁰C for 50 min. The extract was filtered through a 45 µm
The 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) dia­ filter, then the sample extract of 2 µl was injected into the column (Rtx-
mmonium salt (ABTS) was used to calculate the radical scavenging ac­ 5MS (5 % Diphenyl / 95 % Dimethylpolysiloxane - 30 m x 0.25 mm ID x
tivity as described by Freire et al. (2017). The ABTS+solution was 0.25 µm) at 280⁰C, helium used as a carrier gas. A 1 µL injector in split

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Fig. 1. Effect of factors like PCP and LGG microbial inoculums (MI) on response variables like (A) pH, (B) viscosity, and (C) Lactocaseibacillus rhamnosus (LGG) viable
load using optimal mixer design.

mode with split ratio 1:50 and the initial column temperature was titratable acidity, and LGG viable cell count.
100 ◦ C rising 280 ◦ C at a rate of 5 ◦ C/min, the temperature was main­
tained for 3 min and changed to 150 V at 4 ◦ C/min. The temperature was Data analysis
further increased to 280 ◦ C at a rate of 15 ◦ C/min with a hold time of 35
min. The ion source of the mass spectrometer was held at 230 ◦ C with an Physicochemical characteristics of probiotic beverages were
interface temperature of 270 ◦ C. The MS spectra will be observed in full analyzed using one-way ANOVA using Tukey’s methods with a proba­
scan mode from m/z 40 to 650 at a scan time of 250 ms. Detection was bility P < 0.05 in SPSS software 20.0 (International Business Machines
performed in full scan mode. The identification of the compounds was Corp., Armonk, New York). All the experiments were triplicated to
achieved by comparing obtained mass spectra of unknown peaks with reduce the error. Similarity maps of images were drawn using the
those stored in the NIST08 (National Institute of Standards and Tech­ component scores. Interpretation of components was obtained by
nology) and Wiley mass spectral electronic libraries. looking at the linear correlation between the original variables (called
loading factors).
Electronic-nose (E-nose) aroma profile
Results and discussion
The volatile compounds in the probiotic beverage were analysed
using E nose based on the flash GC system (Heracles Neo II, Alpha MOS, Optimization of PCP added white finger millet based probiotic beverage
Toulouse, France) as described by Sengar et al. (2022). A 2 g of (PAPMB)
freeze-dried sample was placed in 20 mL standard vials with 20 mm
polytetrafluoroethylene (PTFE) septa. Samples were incubated at an To optimize the pineapple core added millet-based based probiotic
oven temperature of 40 ◦ C for 20 min with pulsed agitation at 500 rpm beverage d-optimal mixture design was used. A different component of
(5 s ON: 2 s OFF). The accumulated gases during incubation were the mixture (PCP % and LGG inoculum %) was taken as the factors, and
collected and injected (5 mL) into GC at 50⁰C for 90 s with a flow rate of the responses (pH, viscosity, LGG viable load, TPC, DPPH, ABTS) were
500 µL/s. These were injected into the GC injector at 200⁰C, injection taken as functions. Optimization was done based on the response vari­
volume of 2500 µL at the rate of 125 µL/s (Dharini et al., 2022; Srini­ able. A polynomial model was created using Response surface method­
vasreddy et al., 2022). Volatile compounds and their odor descriptions ology (RSM) relating the response of the probiotic beverage individually
were identified using the database AroChemBase (version 8, Alpha MOS, to different factors. The effect of different factors on the response vari­
Toulouse, France). The chromatographic approaches were performed able is shown in Fig. 1. The response surface graphs (Fig. 1(A)) present
using Alpha Soft (version 17, Alpha MOS, Tou-louse, France). the effect of PCP and LGG on the pH of PAMPB. The results revealed that
the pH value decreased as PCP and LGG increased, and pH was signifi­
Shelf life evaluation cantly (p < 0.05) affected by linear terms of PCP and LGG. The decrease
in pH denotes the production of organic acids during fermentation
The fermented probiotic beverage was subjected to a shelf life study which indirectly denotes the higher rate of fermentation at higher PCP
as described by Gupta et al. (2010). The probiotic beverage and control and LGG conditions. It may be due to the action of PCP as an extra
sample was stored at 4⁰C for 21 days, every 3 days once one bottle of carbon source and provide fructooligosaccharides, which act as pre­
beverage was withdrawn from each sample and analyzed for pH, biotics to accelerate the growth of probiotics and enhance the

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Fig. 2. Changes in (A) pH and titratable acidity (TA), (B) Lactocaseibacillus rhamnosus (LGG) and Reducing sugar (RS) during the fermentation period.

