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LWT - Food Science and Technology xxx (2018) xxx-xxx

LWT - Food Science and Technology xxx (2018) xxx-xxx Contents lists available at ScienceDirect LWT -

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LWT - Food Science and Technology

journal homepage: www.elsevier.com

Science and Technology journal homepage: www.elsevier.com Influence of acerola pulp concentration on mead production

Influence of acerola pulp concentration on mead production by Saccharomyces

cerevisiae AWRI 796

Thaíse Souza Amorim a , Solimar de Brito Lopes b , Jose Ailton Conceição Bispo ,

b⁠

b⁠

Carlos Francisco Sampaio Bonafe c , Giovani Brandão Mafra de Carvalho , Ernesto Acosta Martínez

b⁠ ,⁠

a Departamento de Ciências Biológicas, Universidade Estadual de Feira de Santana (UEFS), Av. Transnordestina s/n, Bairro Novo Horizonte, 44036-900, Feira de Santana, BA, Brazil

b Departamento de Tecnologia, Universidade Estadual de Feira de Santana (UEFS), Av. Transnordestina s/n, Bairro Novo Horizonte, 44036-900, Feira de Santana, BA, Brazil

c Departamento de Bioquímica e Biologia Tecidual, Instituto de Biologia, Universidade Estadual de Campinas (UNICAMP), Rua Monteiro Lobato, 255, 13083-862, Campinas, SP, Brazil

ARTICLE

INFO

Keywords:

Acerola

Honey

Mead

Saccharomyces cerevisiae

ABSTRACT

In this work the influence of acerola (Malpighiae marginata DC) pulp, at concentrations of 0, 10, 15, 25 and

30%, on mead production by Saccharomyces cerevisiae AWRI 796 was evaluated. A novel approach based on

cell growth to obtain high accuracy fits for the data and to estimate optimization parameters such as Gibbs free

energy was used. Fermentation was done using 10

of increasing concentrations of acerola pulp progressively enhanced the cell growth, with the highest cell con-

cells/mL. Under these conditions, the lowest Gibbs free energy of growth

was −4.81 kJ/mol after fermentation for 288 h at a pulp concentration of 18%. The velocity of substrate con-

centration reached being 2.09 × 10

cells/mL at pH 5.0 and 30 °C for up to 288 h. The addition

7⁠

8⁠

sumption was −14 × 10 3

production of 15.2% v/v or 120.1 g/L.

A.U./h whereas the velocity of ethanol formation was 17 × 10 3 A.U . /h with ethanol

1. Introduction

Mead is perhaps the oldest alcoholic beverage and is produced from

honey. Mead contains many nutritional elements required by the body

and has favorable effects on digestion, metabolism and the treatment

of anemia and chronic diseases of the gastrointestinal tract (Gupta &

Sharma, 2009). Honey, a natural food produced by honey bees from

flower nectar or plant secretions, contains a complex mixture of carbo-

hydrates (mainly glucose and fructose), organic acids, lactones, amino

acids, minerals, vitamins, enzymes, pollen, wax and pigments (Lemos,

Santos, & Santos, 2010). This composition accounts for the widespread

use of honey to sweeten food and improve the palatability of medicines.

Honey is generally fermented using the yeast Saccharomyces cere-

visiae, the most important microorganism in alcohol fermentation be-

cause of its high capacity for fermentation, its high tolerance to ethanol

and other inhibitors formed during the pre-treatment of raw materi-

als and during fermentation, and its capacity for rapid growth under

Corresponding author. Email address: ernesto.amartinez@yahoo.com.br (E.A. Martínez)

https://doi.org/10.1016/j.lwt.2018.07.009

Received 5 October 2017; Received in revised form 21 March 2018; Accepted 7 July 2018 Available online xxx 0023-6438/ © 2018.

anaerobic conditions (Knauf & Kraus, 2006; Lima, Basso, & Amorim,

2001).

Mead production is still largely an empirical and homemade process.

In the last 15 years, there have been attempts to improve the process

of mead production, such as the use of a better fermentation agent

(Fernandes, Locatelli, & Sacartazzini, 2009; Pereira, Mendes-Ferreira,

Oliveira, Estevinho, & Mendes-Faia, 2013, 2009), the use of a wort

formulation with salts (Fabian, 1935; Ferraz, 2015; Pereira,

Mendes-Ferreira, Estevinho, & Mendes-Faia, 2015; Steinkraus & Morse,

1966), and the use of mixtures of honey with fruits and vegetables such

as banana, berry, soya, grape, black rice, apple, coconut milk (Balogu

& Towobola, 2017; Ferraz, 2015; Gupta & Sharma, 2009; Koguchi,

Saigusa, & Teramoto, 2009), organic acids (Mendes-Ferreira et al., 2010;

Sroka & Tuszyński, 2007) and pollen (Roldán, van Muiswinkel, Lasanta,

Palacios, & Caro, 2011). Some investigations have also sought to im-

prove the quality of honey used (Kempka & Mantovani, 2013; Koguchi

et al., 2009), in addition to attempts to improve the fermentation

process (Czabaj, Kawa-Rygielska, Kucharska, & Kliks, 2017; Gomes,

2010; Gomes et al., 2013; Iglesias et al., 2014; Morse & Steinkraus,

1971; Navrátil, Šturdík, & Gemeiner, 2001; Pereira,

T.S. Amorim et al.

