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Assessment of Date Wine Clarified Using Commercial Pectinase and Aspergillus tamarii Strains Immobilized on Biomatrix

1Emecheta, R. O., *2Maduka, N., 3Ahaotu, I., 4Odu, N. N.,

1Department of Biology/Microbiology, School of Science and Industrial Technology, Abia State Polytechnic, Aba, Nigeria.

2Department of Biological Sciences, Faculty of Natural and Applied Sciences, Wellspring University, Benin City, Edo State, Nigeria 3Department of Microbiology, Faculty of Science, University of Port Harcourt, Choba, Rivers State Nigeria.

4Department of Microbiology, Faculty of Science, Rivers State University, Nkpolu-Oroworukwo Port Harcourt, Rivers State, Nigeria

Corresponding author: maduks.mn@gmail.com

Abstract: This study was aimed at evaluating the use of commercial pectinase and pectinase-producing organisms (Aspergillus tamarii strain DTO: 129–ES and Aspergillus tamari isolate TN – 275) immobilized on a papaya trunk for clarification of date wine. Yeasts (Saccharomyces cerevisiae) and Aspergillus sp. used in this study was isolated from palm wine and yam peel, respectively by means of standard microbiological methods. Physicochemical tests were also performed using Standard methods. Date must obtained from date fruit was fermented by S. cerevisiae for 28 Days at room temperature (28±2 oC) and physicochemical parameters of the fermenting date must was monitored at 24 h intervals followed by clarification of date wine. Results obtained from this study showed that during fermentation of date must, its sugar content and specific gravity which ranged between 18-8 % and 1.055-1.00, respectively steadily reduced. Similarly, there was also reduction in temperature, total dissolved solid, pH, viscosity and reducing sugar of the product which ranged between 29-25.8 oC, 1810-1121mg/l, 6.1-3.9, 179-106 cps and 42.0-1.00 %, respectively. On the contrary, there was steady increase in alcohol content and total titratable acidity of fermenting date must which ranged between 0.5-7.8 % and 0.318-0.606 %, respectively. At wavelength 700-840 nm, average absorbance value of white wine (control), date palm wine clarified using commercial pectinase, immobilized Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES was 0.675, 0.624, 0.2461 and 0.2415 nm, respectively. Key word: Aspergillus tamari, Clarification, Immobilized, Biomatrix, Pectinase, Date wine.

Author(s) agree that this article remain permanently open access under the terms of the Creative Commons

Attribution http://creativecommons.org/licenses/by/4.0/

Vol. 3(1), pp. 019-031, August 2019

ISSN 2315-6287 DOI: 10.14412/SRI2018.031 Copyright© 2018

Author(s) retain the copyright of this article Available online at http://www.scienceparkjournals.com/SPJABR

Science Park Journal of Applied Biotechnology Research

Maduka et al 2.