production of organic acids, especially lactic acid. Similar effects were the fermentation of PAMPB. Initially, the control and PAMPB have a
also reported by Srinivasreddy et al. (2022) in a pineapple peel added similar growth rate of L. rhamnosus; after 90 min, the growth rate of the
probiotic beverage. The value of viscosity varied from 35 to 48 cP, for probiotic was increased than the control. After 300 min in both samples,
the addition the PCP and LGG viscosity increased significantly with the growth rate was decreased and maintained at the same level; maybe
fermentation time (Fig. 1(B)). it denoted the lag phase of the L. rhamnosus due to the acid condition
Compared to LGG, the higher the PCP addition, higher the change in and other environmental factors. When the microbial load increased, the
viscosity was observed, it may be due to the dietary fibers in PCP. The increase in reducing sugar content was observed in PAMPB. Initially,
presence of xylooligosaccharides strengthens the protein network and small changes in RS were observed; when the LGG growth accelerated
enhances the water retention capacity (Van Doan et al., 2021). Also, after 90 min of fermentation, the production of reducing sugars also
higher probiotic biomass at higher PCP content enhances the biomass increased gradually. At some point (after 300 min) small negative de­
density and helps to improve the viscosity. The fruit waste enhances viation from its original line was observed; it may be caused by the
microbial growth due to the presence of fructose and dietary fiber, when consumption of RS by the L. rhamnosus. This deviation was higher in the
the PCP concentration increased from 1 % to 3 % at constant LGG PAMPB sample because the LGG load at PAMPB (11.54 log CFU/mL) is
inoculation (1 %), the viable probiotic count increased from 10.6 to 11.2 much higher than in the control (9.53 log CFU/mL). Similar kind of
log (CFU/mL) (Fig. 1(C)). The positive enhancement in probiotic growth changes in pH, TA, and probiotic load were observed by Yahyaoui et al.
by PCP may be due to the growth factors like dietary fibers, minerals, (2017) in the traditional cereal food “Bassica” using L. rhamnosus.
and proteins in PCP. Similarly, Sah et al. (2016) observed an increase in
the probiotic population from 0.3 to 1.4 log cycle in yogurt by the
addition of pineapple co-product. Based on the experimental data and Effect of PCP on a response variable
goal setting the best combination of PCP and LGG concentration for
producing PAMPB was identified. Minimization of overall fermentation The response variables like TA, pH, viscosity, and LGG viable load
time and maximization of TPC and AOA at the goal set point of viscosity, (log CFU/mL) at different factors are listed in Table 1(A). Several studies
pH, and probiotic load were taken for the optimization. The recom­ reported that adding fruit by-products stimulates the growth of pro­
mended pH and probiotic load in probiotic products are 4 to 4.5 log biotics like LGG due to the availability of free carbon sources like
(CFU/mL), respectively. Based on that, the optimized composition for fructose and soluble dietary fiber (Chawafambira et al., 2020; Liu et al.,
the PAMPB formulation was 14:5:2:2 for WFM, sugar, PCP, and LGG 2021). Results show that the increase in addition of PCP and LGG %
inoculums, respectively. accelerates the reduction of the pH of the probiotic beverage, the pH and
TA of the probiotic beverage vary from 3.84 to 4.9 and 0.43 to 0.93 %,
Changes in pH, TA, reducing sugar, and microbial load during fermentation respectively after 6 h of fermentation. The addition of PCP significantly
time in optimized PAMPB reduces the pH and increases the TA of the millet probiotic beverage.
The changes in pH and TA during PAMPB fermentation are shown The production of lactic acid during fermentation by LGG reduces the
respectively in Fig. 2(A). In both control and PAMPB, the abrupt pH pH. A similar kind of result was reported by Yahyaoui et al. (2017) in
reduction from 6.8 to 6.2 (control) and 6.4 to 5.82 (PAMPB) was beetroot probiotic beverages. On the same hand, the LGG viable load
observed from the time of inoculums addition until 30 min. Once the pH was considerably higher in the PCP-added sample; the addition of PCP
of a probiotic beverage reaches nearly 5.4 the decline of pH slows down and LGG % have a positive trend on LGG viable load. The maximum
and remains in the linear range. In the case of PAMPB, the second sharp reduction of pH, higher TA (%), and LGG viable load were obtained at a
drop in pH was identified after 150 min of fermentation. On the other 3:3 (PCP: LGG %) combination. The increasing glucose conversion rate
hand, the titratable acidity keeps on increasing throughout the to gluconic acid may be due to the higher lactobacillus concentration.
fermentation time. A gradual increase in titratable acidity was observed This gluconic acid again converts into acetic acid and increases the pH of
until 120 min after a sudden decrease of TA occurred in both controls in the probiotic beverage. Increasing titratable acidity was observed while
PAMPB; the reduction in TA was noted. The changes in the trend of pH pH decreased. Sah et al. (2016) stated that the PCP itself contains a
reduction and TA (%) may be because of the generation of organic acids considerable amount of fermented sucrose, so the PCP concentration
in the initial period, when time goes on, the stability of the organic acids provides the prebiotic fructooligosaccharides to the LGG, improving the
is maintained in further hours of the fermentation period. The same kind lactic acid production, further reducing the pH of the probiotic
of trend line of pH in soy whey-based novel functional beverages were beverage. The viscosity of the samples significantly (p < 0.05) varied
observed by Tu et al. (2019). from 35 to 48 cP. The viscosity results depict that the addition of PCP
Fig. 2(B) shows the reducing sugar and LGG viable load trend during alters the intermolecular interaction among molecules, and the higher
the PCP, the higher the viscosity range of probiotic beverages identified.

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 B. Malini et al.
Table 1
(A) Physiochemical properties of beverages prepared from various PCP and LGG concentrations, (B) Proximate composition of pineapple core powder (PCP), white finger millet (WFM), and PCP added white finger millet
probiotic beverage (PAMPB).
PCP (%) : LGG (%) pH TA % LGG viable load V (cP) TSS (⁰B) TPC (mg TFC (mg ABTS (%) DPPH (%) RS (%) WHC (%) SI (%)
(CFU/mL) GAE/g) QE/g)