Mendes-Ferreira, Oliveira, Estevinho, & Mendes-Faia, 2014) as well as the quality (Ukpabi, 2006) and sensory parameters (Gomes et al., 2015; Rivaldi, Silva, Coelho, Tomazi, & Maciel, 2009; Roldán et al., 2011; Vidrih & Hribar, 2007) of the final product.

Honey-based drinks have little body and are very sweet, so fruits in

the form of juices, pulps or pieces can be added to contribute to acidity,

and growth factors can be used to enhance the fermentation and senso-

rial characteristics (Gupta & Sharma, 2009; Koguchi et al., 2009). Fruits

and their pulps have been highly recommended because of their rich-

ness in carbohydrates, fibers, minerals, vitamin C, carotenoids, pheno-

lic substances and sulfuric substances, and also because of their antiox-

idant action that helps to maintain a balance between production and

elimination of reactive oxygen species and other related compounds,

thereby attenuating free radical-induced damage to cells (Maia, Sousa,

& Lima, 2007). Acerola pulp, which is very appreciated for its flavor

and color (Johnson, 2003; Pino & Marbot, 2001), has a high level of

ascorbic acid (10004500 mg/100 g of fruit) and a total phenolic con-

tent of 835 ± 32 mg and 449 ± 10 mg/100 g for aqueous and hydroalco-

holic extracts, respectively. These levels of ascorbic acid and phenolic

compounds indicate a high antioxidant capacity (Vieira, Sousa, & Lima,

2011). In this work, we examined the influence of acerola pulp concen-

tration on mead production by S. cerevisiae AWRI 796.

2. Materials and methods

2.1. Raw material

Wild flower honey was acquired through the Beekeeping Cooper-

ative of Ribeira do Pombal (COOARP) located in the city of Ribeira

do Pombal, BA, Brazil. Commercial acerola pulp [4.0 ± 0.2% reducing

sugars, 96.0 ± 0.2% non-reducing sugars, 685.00 ± 49.81 mg vitamin C/

Brix, pH 3.23 and 0.61 ± 0.09% proteins, assayed as

previously described by Bastos, Martinez, and Souza (2016)] was pur-

chased in markets at Feira de Santana city, BA, Brazil.

100 g, 8.69 ± 0.45

o⁠

2.2. Yeast propagation

The experiments were done with S. cerevisiae strain AWRI 796, a

commercial yeast from Mauri Yeast (Burns Philp, Sydney, Australia)

that was obtained as freeze-dried cultures from the Australian Wine Re-

search Institute culture collection (Adelaide, Australia). A 30 °Brix blend

containing distilled water and honey was prepared and autoclaved at

120 °C for 15 min in a 125 mL Erlenmeyer flask containing 50 mL of

medium. For all experiments, a starter culture was prepared by adding

0.2 g of lyophilized yeast (S. cerevisiae AWRI 796) to a shaker (Tecnal

TE-420) followed by incubation at 30 °C and 150 rpm for 2448 h. The

initial appropriate amount of inoculum (at least 10

termined by counting in a Neubauer chamber (Cadena-Herrera et al.,

2015).

cells/mL) was de-

8⁠

2.3. Preparation and treatment of must

All fermentations were done in triplicate using a system that con-

sisted of 500 mL flasks containing 250 mL of wort mixture and fitted

with an airlock valve used to release CO

tion. This CO 2 displaced the air and served as a blanket, allowing the

yeast to grow under anaerobic conditions. Water supplemented with

yeast extract (5 g/L), malt extract (5 g/L), peptone (10 g/L), magne-

sium chloride (0.05 g/L), ammonium sulfate (0.3 g/L) and dibasic am- monium phosphate (0.05 g/L) was autoclaved at 110 °C for 10 min. Fur- ther addition of honey resulted in final sugar concentrations of 50.9 g/L

produced during fermenta-

2⁠

2

LWT - Food Science and Technology xxx (2018) xxx-xxx

for sucrose, 99.9 g/L for glucose and 118.9 g/L for fructose. Different quantities of acerola pulp were then added to obtain final pulp concen- trations of 10, 15, 20, 25 and 30%. Each wort sample was adjusted to 30 °Brix by adding honey and the pH of the wort was adjusted to pH

5.0 using calcium carbonate or lactic acid. A yeast inoculum (10 8 cells/

mL) was added to each assay to yield an initial cell concentration of

10 7 cells/mL in must. During alcohol fermentation, the flasks were

incubated at 30 °C in a biochemical oxygen demand (BOD) incubator

(model Q315M25, Quimis greenhouse incubator, Diadema, SP, Brazil)

without shaking. The total length of fermentation was 288 h.