INTRODUCTION

In the last few decades, increasing cost of imported wines has been a financial burden to Nigerian consumers. To reverse the trend, attention is shifting towards using locally sourced raw materials abundant in supply for wine production. Wine is defined as a fermented beverage produced using only grapes. According to Jagtap et al. [1], abundant and over ripen fruits are suitable for fruit wine production. So far, many wine industries have used different fruits such as cherry, mango, kiwi, banana, apple, orange, cashew nut, jackfruit etc as substrate for fruit wine production. The product name becomes the fruit name prefixed to wine. Since date palm produce surplus over ripen sweet fruit and the tree grows abundantly in Northern Nigeria, it has great potential for industrial use in fruit wine production [2]. Date palm also known as the tree of dates (Phoenix dactylifera L., Family Arecaceae) is a sweet fruit. In the Middle East and North Africa, it has played key role in maintaining food security [3]. Production of date palm started in Nigeria sometime in 17th century. Pilgrims involved in trans-Saharan trade introduced date palm from North Africa to Nigeria. Presently, the crop is cultivated in Northern parts of Nigeria [2]. In Nigeria, date palm is locally incorporated into native beer to reduce intoxication. Bhusari et al. [4] produced fruit wine using Saccharomyces cerevisae and the product had an acceptable colour, aroma, taste and sweetness on a scale of 10. Awe and Nnadoze [2] produced a similar product which recorded lower sensory acceptability of 71.8 % compared with 78.3 % for imported white wine. This could be partly because date fruit wine was cloudy as a result of pectin (polysaccharide found in plant cell walls) that was present in the product. In wine tasting, the product is considered “clear” when there are no visible particles suspended in the liquid especially white wines that have some degree of transparency. Therefore, the process of removing materials such as dead yeast cells, bacteria, tartrates, proteins, pectins, tannins, phenolic compounds, as well as pieces of grape skin, pulp, stem, sand and gums suspended in wine that makes it cloudy before bottling is known as clarification. This process involves filtration, flotation and the use of fining agents or pectinase (pectin-splitting enzymes) [5]. Brown and Ough [6] reported that adding pectinase in grape juice maceration increased clarity and filterability of the juice by 100 %. Studies have demonstrated that depectinisation of fruit wines using pectinases is efficient in reducing its turbidity [7, 8]. Pectinases degrade pectins, resulting in viscosity reduction and cluster formation which facilitates separation through centrifugation or filtration. Since Date wine fermented using wine yeast (Saccharomiyces cerevisiae) is cloudy and less acceptable to consumers, this study seeks to clarify the product as a strategy to promote acceptability thereby increase large scale production in order to reduce importation of white wine. MATERIALS AND METHODS Dried dates were purchased from Ariaria International Market, Aba, Abia State using sterile plastic bags. The fruits were aseptically transferred to Food Microbiology Laboratory, Abia State Polytechnic, Aba for analysis. Yam peel was obtained from a roadside roasted yam seller and fresh palm wine was obtained from a palm wine tapper in Ekeakpara village, Osisioma L.G.A in Abia State, Nigeria. Isolation of yeast from palm wine

Yeast was isolated from fresh palm wine using the method described by Nwachukwu et al. [9] with slight modification. The fresh palm wine was allowed to sediment and ferment naturally for 24 h at

3. S. P. J. App Bio Res room temperature (28±2 oC). One milliliter (1 ml) fermented palm wine was serially diluted (10 fold dilution) up to 10-6 using normal saline. Approximately 0.1 ml from 104 dilution was inoculated onto malt extract agar in triplicate. The plates were incubated at 28±2 oC for 72 h. Morphology of each fungal isolate on the plates were noted. Repeated subculture of the fungal isolates was performed to obtain pure culture. Microscopic observation and biochemical identification using lactophenol cotton blue stain was carried out on each fungal isolate. Isolation of Aspergillus species from yam peel

Slightly modified method described by Kwatia et al. [10] was adopted. One gram (1 g) of composite ground yam peel was homogenized in 9 ml distilled water. One milliliter (1 ml) of the homogenized sample was subjected to ten-fold serial dilution. From 10-3and 10-5 dilution, 0.1 ml solution was inoculated onto solidified potato dextrose agar (PDA) and Sabouraud dextrose agar (SDA) using spread plate method in triplicates. The plates were incubated at room temperature (28±2 oC) for 5 – 7 days. Fungal growth on the agar plates were subcultured on freshly prepared SDA and PDA plates using the same condition. The pure isolates obtained were identified using cotton blue lactophenol staining technique. Morphological characteristics of the isolates were noted and then compared with a chart described by Cheesbrough [11]. Pure cultures were subcultured in PDA slants for further use. Inoculum development

The method described by Okeke et al. [12] with slight modifications was adopted. Two hundred

grams (200 g) of date fruits were sorted, washed and soaked with 500 ml potable water for 45 min.

The soaked sample was macerated, destoned and the must filtered using muslin cloth. Two hundred

milliliter (200 ml) of the juice was sterilized by autoclaving (121 oC for 15 min.; 15 psi) and allowed to

cool. Three (3) loopful of yeast culture isolated from palm wine were inoculated into the sterilized juice

and incubated for 48 h at room temperature (28±2 oC).