0:1 4.90 0.43±0.03h 9.54±0.04h 35.36 8.35±0.02b 15.28 5.13±0.1f 54.15±0.03d 51.72 0.67±0.01e 18.33 4.567±0.01a
±0.01a ±0.04h ±0.04e ±0.03e ±0.02b
1:1 4.80 0.51 10.64±0.02g 36.38±0.02g
5.80±0.03c 17.47 8.24±0.01d 69.34±0.01c 64.12 0.68±0.01e 41.45 3.411±0.00b
±0.02a ±0.01g ±0.15c ±0.1d ±0.02a
1:2 4.71 0.59±0.00f 11.03±0.02f 38.82 8.51±0.01b 17.62 8.19±0.01e 69.53±0.01c 64.25 0.77±0.01d 41.48 2.960±0.02bc
±0.01b ±0.02f ±0.02b ±0.2d ±0.01a
1:3 4.21 0.77±0.01d 11.36±0.01d 43.55 8.75±0.01b 18.34 8.24±0.01d 69.92±0.01c 64.94 0.89±0.01d 41.43 1.843±0.01c
±0.02d ±0.02d ±0.01a ±0.1d ±0.02a
2:1 4.81 0.54 11.03±0.02f 41.08±0.01e 8.48±0.01b 17.33 8.59±0.03c 70.84±0.01b 69.85 1.07±0.01c 41.52 2.767±0.02c
±0.01a ±.00g ±0.01d ±0.1c ±0.01a
2:2 4.52 0.67±0.01e 11.34±0.01d 43.48 8.60±0.01b 17.64 8.59±0.01c 70.92±0.01b 70.12 1.35±0.01b 41.52 2.863±0.01c
±0.01c ±0.01d ±0.01b ±0.1b ±0.01a
2:3 3.95 0.86±0.01b 11.72±0.02c 45.13 9.04±0.01a 18.36 8.58±0.01c 71.03±0.02ab 70.22 1.38±0.01b 41.52 1.550±0.02d
±0.15e ±0.01c ±0.02a ±0.03b ±0.01a
6

3:1 4.20 0.81±0.00c 11.14±0.01e 45.05 8.85±0.01b 17.45 9.52±0.01b 71.54±0.01a 72.37 1.42±0.01a 41.54 2.134±0.00c
±0.05d ±0.02c ±0.10c ±0.02a ±0.01a
3:2 4.35 0.74±0.01d 11.85±0.02b 46.37 8.64±0.01b 17.68 9.56±0.02b 71.67±0.01a 72.46 1.47±0.02a 41.57 2.072±0.01c
±0.04d ±0.02b ±0.01b ±0.02a ±0.00a
3:3 3.84 0.93±0.01a 12.54±0.01a 48.22 9.23±0.02a 18.51 9.68±0.04a 71.82±0.01a 72.54 1.47±0.02a 41.53 1.371±0.00d
±0.04e ±0.02a ±0.05a ±0.00a ±0.02a

Parameters Pineapple core powder (PCP) White finger millet (WFM) White finger millet beverage (Control) Optimized white finger millet beverage (PAMPB)

Moisture 6.49±0.09c 6.17±0.04c 84.91±0.27b 86.70±0.29a

Protein 3.79±0.21d 7.33±0.11a 4.38±0.03c 5.32±0.07b
Fat 1.47±0.13a 1.10±0.01b 0.82±0.02c 0.81±0.02c
Ash 2.19±0.03a 0.86±0.06b 0.42±0.01c 0.45±0.01d
Crude fiber 1.26±0.03b 5.12±0.01a 0.78±0.03c 0.80±0.02c
Carbohydrate 53.83±0.51b 71.13±0.17a 9.30±0.10c 8.22±0.07d

Values are represented as mean ± standard deviation.

Different superscripts in the same column differ significantly (p < 0.05).