2.4. Monitoring of fermentation

At the end of each 24-h period, samples were obtained aseptically to

assess the fermentation and growth parameters. Cells were counted with

a syringe-type system and cell viability was assessed using methylene

blue in a Neubauer chamber. The concentration of soluble solids ( o Brix)

was determined with a digital portable refractometer (Reichert Tecnal

AR-200, Piracicaba, SP, Brazil). The alcohol content (% v/v) was mea-

sured with a DDM 2911 bench densitometer from Rudolph Analytical

Research (De Melo, Araújo, Bispo, Del Bianchi, & De Carvalho, 2017).

2.5. Thermodynamic analysis

The results were analyzed thermodynamically essentially as de-

scribed elsewhere (Bispo, Bonafe, Santana, & Santos, 2015, 2012). Some

modifications were made to the previous procedure to allow the in-

clusion of more than two species states and the occurrence of reaction

rates involving distinct stoichiometric coefficients between reagents and

products. In this case, the experimental results and 3D surface plots

were fitted using Eq. (2) of Bispo et al. (2015) described below. Eq. (2)

was applied to n equal to the number of experimental points for yeast

cell growth (y) and time (x).

points for yeast cell growth ( y ) and time ( x ). (1) Considering the

(1)

Considering the process of yeast duplication during growth, the equi-

librium constant should be expressed as:

(2)

where L is the initial concentration of yeast species prior to division

corresponds to the yeast generated by division. Note that

the superscript (*) is used here only to differentiate the starting yeast

species from those arising from duplication. In addition, Eq. (2) gives

only a simplified reaction equation without the presence of other reac-

tants, e.g., substrate, and reaction products such as ethanol and carbonic

gas. However, the process described below allows independent charac-

terization of these other compounds.

Thus, considering the process described by relationship (2) and the

extent of process parameter (x L ) (Bispo et al., 2015), the concentration

) of each species at time zero (t 0 ) and any time (t) can be

(c L ) and (c L*

described as:

while L

be (c L ⁠ ) and (c L * ⁠ described as: while L * ⁠

*

(3)

(3)

(4)

(4)

whereas at time (t), the relationship would be

(5)

(5)

T.S. Amorim et al.

where corresponds to the initial yeast concentration.
where
corresponds to the initial yeast concentration.

(6)

As previously described [eq. (1) in Bispo et al. (2015)], from Eqs. (4)

and (5) the Gibbs free energy of yeast growth/duplication related to eq.

(1) can be defined as:

growth/duplication related to eq. (1) can be defined as: (7) where R is the gas constant

(7)

where R is the gas constant and T is the absolute temperature (K).

Thus, the degree of growth for an initial yeast cell concentration (c L 0

) is given by:

initial yeast cell concentration ( c L 0 ) is given by: (8) Introducing Eq. (8)

(8)

Introducing Eq. (8) into Eq. (7) yields the following relationship that

directly correlates the degree of growth (G ) with the Gibbs free energy

of growth/duplication:

G ) with the Gibbs free energy of growth/duplication: (9) This equation shows that the energy

(9)

This equation shows that the energy change of reaction is obtained

from the degree of growth, the initial yeast concentration and the spec-

ified stoichiometry of reaction. Analogously, considering now the con-

version of substrate (glucose/fructose) into ethanol, the same procedure

should be applied. Thus,

(10)

which yields

same procedure should be applied. Thus, (10) which yields where the subscript R in and ΔG
where the subscript R in and ΔG R⁠
where the subscript R in
and
ΔG R⁠

α R

(11)

are related, respectively, to the

Gibbs free energy and degree of reaction during ethanol formation, and

s 0 is the initial substrate concentration.

In addition, the differentiation of the degree of reaction with respect

to time, which can be determined based on cell growth, substrate deple-

tion and ethanol formation, yields

(12) Eq. (12) provides ν , and , additional important parameters ob- G⁠ ν S⁠
(12)
Eq. (12) provides ν
,
and
, additional important parameters ob-
G⁠
ν S⁠
ν P⁠

tained from the experimental data that are directly related to the veloc-

ities of growth, substrate depletion and product formation, respectively.