Preparation and fermentation of must for date palm wine production

Three kilograms (3 Kg) of Date palm fruits were sorted, washed and soaked with 7.5 litres of potable water for 45 min. The soaked sample was macerated, de-stoned and the must filtered using muslin cloth. Two hundred milliliter (200 ml) of the juice was sterilized by autoclaving (121 oC for 15 min.; 15 psi) and allowed to cool. The inoculum was poured inside the must and the setup was allowed to ferment for 28 Days at room temperature (28±2 oC). Aliquots of the fermenting must was removed every 24 h for physicochemical analysis and sensory evaluation. Physicochemical analysis of date must pH determination Electrochemical method was adopted using pH meter (Hanna H1 98107). Ten milliliter (10 ml) of the sample was poured into 100 ml beaker and pH meter was dipped into the sample in such a way that the electrode was in contact with the sample. The value on the display was noted at 20 oC.

Determination of temperature

Ten milliliter (10 ml) of the sample was poured into 100 ml beaker and mercury-in-glass thermometer was dipped inside the sample. Temperature was determined by reading the rise in the liquid inside the glass thermometer.

Maduka et al 4.

Determination of titrable acidity

Titration method was used to determine the titratable acidity. Ten milliliter (10 ml) of the sample was pipette into 100 ml conical flask. Fifty milliliter (50 ml) of distilled water was added to the sample followed by three drops of phenolphthalein indicator was added to the mixture. The mixture was titrated against 0.1 M NaOH. A slight change of pink colour indicates the end point. Determination of specific gravity

One hundred and fifty milliliter (150 ml) of the sample was poured into 250 ml measuring cylinder. Hydrometer was dipped into the sample causing it to suspend. Then specific gravity was determined by reading the calibration on the hydrometer. Determination of alcoholic content

To determine the % (v/v) alcohol content of the sample, specific gravity, temperature and alcohol correlation was used. One hundred milliliter (100 ml) of the sample was poured into a measuring cylinder and hydrometer was dipped inside the cylinder, causing it to suspend due to alcohol–specific gravity correlation. The calibration was read while avoiding the alcoholmeter from lapping to the wall of the cylinder. Conversion was made using Alcohol-specific gravity-temperature chart. Determination of sugar content

Brix value method by means of refractometer was used to determine the sugar content of the samples following the procedure described by Oti [13]. The refractometer was reset to zero using distilled water. Using a dropper, the sample was placed on the slide of the refractometer and the brix value was viewed from the lens of the refractometer.

Determination of viscosity

The viscosity of date must was determined using a viscometer. Fifty milliliter (50 ml) of sample was poured into the breaker then robot 2 was used at the speed of 30. The correct reading was taken when it was above 10 %. The robot number was selected and the speed of reading was recorded. Determination of total dissolved solid

TDS meter-HannaHI9830 (instrumentation) was used to determine total dissolved solid (TDS) in date must. Fifty milliliter (50 ml) sample was poured into a clean 100 ml beaker. The TDS meter (HI98301) was dipped into the sample, causing the electrode to come in contact with the sample. The TDS value displayed on the equipment was recorded as the result in mg/l.