Food Chemistry Advances 4 (2024) 100593

Values are represented as mean ± standard deviation.
Different superscripts in the same row differ significantly (p < 0.05).
B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Arya and Shakya (2021) observed a similar kind of increasing viscosity inoculums (%) have the maximum impact on reducing sugar. When the
trend in prebiotic multigrain functional beverages. TSS and viscosity are higher LGG inoculum (%) was added to the sample, the reducing sugar
related to the stability of the bioactive compounds in plant-based content in the probiotic beverage decreased. The reduction in reducing
products. sugar is due to the consumption of sugar molecules by LGG for their
metabolic activity was noted by Tu et al. (2019). Similarly, Panghal,
Effect of PCP on physicochemical property Virkar and Kumar (2017) noted the reducing trend of sugar content in
beetroot juice due to microbial fermentation.
The effect of addition of PCP on different physicochemical properties
of beverages: proximate composition, TA (%), TSS (⁰B), viscosity, total Effect on antioxidant activity, total phenolic content, and total flavonoid
phenolic, antioxidant activity, total flavonoid content, reducing sugar, content
water holding capacity, and suspension stability are shown in Table 1. The antioxidant activity of probiotic beverages was calculated by
both DPPH and ABTS radical scavenging activity methods. In both
Effect on proximate composition methods, the antioxidant activity of the probiotic beverage was signif­
The proximate composition of raw materials and millet probiotic icantly (p < 0.05) different from the control sample. The control con­
beverages is listed in Table 1(B). A similar kind of proximate composi­ tains a DPPH activity of 51.72 %, and the PCP incorporated probiotic
tion of White Finger millet and PCP is obtained by Navyashree et al. beverage’s DPPH range from 64.12 to 72.54 %. Similarly, the control
(2022b) and Sengar et al. (2022). The PCP contains fat, ash, and crude contains 54.15 %, and PCP incorporated samples contain nearly 69.34 to
fiber content of 1.47 %, 2.19 %, and 1.26 %, respectively; similar kind of 71.82 % of ABTS activity. At different PCP and LGG inoculums, a sig­
results were obtained by Sengar et al. (2022). Ash content indicates the nificant rise in antioxidant activity was observed while rising in the PCP
presence of mineral, its value is less compare to the pineapple crown incorporation irrespective of the LGG inoculum content. The results
co-product (Byresh et al., 2023). Crude fiber content represents the represent the increase in antioxidant activity of the millet probiotic
presence of cellulose, hemicellulose, and holocellulose in PCP. Crude beverage during PCP incorporation. Pineapple co-products contain
fibers are a good source to enhance the growth and survival of probiotics phenolic compounds and ferulic acid, which can act as an antioxidant.
during storage (Sah et al., 2016). Results represent that the addition of Saraswathy et al. (2017) reported that the Inhibition Concentration
PCP does not affect the probiotic beverage’s fat, ash, and crude fiber (IC50) value of antioxidant activity of dried pineapple co-product ex­
content, so the control and PAMPB contain no significant difference tracts was 0.8–1.3 mg/mL, which denotes the higher antioxidant ac­
between them A significant increase in crude fiber content was observed tivity of the co-products. It suggests that PCP can retard the OH induced
in the cereal bars and probiotic beverage incorporated with pineapple destruction and beverage spoilage. The antioxidant activity of the
co-products like peel and crown, respectively (Damasceno et al., 2016; product has a positive correlation with the TPC and TFC. The maximum
Byresh et al., 2023). However, the moisture content of the PCP added antioxidant activity was obtained in the 3 %:3 % PCP: LGG combination.
PAMPB (86.70±0.29 %) comparatively increased than the control In the case of TPC and TFC, even the low (1 %) addition of PCP
sample (84.91±0.27 %); it may be due to the hygroscopic nature of PCP, creates a significant rise in the probiotic beverage than the control
so it has a strong bond with the moisture molecules. The ratio of soluble sample (Table 1(A)). TPC and TFC in PAMPB increased from 17.47 to
and insoluble fiber in the pineapple co-products significantly affects the 18.51 mg GAE/g and 8.24 to 9.28 mg QE/g, respectively. Results indi­
water retention capacity of beverages. A high carbohydrate content cated that PCP-added probiotic beverages had higher TPC and TFC than
(53.83±0.51 %) in PCP indicates the presence of high sugar content, the control sample, even at low (1 %) incorporation of PCP. Further
which was comparatively lower than other pineapple co-products like addition of PCP again increases the TPC and TFC. The PCP contains
peel (75.64 %) and crown (72.91 %). A previous study on pineapple peel phenolic and flavonoid compounds like catechin, epicatechin, gallic
powder (PPP) by Srinivasreddy et al. (2022) reported that PPP does not acid, and ferulic acid. The phenolic compounds are composed of the
affect the carbohydrate content, but in this significant decrease in car­ number and position of hydroxyl groups and can donate hydrogen
bohydrate content was identified then the control sample (from 9.30 atoms, this unpaired electron supporting capacity exhibits its antioxi­
±0.10 % to 8.22±0.07 %), the consumption of mono or disaccharides by dant property. The higher antioxidant activity in PAMPB may be due to
the LGG can lead to the low carbohydrate content. A reduction in car­ the presence of higher phenolic and flavonoid compounds. Most flavo­
bohydrate content in PAMPB might indirectly represent the higher noid compounds in plant materials are combined with protein, fat, and
amount of probiotic load. Significant (p < 0.05) improvement in protein other soluble fibers. The metabolic activity of probiotic LGG can cause
content was observed in PAMPB than the control sample. the release of phenolic and flavonoid compounds, the breakdown of
In the study by Chawafambira et al. (2020), the L. rhamnosus fer­ complex phenolic and antioxidant compounds by enzymes released
mented fruit jam contains higher carbohydrate, protein, ash, and crude during fermentation can further increase the TPC and TFC. For example,
fiber; reduction in fat content than the control sample. Hence, pineapple Lactobacillus breaks complex tannin acid into simpler phenolic com­
co-products (core) contain a considerable amount of minerals (Ca, Mg, pounds like gallic acid and pyrogallol (Byresh et al., 2023). Further, the
K, and P), and dietary fiber, it can be considered a potential prebiotic phenolic compounds in PCP have potential anti-thrombotic and
component for probiotic bacteria; also, pineapple core contains essential anti-inflammatory properties.
amino acids like valine, isoleucine, lysine, histidine, and threonine (Sah Chandrasekara and Shahidi (2012) stated that during microbial
et al., 2016; Sengar et al., 2022). Sah et al. (2016) reported that pine­ fermentation breakdown of these plant molecules by the β-glucosidase
apple co-product contains dietary fiber content in the range of enzyme, which was secreted during the fermentation process, helps to
44.90–58.48 g/100 of dry matter, which was near to the fiber rice increase the TFC in the fermented samples. In other cases, the acid
fraction in wine pomace and passion fruit seed fibers, the high fiber produced during the metabolic activity of microorganisms produces new
content and essential mineral contents help to utilize pineapple flavonoid compounds (Tu et al., 2019). Breakdown or release of
co-product as a prebiotic source for probiotic bacteria like LGG. phenolic and flavonoid molecules in PCP during fermentation by the
metabolic activity of lactic acid bacteria enhances the TPC and TFC in
Effect on TSS and reducing sugar PAMPB, most of the phenolic compounds contribute to the antioxidant
The TSS for all combinations significantly (p < 0.05) varied from properties which helps to enhance the antioxidant property of the same.
5.80 to 9.23 ⁰B. In the PCP-added samples, the increasing trend of TSS Similar kind of results were found in pineapple co-product added to
was identified. The reducing sugar content significantly (p < 0.05) green tea, where a significant improvement in TPC content was noted by
varied from 0.67 to 1.47 %, and adding PCP and LGG inoculums % Kaur and Mittal (2018). Hadidi et al. (2020) reported that the poly­
decreased the reducing sugar content. But compared to PCP, LGG saccharides present in pineapple core provide excellent scavenging

7
B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Fig. 3. Fourier transform infrared (FTIR) analysis of pineapple core powder added millet probiotic beverage (PAMPB).