3. Results and discussion

Fig. 1a shows the experimental results (symbols) and curve fits

(lines) obtained with eq. (1) and Fig. 1b shows the respective percent- age residuals determined from the fitted curves. As indicated in these

LWT - Food Science and Technology xxx (2018) xxx-xxx

3

figures, there was good agreement between the theoretical curves and the experimental data, with random residual distribution and correla-

tion coefficients (R 2 ) >0.9 (data not shown). The use of a surface plot (Fig. 1c) allowed better visualization of the relationship between cell

growth and the pulp concentration and duration of fermentation. The

influence of these parameters can be evaluated by analyzing the contour

plots in relation to the cell concentration axis. Fig. 1d shows four regions

of high cell growth (2 × 10

pulp concentration of 17%, after 168 h of fermentation at a pulp concen-

tration of 24%, after 216 h of fermentation at a pulp concentration of

10% and after 288 h of fermentation at a pulp concentration of 18%.

According to Pereira et al. (2013), the maximum cell biomass and the

maximum number of CFUs (S. cerevisiae Lalvin QA23 and ICV D47) were

obtained at a pitching rate (yeast net growth) of 10

al. (2015), subsequently examined effect of supplementing honey-must

with minerals and/or vitamins on the growth of yeast strains QA23 and

ICV D47. The growth behavior of strain QA23 was similar in all fermen-

tations and the population almost reached 10

pendently of the honey-must composition, the stationary phase of strain

ICV D47 started after 48 h of fermentation and after the population had

CFUs/mL; the number of CFUs/mL was slightly lower

in fermentations with vitamins and salts + vitamins. For both strains

and in all fermentations the population remained constant between 48 h

and 168 h, and then decreased slightly after 288 h (Pereira et al., 2015).

Gomes et al. (2015) observed that in sweet and dry mead fermenta-

tions there was a lag phase of 10 h followed by an exponential phase

of 45 h duration.

Fig. 1e shows the cell concentration as a function of pulp concen-

tration (0, 5, 10, 15, 20, 25 and 30%) at distinct times of fermenta-

tion. The cell growth seen here agreed with the steady progressive mi-

crobial growth (from 1.0 × 10 6 cells/mL to 1.6 × 10 8 cells/mL) observed

during 60 days of fermentation of a honey-coconut milk blend using S.

cerevisiae (Balogu & Towobola, 2017). Cell growth from 10 5 cells/mL to

cells/mL was also seen during honey fermentation supple-

mented with nitrogen and organic acids (Mendes-Ferreira et al., 2010).

According to Pereira, Dias, Andrade, Ramalhosa, and Estevinho (2009),

all yeast strains showed the same behavior at 10% (v/v) ethanol, al-

though there was a decrease in cell viability and none of the strains

grew at ethanol concentrations of 1520% (v/v).

Fig. 1f shows the degree of growth when the data were normalized

from 0 to 1. From these results and Eq. (9) it was possible to calcu-

late the Gibbs free energy of growth/duplication (Fig. 1g). Cell growth

was spontaneous under all conditions studied, with values ranging from

3.2 to 4.8 kJ/mol. Comparison of Fig. 1d,h indicated that the fermen-

tation time and pulp concentration that yielded the highest cell growth

cells/mL) were those that yielded the lowest Gibbs free en-

ergy of growth (−4.81 kJ/mol). For 1.84 × 10 8 cells/mL, the process was

also spontaneous, with a free Gibbs energy of growth of −4.51 kJ/mol.

The surface profile for the velocity of growth during fermentation

can be obtained from Eq. (12) and is shown in Fig. 2a. Fig. 2b and c

shows that for 1.84 × 10 8 cells/mL a higher growth rate, corresponding

to 0.0315 A.U./h, was obtained after 48 h of fermentation at a pulp

concentration of 17%. These figures also indicate regions of negative

growth that reflect greater cell death relative to cell division.

Application of the analytical approach described above to the eval-

uation of soluble solids in solution (S.S., °Brix) is shown in Fig. 3a

and b. The optimal conditions should contemplate an economy in the

most expensive component in mead fermentation, namely honey. Fig. 3c,d,e shows that such optimization is reached at 1.84 × 10 8 cell/mL after 168 h of fermentation at a pulp concentration of 24% and at 2.09 × 10 8 cells/mL after 288 h of fermentation at a pulp concentration

(2.09 × 10

1.52.8 × 10 7

reached 78 x 10

CFUs/mL after 48 h. Inde-

CFUs/mL. Pereira et

cells/mL) after 48 h of fermentation at a

8⁠

8⁠

8⁠

7⁠

8⁠

T.S. Amorim et al.