5. S. P. J. App Bio Res

Determination of reducing sugar

The test for reducing sugar was carried out using Lane and Eynon method. The solution was clarified by adding 5 ml zinc acetate solution followed by 5 ml of potassium ferrocyanide solution and filtered. Fifty milliliter (50 ml) of the filtrate was transferred into 100 ml volumetric flask and made up to the mark. Five milliliter (5 ml) of Fehlings solution A and 5 ml of Fehlings solution B were pipette into the conical flask. From the burette, 15 ml of the filtrate was added to the conical flask containing Fehlings solution and boiled until brick red colour was seen. Four drops of methylene blue indicator was immediately added while boiling and titration continued with constant shaking until brick red colour was observed and the titre value (vml) recorded. The titration was repeated twice vml equal (v-2) ml. From the sugar table, the milligram of sugar (as fructose) in the titrated volume of the sugar solution was noted. After Inversion 40 ml of clarified sugar was transferred into 100 ml volumetric flask followed by addition of 7 ml of concentrated hydrochloric acid and immersed in waterbath set at 60 oC for 12 min. The flask was removed from waterbath and allowed to cool. The mixture was then neutralized using 5 M NaOH using phenolphthalein indicator and then made up to mark using distilled water. Fifty milliliter (50 ml) of neutralized solution was pipette into 100 ml volumetric flask and made up using distilled water. The same titration before inversion was carried out for the inversed solution using 10 ml/Fehlings solution was repeated. Note: 1 ml of Fehling’s solution = 4.95 mg glucose; = 5.25 mg fructose; = 5.09 mg invest sugar; = 7.68 mg lactose; = 6.46 mg lactose; = 4.75 mg sucrose. From the original small increments of about 0.2 ml standard, invert sugar was added to the boiling solution until the blue colour lessened, then drop wise until the final disappearance of blue colour leaving absolute colour of copper 1 oxide. With the sample solution in the burette, titration was repeated using equal volume of Fehlings solution and water but 25 ml of sample solution for the trial run. The final accurate titration (vi) was carried out using a volume of sample that is 1ml less than the volume used in initial titration. Calculation:

% reducing sugar as invert = 𝑉𝑜 𝑥 25 𝑥 𝑓

𝐶 𝑋 𝑉1

Where: F is a correction factor to be applied if large amounts of sucrose are present e.g. analysis of cane or beef sugar products. C is the % m/v of sample.

Clarification of date palm wine using commercial pectinase Commercial pectinase was purchased from Federal Institute of Industrial Research Oshodi (FIIRO) Lagos Nigeria and maintained at 45 oC to 55 oC and optimum pH of 4 - 5. The freshly prepared date palm wine which had undergone 28 Days fermentation was pasteurized (45 - 55 oC for 2 h ) to deactivate all enzymes that could be present in the wine. Five milliliter (5 ml) of commercial pectinase was pipette into 500 ml date palm wine maintained at 50 oC inside water bath for wine clarification. The setup was allowed to stand for 10 h. The clarified wine was filtered, bottled and allowed to age.

Clarification of date palm wine using two immobilized strains of Aspergillus tamarii on biomatrix

The processes involved in clarification of date palm wine by separately using immobilized Aspergillus tamarii TN-275 and A. tamarii strain DTO:129-ES are as follows:

Maduka et al 6.

Preparation of immobilizing material (SFN) Biomatrix which constitute trunk of structural fibrous network (SFN) of papaya was debarked, cut into small pieces (2 cm by 4 cm), thoroughly washed and soaked for 2-3 h inside distilled water changed 3-4 times. The soaked pieces of papaya were removed from the solution and oven dried to a constant weight at 80 oC and then kept inside a dessicator to cool.

Inoculum development

Date must prepared by dissolving 50 g date fruit into 50 ml distilled water was poured into two separate conical flasks and autoclaved at 121 oC at 15 psi for 15 min. and then allowed to cool. Aspergillus tamarii isolate TN-275 and Aspergillus tamarii Strain DTO:129-ES were separately inoculated into the two flasks and incubated at room temperature (28±2 oC) for 48 h. The content of each flask were standardized using 0.5 McFarland Standard in line with the procedure described by Chapin and Lauderdale [14] and ASM Manual of Clinical Microbiology [15].

Immobilization of Aspergillus tamarii TN-275 and A. tamarii strain DTO:129-ES on a biomatrix