activity on DPPH and hydroxyl radicals. Sah et al. (2016) reported that
Table 2
in yogurt, the potent antioxidant peptides from milk proteins acted as
Sugar profile in PAMPB by HPLC.
hydrogen or electron donors in the presence of pineapple co-product
(peel and pomace powder) and formed stable products; also found a Compound Control (mg/L) Optimized PAMPB (mg/L)
significant difference in *OH scavenging activity based on the different Glucose 169.613 279.216
part of pineapple co-product powder. Galactose 770.573 547.955
Fructose 86.337 337.202
Sucrose 14,375.056 9027.896
Effect on water holding capacity and suspension stability Maltose 57.827 63.326
The water holding capacity and suspension stability of the probiotic
beverage can help to study the stability of the beverage. The PCP-added
samples contain significantly higher WHC (41.45 %) than the control wavelength of 3514 cm− 1C– –C–H stretch due to the metabolic activity
(18.33 %). However, the different level of PCP and LGG inoculum of L. rhamnosus was not identified in the results. To fingerprint the
concentration does not affect the WHC of the PAMPB. Results state that bacteria, the specific markers are identified at 1640–1730 cm− 1 (C––O
the addition of PCP alters the binding nature of the WFM to the water stretching vibrations). The water molecules provide the stretching at
molecules during fermentation. The effect of PCP on the suspension 3572.17 cm− 1 (Vodnar et al., 2010). The carbonyl stretch peaks were
stability of the control and PAMPB was studied for 10 days by using the found at 1726.29 cm− 1 in the PAMPB and higher intensities in the
separation index (SI, %). The SI of the control sample and PAMPB lies control samples (Adeyinka et al., 2016). The PAMPB consists of the
between 4.567 % and 3.411–1.371 %. It was noted that the control alkane (C–H) group at 858.32 cm− 1, which was absent in the control
sample had two layers of water and bulky solids during storage of 10 sample. Moreover, the vibration at 1629.85 cm− 1 denotes cyclic alkenes
days. In the PCP-added samples, the separation was minimum compared stretching (C––C). Vibrations from 925 cm− 1 and 669 cm− 1 detail the
to control samples. So, the WHC and SI of the PAMPB imply the stability angular deformation of aromatic hydrogens. The asymmetric stretch of
of the probiotic beverage during the storage condition and the higher C–H bonds at 3625–3846 cm− 1 was only identified in the PCP-added
water holding capacity. The fructose-fructose and fructose-glucose samples. The changes between control and PAMPB samples indicate
glycosidic bonds contain glycosidic linkages, which can provide better the presents of new compounds or structural changes of the existing
stability (Arya & Shakya, 2021). compounds due to the addition of PCP in the millet beverage.

Fourier transform infrared spectroscopy (FTIR) analysis Analysis of sugar compounds using HPLC

The FTIR analysis identified modification in a functional group due To analyze the changes in the sugar content in the probiotic
to the addition of PCP. The functional groups were identified by beverage, HPLC analysis was performed, and the results were presented
comparing the wavelength (cm− 1) and transmittance percentage in Table 2. The sucrose content was identified as 14,375 mg/L and 9027
(Fig. 3). Maximum transmittance observed in the wavelength range of mg/L in the control and PAMPB, respectively. It shows the lactobacillus
349–352 cm− 1 denotes the hydroxyl (O–H) group presence in both the preference toward sucrose for their consumption. But the disaccharide
control and PAMPB samples. Similar to the alcohol, aldehyde, and esters maltose content was higher than in control samples. Alemneh et al.
groups were identified in the 1145.72, 1076.26, and 925.83 cm− 1 in the (2021) explain the likability of lactobacillus species (LGG) towards
pineapple-added samples but not in the control samples. A similar kind monosaccharide (glucose) than disaccharide (maltose), but in this study,
result was identified by Srinivasreddy et al. (2022) in pineapple peel the sucrose content was getting reduced to the fermentation by LGG
powder added millet-based probiotic beverage, but vibration at a growth. However, the monosaccharides like galactose, glucose, and

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Table 3
Chemical composition of PAMPB by GCMS.
Compound Peak (min) Area (%) Control Peak (min) Area (%) PAMPB

Thymine – – – 3.7704 4.04 ✓

D-Alanine, N-propargyloxycarbonyl-, isohexyl ester 3.827 10.15 ✓ – – –
Butanoic acid – – – 4.9777 0.55 ✓
5-Hydroxymethylfurfural – – – 6.0764 3.36 ✓
Melezitose 6.268 2.15 ✓ – – –
D-Tagatose – – – 6.8946 1.17 ✓
5-Chlorovaleric acid – – – 6.5799 0.54 ✓
2,4,4-trimethylpentyl ethylphosphonofluoridate 8.126 8.60 ✓ – – –
Z-8-Methyl-9-tetradecenoic acid 8.377 2.00 ✓ – – –
Sucrose 9.185 3.84 ✓ – – –
β-d-Glucopyranose – – – 9.2178 1.97 ✓
Lactose 9.936 1.76 ✓ – – –
3-Deoxy-d-mannoic lactone – – – 11..6329 9.82 ✓
D-Glucopyranuronic acid 12.195 1.34 ✓ 9.7900 1.94 ✓
N-8-Guanidino spermidine – – – 13.9327 0.44 ✓
n–Hexadecanoic acid 15.684 17.54 ✓ 15.6322 0.10 ✓
d-Mannose 15.804 1.30 ✓ – – –
6-Octadecenoic acid 18.167 38.81 ✓ 18.3845 0.22 ✓
9-Octadecenamide 24.2479 0.58 ✓
Acetate – – – 27.957 6.91 ✓
γ-Sitosterol 37.478 5.38 ✓ 37.3302 1.06 ✓