LWT - Food Science and Technology xxx (2018) xxx-xxx

et al. LWT - Food Science and Technology xxx (2018) xxx-xxx Fig. 1. A) Cell concentration

Fig. 1. A) Cell concentration [X] (symbols) as a function of fermentation time (h) and pulp concentration (%). Lines were fitted using the function described in Eq. (1). b) Residual plot

for the data fitted to the lines in panel a. c) Surface plot and d) contour plot of cell concentration (S. cerevisiae AWRI 796) as a function of fermentation time (h) and pulp concentration

(%). e) Cell concentration [X] as a function of pulp concentration (%) in each fermentation time (h). f) Surface plot of the degree of growth as a function of fermentation time (h) and pulp

concentration (%). g) Surface plot and h) contour plot of the Gibbs free energy of growth (

and h) contour plot of the Gibbs free energy of growth ( Joule/mol) as a function

Joule/mol) as a function of fermentation time (h) and pulp concentration (%).

of 18%. These optimal conditions represent an economy in the use of

honey (Fig. 3d and e). Fig. 3fh corroborates these results since the °Brix

decreased over time and the negative velocity of reaction reached at

40 h indicated high substrate consumption and ethanol formation (Fig.

3h).

Fig. 4 shows a similar analysis for substrate consumption and ethanol production. Fig. 4c,f shows a velocity of substrate consumption of −1.4 × 10 2 A.U./h, whereas the velocity of ethanol formation was

1.7 × 10

glucose => 2 ethanol), these data suggest that 39% of the substrate

was used to produce other cell compounds.

Substrate consumption was 49.1% and ethanol production was 14.7% (v/v; 116.2 g/L) in the absence of acerola pulp (Figs. 3a and

5a-d). The addition of 1030% acerola pulp enhanced substrate con- sumption (5558%) and increased ethanol production (15.216.6%,

or

v/v,

A.U./h. Based on the stoichiometry of the reaction as 1:2 (1

−⁠

2

4

T.S. Amorim et al.

LWT - Food Science and Technology xxx (2018) xxx-xxx

et al. LWT - Food Science and Technology xxx (2018) xxx-xxx Fig. 2. A) Surface plot

Fig. 2. A) Surface plot of the velocity of growth (v

surface plot of the velocity of growth (V

G , expressed in absorbance units (A.U.)/h) as a function of fermentation time (h) and pulp concentration (%). b) Contour plot and c)

G⁠

) as a function of fermentation time (h) and pulp concentration (%).

120127 g/L) after 288 h of fermentation (Figs. 3a and 5a-d). Similar

values for substrate consumption (58.5%, 54.5% and 42.9%) and lower

ethanol production (9 g/L, 8 g/L and 6.5 g/L) were obtained during the

fermentation of mead (16

respectively, using S. cerevisiae at pH 5.45 and 36 °C for 168 h (Kempka

& Mantovani, 2013). Multiflorous and honeydew diluted in water (36 o Brix) incubated with Saccharomyces bayanus and S. cerevisiae resulted in a substrate con- sumption of 49.759.6% and ethanol content of 11.9816.53% (v/v) at the end of mead production (720 h) (Czabaj et al., 2017). In con

Brix) with honeydew, wild and angico honey,

o⁠

trast, higher substrate consumption (88.5%) and greater ethanol pro-

duction (8%, v/v) were reported by Ilha et al. (2008) for 21 o Brix af-

ter 84 h at pH 4.5 and 25 °C. Gomes et al. (2015) also observed high

rates of sugar consumption (80%) and low ethanol production (7.54

and 13.54%) with S. cerevisiae (pH 3.5, 35 °C) after sweet and dry

mead fermentations for 79 h and 198 h, respectively. Similar high sub-

strate consumption (82%) and ethanol production of 10.0310.33% (v/ v) and 9.710.37% (v/v) were obtained in fermentations with S. cere- visiae Lalvin QA23 and ICV D47, respectively (Pereira et al., 2013).

5

T.S. Amorim et al.

LWT - Food Science and Technology xxx (2018) xxx-xxx

et al. LWT - Food Science and Technology xxx (2018) xxx-xxx Fig. 3. A) Soluble solid

Fig. 3. A) Soluble solid (S.S.; symbols) content of mead as a function of fermentation time (h) and pulp concentration (%). Lines were fitted using the function described in Eq. (1). b)

Residual plot for the data fitted to the lines in panel a. c) Surface plot and d) contour plot of soluble solids (S.S.) as a function of fermentation time (h) and pulp concentration (%). e)

Soluble solid (S.S.; symbols) content as a function of pulp concentration (%) in each fermentation time (h). f) Surface plot of the degree of reaction as a function of fermentation time (h) and pulp concentration (%). g) Surface plot and h) contour plot of the Gibbs free energy of reaction (ΔG R , Joule/mol) as a function of fermentation time (h) and pulp concentration (%).