Four (4) pre-weighed pieces of structural fibrous network (SFN) of papaya wood were put inside 200 ml date palm wine must contained in two Erlenmeyer flasks (500 ml) and sterilized at 121 oC at 15 psi for 15 min. Each of the flasks was separately inoculated with Aspergillus tamarii isolate TN – 275 (RF10) and Aspergillus tamarii strain DTO: 129 – ES (RF11) inoculums and incubated for 48 h at room temperature (28±2 oC). The flask containing 200 ml date palm must without biomatrix (SFN) added served as control. Under aseptic conditions, SFN that had entrapped fungal hyphae was removed from the flasks and washed thoroughly using date must to remove free hyphae. Compound microscope was used to verify presence of hyphae in the date must. Clarification of date palm wine using immobilized Aspergillus tamarii TN-275 and A. tamarii strain DTO:129-ES on a biomatrix Structural fibrous network (SFN) separately entrapped with hyphae of Aspergillus tamarii isolate TN-275 and Aspergillus tamarii strain DTO:129-ES were transferred separately to 100 ml fresh date wine must and incubated at room temperature (28±2 oC) for 7 Days. Both free and SFN – immobilized biomass of the two strains of Aspergillus were harvested from the medium, washed twice with distilled water and oven dried at 70 oC to constant weight, respectively. The SFN immobilized biomass was separated from date wine by decantation and clarification. Similarly, free-grown fungal biomass was separated from date palm wine by filtration using pre-weighed filter paper (Whatman № 1).

Measurement of clarity The method adopted was described by Awe [16] which involves using a spectrophotometer. Without agitation of the containers containing the samples, 25 ml of each sample were dispensed into 100 ml conical flask and gradually poured into a cuvette. Wavelength of the spectrophotometer was adjusted before optical density of the sample was analyzed and the reading was recorded.

Statistical analysis One-way ANOVA was used to analyze duplicate results obtained from each analysis with the aid of Statistical Package for the Social Sciences (SPSS) software version 21. The mean (M) and Standard Deviation of the duplicate results were calculated at 95 % confidence interval.

7. S. P. J. App Bio Res

RESULTS The result presented in Table 1 shows cultural and morphological characteristic of fungal isolates used in the study. Table 2 depicts the physicochemical parameters of freshly prepared date must used for production of date wine. Shown in Figure 1-9 is the changes in physicochemical parameters of freshly prepared date must undergoing fermentation for 27 Days monitored at 24 h intervals. Table 3 shows the absorbance value of commercial wine, date wine clarified with commercial pectinase, Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES immobilized on a biomatrix at different wavelength. Shown in Figure 10 is the average absorbance value of Date wine separately clarified using two Aspergillus tamari strains immobilized on biomatrix and white wine (control).

Table 1. Cultural and morphological characteristics of fungal isolates used in the study

Isolate code

Macroscopy Microscopy Presumptive identity

Yeast

,

Flat, moist, rough

glistening, dull and creamy

in colour. Diameter of 2-8

μm and length of 3-25μm.

Rough colonies, presence of buds

(blastoconidia) unicellular, globose

and ellipsoid, elongate in shape,

multilateral (multipolar)

budding, absence of hyphae.

Saccharomyces

cerevisiae

Immobilized cell

Radiate pale green and

conspicuously, echinulate

conidia, having heads with

phialides both directly from

the vesicle. Hyphae are

septate and hyaline.

Spreading yellow-green

colonies, mature vesicle bearing

phialides over their entire surface and

echinulate conidia

Aspergillus spp.

Table 2. Physicochemical parameters of freshly prepared date must

Parameter Result

Appearance/colour Brownish coloured semi-viscous liquid

Odour Characteristic of fresh palm wine

Taste Sugary Texture Smooth and slippery to the tongue

Temperature (oC) 22.8 Specific gravity 1.06 Viscosity (cps) 113 pH 6.3 Sugar content (%) 22 Total dissolved solids (mg/l) 1942 Total titratable acidity (%) 0.282 Alcoholic content (%) Nil

Maduka et al 8.

Figure 1: Change in viscosity during fermentation of date must for 27 Days Figure 2: Change in pH during fermentation of date must for 27 Days

Figure 3: Change in sugar content during fermentation of date must Figure 4: Change in specific gravity during fermentation of date must

for 27 Days for 27 Days

Figure 5: Change in total dissolved solid during fermentation of Figure 6: Change in alcohol content during fermentation of date must date must for 27 Days for 27 Days

0

20

40

60

80

100

120

140

160

180

200

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Vis

cosi

ty (

cp

s)

Day

0

1

2

3

4

5

6

7

1 3 5 7 9 11 13 15 17 19 21 23 25 27

pH

Day

0

2

4

6

8

10

12

14

16

18

20

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Su

ga

r c

on

ten

t (%

)

Day

0.97

0.98

0.99

1

1.01

1.02

1.03

1.04

1.05

1.06

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Sp

ecif

ic g

ra

vit

y (

sp.g

r.)