fructose are increasing. Maybe it is due to the breakdown of di­ compounds like 5‑hydroxy methyl furfural, aroma compounds like n-
saccharides during the fermentation period. The sucrose content in hexadecanoic acid, and 6-octadecenoic acid was identified in the
PAMPB is nearly 38 % lower than in the control sample, but mono­ PAMPB rather than control and confirmed the pineapple flavor in the
saccharides like glucose and galactose were three times higher than in probiotic beverage of PAMPB, these results were correlated with the
the control samples. Results clearly show that in PAMPB, sucrose was aroma profile obtained by E-nose. Other than this, the esters like acetate
consumed by LGG and converted to glucose and galactose. Yahyaoui and hydroxyl ethyl esters were identified in both samples, but the
et al. (2017) identified a similar result in the probiotic fermented cereal pineapple core added sample has a higher ester area percentage than the
breakfast. In the PCP-added samples, the fructose was entirely consumed control sample, it may be due to the formation of esters by fermentation
by the LGG strain. This may be due to the PCP containing fermentable or the accelerated production of ester compounds and organic acids
sugar in the 51.05 mg/mL range, stimulating the breakdown of sucrose while adding PCP. During fermentation, more amounts of organic acids
molecules during fermentation (Hlalukana et al., 2021). The nature like glucuronic acid, formic acid, acetic acid, and citric acid were pro­
carbon source of PCP to microbes (LGG) makes PCP a useful prebiotic duced. These organic acids mainly affect the probiotic beverage’s sen­
source. sory quality and functional activity. Acetic acid is the predominant
organic acid found in fermented beverages; the percentage of acetic acid
varied significantly in the control and PAMPB. Similar kind of results
GC–MS analysis in PAMPB was obtained in most of the probiotic beverages obtained by
L. rhamnosus (Alemneh et al., 2021). Phytosterols like γ-sitosterols
Phytochemical screening on control and PAMPB was done using which have potential anticancer and antioxidant activity were identified
GC–MS to analyze the change in constituents of bioactive and chemical in both control and PAMPB samples, which can help to reduce the
compounds of millet probiotic beverage by the addition of PCP. Based on cholesterol levels in the body (Babu & Jayaraman, 2020).
the results, the presence of several polysaccharides, acids, alkaloids,
amines, and esters was identified and listed in Table 3. The reducing
sugar sucrose in control is not identified in the PAMPB; it denotes the Aroma profile analysis in PAMPB
complete utilization of sucrose by lactobacillus species and improved
fermentation rate. Trisaccharides like melezitose, a rare glucoside pre­ The aroma profile of the samples was analyzed using an E-nose; more
sent in honey, are identified in the PAMPB samples, which shows the than 250 aroma compounds were identified in millet probiotic beverage
structural changes, breakdown, and consumption of sugar molecules. (Table 4). Most of the native volatile compounds of finger millets are
Lactobacillus species does not consume melezitose for their metabolic unidentified, but new aroma compounds, especially many new esters
activity, so it remains in the millet beverage and provides sweetness. and aldehydes were identified in a fermented probiotic beverage. Pro­
Also, it contains potential nutraceutical and prebiotic properties (Abdi biotic beverages from WFM usually produce unfavorable aroma and
et al., 2006; Garcia-gonzalez et al., 2019). Other than this, poly­ flavor, so they are usually aromatized using dried herbs, and spices.
saccharides like lactose and disaccharide mannose were found in control Wang et al. (2014) identified hexanal, 2-pentylfuran, and 3-octen2-one
samples, but PAMPB consists of the hexose monosaccharide tagatose. are the compounds responsible for the dominant millet odor; these
The different structure of reducing sugar helps improve the food sys­ compounds are no longer identified in fermented beverages. Instead,
tem’s antioxidant property, polysaccharides present in PCP exhibit high they structurally modify and produce several esters, aldehydes, ketones,
antioxidant activity and interesting water-binding capacity (Hadidi lactones, and furan compounds. Wei et al. (2011) analyzed the volatiles
et al., 2020). A β-d-glucopyranose manner indicates the breakdown of in pineapple core and pomace, noticed the presence of 44 volatile
galactomannan by enzymes secreted by the lactobacillus enzymes; the compounds, in that 18 were esters, 17 were terpenes, and 4 were al­
newly converted manner act as a new carbon source for the lactobacillus kenes; compare to other waste parts core contains a huge number of
during fermentation which will confirm the prebiotic effect of PCP. A terpenes. The pineapple core aroma was exhibited by ethyl hexanoate to
similar kind of prebiotic effect of pineapple peel powder was observed a greater degree compared to other compounds like decanal, ethyl
by Srinivasreddy et al. (2022), where the enzymes produced D man­ 3-(methylthio)propanoate, and nonanal. The ester compounds respon­
nopyranose from galactomannan. sible for the characteristic aroma in pineapple core like 2-methyl buta­
A specific aroma compound responsible for pineapple flavor noate, ethyl hexanoate, and 2,5-dimethyl-4‑hydroxy-3(2H)-furanone

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Table 4
Qualitative analysis of aroma compounds in PAMPB.
Group RI Compound name CAS Relative peak Sensory
area (%) description