6

T.S. Amorim et al.

LWT - Food Science and Technology xxx (2018) xxx-xxx

et al. LWT - Food Science and Technology xxx (2018) xxx-xxx Fig. 4. A) Surface plot

Fig. 4. A) Surface plot of the velocity of substrate consumption (V

concentration (%) as a function of velocity of substrate consumption (V

tion time (h) and pulp concentration (%). e) Contour plot and f) surface plot of pulp concentration (%) as a function of the velocity of ethanol production (V P ) and fermentation time (h).

Brix⁠

) as a function of fermentation time (h) and pulp concentration (%). b) Contour plot and c) surface plot of the pulp

Brix⁠

) and fermentation time (h). d) Surface plot of the velocity of ethanol production (V P ) as a function of fermenta-

production (V P ⁠ ) as a function of fermenta- Fig. 5. A) Ethanol production [P]

Fig. 5. A) Ethanol production [P] as a function of fermentation time (h) and pulp concentration (%). Lines were fitted using the function described in Eq. (1). b) Residual plot for the data

fitted to the lines in panel a. c) Surface plot and d) contour plot of ethanol production as a function of fermentation time (h) and pulp concentration (%).

Lower ethanol production (10.711.4%) was reported by

Mendes-Ferreira et al. (2010) for honey fermentation after nitrogen supplementation and the addition of organic acids under the follow- ing initial conditions: 22 o Brix, pH 3.504.72, 10 5 cells/mL of S. cere-

visiae

UCD522

and 600 h of fermentation. A mead with an alcohol content of 1017.2%

and a substrate consumption of 60% after nine days was obtained from honey using commercial S. cerevisiae (20 g/L) at an initial pH

of 3.55. Ethanol production of 12.715.0% was reported by Ukpabi

7

T.S. Amorim et al.

(2006) for mead obtained with cassava (Manihot esculenta) floral honey under farm conditions. Vidrih and Hribar (2007), who studied the fermentation of chestnut, lime and honeydew meads, and Gupta and Sharma (2009), who studied home-brewed and commercial meads

made from soya honey, reported ethanol production of 14.2% and

4.611.8 (v/v), respectively.

As shown here, S. cerevisiae AWRI 976 grew at ethanol concentra-

tions up to 16.6% (v/v). This finding suggests that acerola pulp may in-

crease the tolerance of yeast to ethanol.

4. Conclusion

The addition of acerola pulp during mead fermentation enhanced

cell growth (greater cell concentration) and increased substrate con-

sumption and ethanol production. This is the first study to examine the

influence of fruit pulp concentration on mead production and indicates

that acerola pulp could be a useful substitute for honey, thereby reduc-

ing the cost of mead production. A complementary sensorial analysis of

mead produced with acerola pulp is required to demonstrate its suitabil-

ity as a substitute for honey.

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgments

The authors thank the Beekeeping Cooperative of Ribeira do Pombal

(COOARP), Ribeira do Pombal, BA, Brazil for providing the honey and

Dr. Stephen Hyslop for editing the English of the manuscript. This work

was supported by Conselho Nacional de Desenvolvimento Científico e

Tecnológico (CNPq) and Fundação de Amparo à Pesquisa do Estado de

São Paulo (FAPESP), Brazil.

References

Balogu, T.V., Towobola, O., 2017. Production and quality analysis of wine from honey

and coconut milk blend using Saccharomyces cerevisiae. Fermentation 3, 1–9 http//

dx.doi.org/10.3390/fermentation3020016.

Bastos, J.C., Martinez, E.A., Souza, S.M.A., 2016. Physicochemical characteristics of com-

mercial umbu pulp (Spondias tuberosa Arruda Câmara): Concentration effect. J.

Bioen. Food Sci. 3 (1), 11–16 http//dx.doi.org/10.18067/jbfs.v3i1.48.

1 Bispo, J.A.C., Bonafe, C.F.S., Joekes, I., Martinez, E.A., Carvalho, G.B.M., Norberto,

D.R., 2012. Entropy and volume change of dissociation in tobacco mosaic virus probed

by high pressure. Journal of Physical Chemistry B 116, 14817–14828 http//dx.doi.

1

org/10.1021/jp310219k.

The results were analyzed thermodynamically essentially as described in these pa-

pers. In the thermodynamic analysis some modifications were made to the previous

procedure to allow the inclusion of more than two species states and the occurrence

of reaction rates involving distinct stoichiometric coefficients between reagents and

products.