Control PAMPB
Aldehydes 82.09 p-Anisaldehyde 123–11–5 0.135 0.079 Anise; Floral; Hawthorn; Minty; Powdery; Sweet
89.93 Vanillin 121–33–5 – 0.094 Aromatic; Chocolate; Creamy; Pleasant; Sweet; Vanilla
Ketones 17.12 Propan-2-one 67–64–1 65.15 80.36 Fruity, Violet, Apple
21.90 Butane-2,3-dione 431–03–8 – 11.36 Butter; Caramelized; Chlorine; Creamy; Fruity; Pineapple;
Pungent; Spirit; Strong; Sweet
38.47 Pentan-2-one 107–87–9 – 0.063 Acetone; Banana; Etheral; Fruity; Fruity(Sweet); Sweet
42.27 1-Hydroxy-2-propanone 116–09–6 0.011 0.038 Caramelized; Pungent; Sweet
65.03 3-Octanone 106–68–3 0.374 1.26 Butter; Fresh; Fruity; Herbaceous; Mild; Mushroom; Resinous;
Sweet
89.37 2-Tridecanone 593–08–8 – 0.212 Coconut; Dairy; Earthy; Fatty; Fruity; Green; Herbaceous; Milky;
Nutty
99.37 Benzophenone 119–61–9 0.098 0.023 Balsamic; Powdery(Faint); Rose
Alkene 61.48 Myrcene 123–35–3 – 0.057 Balsamic; Etheral; Fruity; Lemon; Plastic; Pleasant; Spicy; Sweet
Esters 17.12 Methyl acetate 79–20–9 0.982 0.979 Blackcurrant; Etherl; Fragrant; Fruity; Fruity(Sweet); Pleasant;
Solvent; Sweet
26.14 Methyl propanoate 554–12–1 0.64 0.64 Apple; Etheral; Fresh; Fruity; Harsh; Rum; Strawberry; Sweet
28.97 Ethyl Acetate 141–78–6 – 0.025 Acidic; Butter; Caramelized; Etheral; Fruity; Green; Orange;
Pineapple; Pungent; Solvent; Sweet
29.48 Ethyl butyrate 105–54–4 – 0.095 Acetone; Banana; Bubblegum; Caramelized; Fruity; Pineapple;
Strawberry; Sweet
31.28 Methyl butanoate 623–42–7 0.042 Apple; Banana; Ester; Etheral; Fruity; Green; Pineapple; Sweet
60.42 Ethyl hexanoate 123–66–0 – 0.426 Anise; Apple; Banana; Berry; Fruity (sweet); Pineapple;
Strawberry; Sweaty; Sweet
76.15 Methyl nonanoate 1731–84–6 0.325 0.092 Coconut; Fruity; Fruity (Sweet); Nutty; Pear; Sweet; Tropical
76.22 Phenylethyl acetate 103–45–7 0.146 0.092 Apple; Apricot; Floral; Fruity; Honey; Sweet; Tobacco; Tropical
79.51 5-Propyldihydro-2(3H)- 105–21–5 – 0.255 Fatty; Fruity; Nutty
furanone
81.62 (Z)− 3-Hexenyl hexanoate 31,501–11–8 – 0.079 Fruity; Grassy; Green; Pear; Pineapple; Prune; Tropical; Waxy;
Winey
84.68 Methyl cinnamate 103–26–4 – 0.062 Balsamic; Cherry; Cinnamon; Fruity; Strawberry; Sweet
103.37 Methyl hexadecanoate 112–39–0 – 0.022 Faint; Fatty; Orris; Sweet
Acids 50.11 Propanoic acid 79–09–4 0.363 0.036 Acidic; Pungent; Pungent (Slightly); Rancid; Soy; Vinegar
61.65 3-methylbutanoic acid 503–74–2 – 0.045 Acidic; Cheese; Fruity; Sour; Sweaty; Tropical
84.03 Decanoic acid 334–48–5 – 0.062 Citrus; Creamy; Fatty; Meaty; Peach; Rancid; Soapy; Sour
Phenols 77.81 m-cresol 108–39–4 0.323 0.031 Phenolic; Woody
Alcohol 17.12 Ethanol 64–17–5 0.982 0.98 Alcoholic, Ethanol, Pungent, Strong
34.41 1-Propanol, 2-methyl 78–83–1 – 0.028 Alcoholic; Bitter; Sweet; Unpleasant; Winey
58.85 2,3-Butanediol 513–85–9 – 0.030 Creamy; Fruity; Odorless; Onion
73.03 Phenol 108–95–2 0.104 0.080 Acrid; Aromatic; Medicinal; Phenolic; Sweet
78.51 Geraniol 106–24–1 0.211 0.692 Citrus; Floral; Fruity; Pleasant; Rose; Sweet
84.16 Nerol 106–25–2 0.227 0.043 Citrus; Floral; Marine; Rose; Sweet
Lactone 83.85 4-Octanolide 104–50–7 0.192 0.062 Caramelized; Coconut; Creamy; Dairy; Fatty; Fruity; Pungent;
Roast; Sweet
86.25 Delta-octalactone 698–76–0 0.239 0.031 Coconut; Dairy; Fatty; Peach; Sweet; Tropical
93.53 Delta-decalactone 705–86–2 0.074 0.029 Coconut; Creamy; Dairy; Fresh; Fruity; Oily; Peach; Sweet
100.07 Dodecan-4-olide 2305–05–7 – 0.026 Fruity; Green; Metallic; Peach; Sweet
Others 28.25 Chloroform 67–66–3 0.022 0.048 Pleasant
45.99 Acetoin 513–86–0 0.049 0.069 Butter; Coffee; Creamy; Dairy; Fatty; Milky
56.85 Formamide, N,N- 68–12–2 0.027 0.179 Amine; Faint; Fishy
dimethyl-
67.15 5-Methylfurfural 620–02–0 0.548 0.031 Acidic; Almond; Burnt Sugar; Caramelized; Coffee; Maple; Spicy
67.42 5-dimethyl-3(2H)- 3658–77–3 – 0.0451 Baked; Burnt sugar; Candy; Caramelized; Cotton candy;
furanone Strawberry; Sweet
86.41 Eugenol 97–53–0 – 0.027 Balsamic; Herbaceous; Honey; Spicy; Sweet
88.31 2-Tridecae 593–08–8 – 0.110 Coconut; Dairy; Fatty; Fruity; Green; Milky; Nutty; Spicy; Waxy