1 Bispo, J.A.C., Bonafe, C.F.S., Santana, K.M.O.V., Santos, E.C.A., 2015. A comparison

of drying kinetics based on the degree of hydration and moisture ratio. Lebensmit-

tel-Wissenschaft und -Technologie- Food Science and Technology 60, 192–198 http//

dx.doi.org/10.1016/j.lwt.2014.07.014.

Cadena-Herrera, D., Esparza-De Lara, J.E., Ramírez-Ibañez, N.D., López-Morales, C.A.,

Pérez, N.O., Flores-Ortiz, L.F., et al., 2015. Validation of three viable-cell counting

methods: Manual, semi-automated, and automated. Biotechnol. Rep 7, 9–16 https://

doi.org/10.1016/j.btre.2015.04.004.

Czabaj, S., Kawa-Rygielska, J., Kucharska, A.Z., Kliks, J., 2017. Effects of mead wort heat

treatment on the mead fermentation process and antioxidant activity. Molecules 22,

803 http//dx.doi:10.3390/molecules22050803.

De Melo, V.F., Araújo, G.S., Bispo, J.A.C., Del Bianchi, V.L., De Carvalho, G.B.M., 2017. Ef-

fect of different concentrations of bush passion fruit pulp and temperature in the pro-

duction of beer. African Journal of Biotechnology 16, 1150–1158 http//dx.doi.org/

10.5897/AJB2016.15613.

Fabian, F.W., 1935. The use of honey in making fermented drinks. Fruit. Prod. J. Am. Food 14, 363–366. Fernandes, D., Locatelli, G.O., Sacartazzini, L.S., 2009. Avaliação de diferentes estirpes da

levedura Saccharomyces cerevisiae na produção de hidromel, utilizando méis residu- ais do processo de extração. Evidência, Joaçaba 9, 29–42. 2 Ferraz, F.O., 2015. Estudos dos parâmetros fermentativos, características físico-quími- cas e sensoriais do hidromel. PhD thesis Universidade de São Paulo, Lorena, SP, Brazil.

8

LWT - Food Science and Technology xxx (2018) xxx-xxx

2 This work is important for the present study because the author used apple pieces as

a supplement in mead production and this provided the idea of using typical fruit

pulp from the northeast of Brazil in mead production. Gomes, T.M.C., 2010. Produção de hidromel: Efeito das condições de fermentação. MSc

dissertation. Instituto Politécnico de Bragança, Bragança, Portugal.

Gomes, T., Barradas, C., Dias, T., Verdial, J., Morais, J.S., Ramalhosa, E., et al., 2013. Opti-

mization of mead production using response surface methodology. Food and Chemical

Toxicology 59, 680–686 http//dx.doi.org/10.1016/j.fct.2013.06.034.

Gomes, T., Dias, T., Cadavez, V., Verdial, J., Morais, J.S., Ramalhosa, E., et al., 2015. In-

fluence of sweetness and ethanol content on mead acceptability. Pol. J. Food and Nu-

trition Sciences 65, 137–142 http//dx.doi.org/10.1515/pjfns-2015-0006.

3 Gupta, J.K., Sharma, R., 2009. Production technology and quality characteristics of

3

mead and fruit-honey wines: A review. Natural Product Radiance 8, 345–355.

This review is important because of it relates the technology of mead production to

the raw materials used, microbiological aspects and the preparation of mead.

Iglesias, A., Pascoal, A., Choupina, A.B., Carvalho, C.A., Feás, X., Estevinho, L.M., 2014.

Developments in the fermentation process and quality improvement strategies for

mead production. Molecules 19, 12577–12590 http//dx.doi.org/10.3390/

molecules190812577.

Johnson, P.D., 2003. Acerola (Malpighia glabra L., M. punicifolia L., M. emarginata D.C.):

Agriculture, production and nutrition. World Review of Nutrition & Dietetics 91,

67–75.

Kempka, A.P., Mantovani, G.Z., 2013. Produção de hidromel utilizando méis de diferentes

qualidades. Rev. Bras. Prod. Agroindustriais 15, 273–281 http//dx.doi.org/10.15871/

1517-8595/rbpa.v15n3p273-281.

Knauf, M., Kraus, K., 2006. Specific yeasts developed for modern ethanol production.

Sugar Ind 131, 753–758.

Koguchi, M., Saigusa, N., Teramoto, Y., 2009. Production and antioxidative activity of

mead made from honey and black rice (Oryza sativa var. Indica cv. Shiun). Journal of

the Institute of Brewing 115, 238–242 http//dx.doi.org/10.5897/AJB10.1516.

Lemos, G.S., Santos, J.S., Santos, M.L.P., 2010. Validação de método para a determinação

de 5-hidroximetilfurfural em mel por cromatografia líquida e sua influência na quali-

dade do produto. Quimica Nova 33, 1682–1685.

Lima, U.A., Basso, L.C., Amorim, H.V., 2001. Biotecnologia Industrial: Processos Fermen-

tativos e Enzimáticos. Blucher, São Paulo 1–43.

Maia, G.A., Sousa, P.H.M.S., Lima, A.S., 2007. Processamento de Sucos de Frutas Tropi-

cais. UFC, Fortaleza 320.

Mendes-Ferreira, A., Cosme, F., Barbosa, C., Falco, V., Inês, A., Mendes-Faia, A., 2010. Op-

timization of honey-must preparation and alcoholic fermentation by Saccharomyces

cerevisiae for mead production. International Journal of Food Microbiology 144,

193–198 http//dx.doi.org/10.1016/j.ijfoodmicro.2010.09.016.

Morse, R.A., & Steinkraus, K.H. (1971). Method of making wine from honey. US patent.

3,598,607.

Navrátil, M., Šturdík, E., Gemeiner, P., 2001. Batch and continuous mead production with

pectate immobilised, ethanol-tolerant yeast. Biotechnology Letters 23, 977–982 http//

dx.doi.org/10.1023/A:1010571208324.

4 Pereira, A.P., Dias, T., Andrade, J., Ramalhosa, E., Estevinho, L.M., 2009. Mead pro-

duction: Selection and characterization assays of Saccharomyces cerevisiae strains.

Food and Chemical Toxicology 47, 2057–2063 http//dx.doi.org/10.1016/j.fct.2009.

4

05.028.

This work examines the ability of S. cerevisiae strains to grow in different conditions.

The authors emphasize the importance of considering the characteristics of the

honey and supplements used in formulating the fermentation medium to achieve the

best results in mead production.

Pereira, A.P., Mendes-Ferreira, A., Estevinho, L.M., Mendes-Faia, A., 2015. Improvement

of mead fermentation by honey-must supplementation. Journal of the Institute of

Brewing 121, 405–410 http//dx.doi.org/10.1002/jib.239.

Pereira, A.P., Mendes-Ferreira, A., Oliveira, J.M., Estevinho, L.M., Mendes-Faia, A., 2013.

High-cell-density fermentation of Saccharomyces cerevisiae for the optimisation of

mead production. Food Microbiology 33, 114–123 http//dx.doi.org/10.1016/j.fm.

2012.09.006.

Pereira, A.P., Mendes-Ferreira, A., Oliveira, J.M., Estevinho, L.M., Mendes-Faia, A., 2014.

Effect of Saccharomyces cerevisiae cells immobilisation on mead production. Lebens-

mittel-Wissenschaft und -Technologie- Food Science and Technology 56, 21–30 http//

dx.doi.org/10.1016/j.lwt.2013.11.005.

Pino, J.A., Marbot, R., 2001. Volatile flavor constituents of acerola (Malpighia emarginata

DC.) fruit. Journal of Agricultural and Food Chemistry 49, 5880–5882 http//dx.doi.

org/10.1021/jf010270g.

Rivaldi, J.D., Silva, M.M., Coelho, T.C., Tomazi, C., Maciel, I., 2009. Caracterização e perfil

sensorial de hidromel produzido por Saccharomyces cerevisiae IZ 888. Brazilian Jour-

nal of Food Technology 7, 58–63.

Roldán, A., van Muiswinkel, G.C.J., Lasanta, C., Palacios, V., Caro, I., 2011. Influence

of pollen addition on mead elaboration: Physicochemical and sensory characteristics.

Food Chemistry 126, 574–582 http//dx.doi.org/10.1016/j.foodchem.2010.11.045.

Sroka, P., Tuszyński, T., 2007. Changes in organic acid contents during mead wort fermen-

tation. Food Chemistry 104, 1250–1257 http//dx.doi.org/10.1016/j.foodchem.2007.

01.046.

Steinkraus, K.H., Morse, R.A., 1966. Factors influencing the fermentation of honey in mead. Journal of Apicultural Research 5, 17–26. Ukpabi, U.J., 2006. Quality evaluation of meads produced with cassava (Manihot escu- lenta) floral honey under farm conditions in Nigeria. Trop. Subtrop. Agroecosystems

6, 37–41.

Vidrih, R., Hribar, J., 2007. Studies on the sensory properties of mead and the formation of aroma compounds related to the type of honey. Acta Alimentaria 36, 151–162 http//

dx.doi.org/10.1556/AAlim.36.2007.2.2.

Vieira, L.M., Sousa, M.S.B., Lima, A., 2011. Fenólicos totais e capacidade antioxidante

in vitro de resíduos de polpas de frutas tropicais. Rev. Bras. Frutic., Jaboticabal 33, 888–897 http//dx.doi.org/10.4260/BJFT2011140300024.