were identified in the PAMPB, which helps to mask the millet flavor in identified in the PAMPB. Acetoin was identified in both probiotic bev­
the pineapple core added beverage (Wei et al., 2011). The amino acid erages; it is an organic compound excreted by fermentative bacteria to
conversion explains the production of several new acids and aromatic prevent the over-acidification of the cytoplasm (Bajpai-Dikshit et al.,
compounds which are not available in WFM and PCP during probiotic 2003).
growth time, including acidic acid, butanoic acid, hexatonic acid, and Several aromatic ketones like benzene and 1‑hydroxy-2-propanone
heptatonic acid. Like the findings the fermented probiotic beverage were identified in both fermented beverages, a butane-2,3‑dione will
exhibited several new acidic and aromatic compounds than the pure provide the buttery flavor in fermented beverages, pentan-2-one, and 2-
apple juice due to the amino acid conversion during fermentation tridecanone was present in pineapple core added probiotic beverage. A
(Ellenderson et al., 2012). The L. rhamnosus fermentation leads to the lactone compound like delta-octalactone, and delta-decalactone, which
generation of different alcohol compounds like propane-2-one, ethanol, will be presented in fermented products and provide additional flour
and n-butanol. Within the volatile compounds, propane-2-one occupies was found in both control and PAMPB. In addition to that, the PAMPB
65.15 % and 80.36 % in control and PAMPB, respectively. Some cyclic consists of Dodecan-4-olide and gamma-nonalactone compounds.
ester compounds like 5-Propyldihydro-2(3H)-furanone were found in Alkene in PAMPB like myrcene has an antidiabetic, anti-inflammatory,
both samples. Aromatic compounds like eugenol and geraniol were and anticancer effect (Srinivasreddy et al., 2022). The results show

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B. Malini et al. Food Chemistry Advances 4 (2024) 100593

Fig. 4. Effect of storage time at 4 ◦ C on the (A) pH and titratable acidity (TA), (B) Lactocaseibacillus rhamnosus (LGG) in pineapple core powder added millet probiotic
beverage (PAMPB).

that most of the indigenous aroma compounds of WFM were metabo­ pineapple core ester compound ethyl hexanoate. The shelf life of the
lized to low or undetectable levels, meanwhile, the new aroma com­ probiotic beverage was observed for 21 days under storage conditions. A
pounds especially esters, terpenes, and aldehydes formed; further, the notable decrease in probiotic load and pH of the PAMPB was observed.
addition of PCP enhances the aroma profile by providing the dominant The potential application of millets in probiotic beverages is about to
pineapple core flavor by ethyl hexanoate. Hence, the millet flavor was explore in future markets. The results provide scientific evidence for the
replaced by the new fruity flavor which enhanced the pleasant smell due PCP added millet probiotic beverages as a better probiotic product in a
to the presence of aldehydes, including nonanal and undecanal. food market. In conclusion, the utilization of PCP in millet probiotic
beverage has potential health benefits and increases the beverage’s
overall quality. Further studies to stabilize the viability of probiotic
Viability of LGG in PAMPB during conservation under storage condition and the effect of other pineapple co-product like
pomace and crown in millet beverages further helps to compare the
The PAMPB was prepared under the optimized condition and the suitability at the commercial level and analyze the advantages over one
control was stored at 4⁰C for 21 days to study the viability of LGG under other.
storage conditions. Every 3 days interval pH, TA, and LGG viable load
was noted down, and the variation in parameters under storage condi­ CRediT authorship contribution statement
tions was shown in Fig. 4. Fig. 4(A) shows the significant reduction of
LGG from 9.54 to 8.53 log (CFU/mL) in control and from 11.54 to 9.39 B Malini: Methodology, Formal analysis, Investigation, Writing –
log (CFU/mL) in PAMPB at 21 days. Fig. 4(B) shows the reduction in pH original draft. C K Sunil: Conceptualization, Visualization, Methodol­
and increase in TA % during refrigerated storage of probiotic beverages. ogy, Validation, Supervision, Resources, Funding acquisition, Writing –
The pH of the beverage continued to decrease significantly (p <0.05) review & editing. Ashish Rawson: Supervision, Writing – review &
from 4.55 – 4.10 and 4.37 – 3.90 in control and PAMPB, respectively, editing. R Vidyalakshmi: Supervision, Writing – review & editing. N
during the 21 days storage period. The average change in pH per day is Venkatachalapathy: Supervision, Writing – review & editing.
0.06 in both beverages. The TA (%) of the fermented beverages
increased throughout the storage period. The pH and TA of the probiotic
beverage’s changes were significant (p < 0.05) in the storage period. Declaration of Competing Interest
Rozada et al. (2009) reported a similar kind of pH reduction in
malt-based beverages fermented by Bifidobacterium. The reduction in All the authors enlisted declare that there are no conflicts of interest
LGG may be caused by the pH and storage temperature hurdle effect. to disclose.
However, L. rhamnosus has a higher survival rate under higher acidic
storage conditions at 4⁰C; similar kinds of results were stated by Gupta Data availability
et al. (2010) in an L. plantarum related study.
Data will be made available on request.
Conclusion

Acknowledgment
This study provides insight and an overview of changes in probiotic
millet beverages due to the incorporation of PCP. The development of a
The work was funded by the Ministry of Food Processing Industries,
white finger millet probiotic beverage with PCP was optimized suc­
Government of India (Q-11/10/2020-R&D).
cessfully using the D optimal mixture design. The optimized condition,
the addition of 2 % PCP and 2 % LGG inoculum with WFM probiotic
beverage, provides the appropriate probiotic (LGG) load in the limited References
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