Day

0

200

400

600

800

1000

1200

1400

1600

1800

2000

1 3 5 7 9 11 13 15 17 19 21 23 25 27

TD

S (

mg

/l)

Day

0

1

2

3

4

5

6

7

8

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Alc

oh

ol (%

)

Day

9. S. P. J. App Bio Res

Figure 7: Change in total titratable acidity during fermentation of Figure 8: Change in reducing sugar during fermentation of

date must for 27 Days date must for 27 Days

Figure 9: Change in temperature during fermentation of date must for 27 Days

Table 3. Absorbance value (nm) of commercial wine, date wine clarified with commercial pectinase, Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES immobilized on a biomatrix

Wavelength

(nm)

Commercial wine

(Control)

Date wine clarified

with commercial

pectinase

Date wine clarified with

(RF10)

Immobilized on a

biomatrix

Date wine clarified

with (RF11)

Immobilized on a

biomatrix

700 0.780±0.030d

0.740±0.0028c

0.287±0.0042b

0.266±0.0028a

720 0.740±0.0014c

0.698±0.0042b

0.222±0.0028a

0.221±0.0056a

740 0.699±0.0035b

0.663±0.0028a

0.975±0.0057c

0.969±0.0056c

760 0.683±0.0014c

0.628±0.0028b

0.139±0.0042a

0.136±0.0028a

780 0.663±0.0028c

0.598±0.0042b

0.111±0.0028a

0.110±0.0056a

800 0.660±0.0028c

0.579±0.0042b

0.091±0.0049a

0.090±0.0028a

820 0.598±0.0057c

0.559±0.0057b

0.078±0.0028a

0.075±0.0042a

840 0.579±0.0035c

0.527±0.0028b

0.066±0.0057a

0.065±0.0028a

Values are duplicates of samples. Mean scores with same superscripts across the row are not significant at p<0.05, while

others with different superscripts are significant. Where RF10 - Aspergillus tamarii TN275 and RF11 - Aspergillus tamarii strain DTO:129-ES.

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

1 3 5 7 9 11 13 15 17 19 21 23 25 27

TT

A (

%)

Day

0

5

10

15

20

25

30

35

40

45

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Red

ucin

g s

ug

ar (

%)

Day

24

25

26

27

28

29

30

1 3 5 7 9 11 13 15 17 19 21 23 25 27

Tem

pera

ture (

oC

)

Day

Maduka et al 10.

Figure 10: Average absorbance value of date wine separately clarified using two Aspergillus tamari strains

immobilized on biomatrix and white wine (control)

DISCUSSION The result obtained from this study shows that sugar content (22 %) and dissolved solids (1942 mg/l) present in fresh date must was relatively high. This might have resulted in cloudiness of date must which could be less acceptable to consumers unless it is subjected to clarification. The date must was viscous (113 cps) and alcohol was not detected in the product. Meanwhile, total titratable acidity and pH of the product was 0.282 % and 6.3, respectively. These values are indication that low quantity of slightly acidic substances was present in freshly prepared date must. Temperature of the product (22.8 oC) was close to room temperature (28±2 oC). This could be as a result of reduced microbial activities in the freshly prepared date must. Fruit juices are naturally cloudy due to the presence of polysaccharides (pectin, cellulose, hemicelluloses, lignin and starch), proteins, tannins and metals [17]. Absorbance value of wine tested using spectrophotometer is an indication of wine clarity. This study has shown that at wavelength 700-840 nm, there was no significant difference between absorbance value of date palm wine separately clarified with Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES with the exception of their absorbance value at wavelength 700 nm. Meanwhile, average absorbance value of date fruit wine clarified using Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES at wavelength 700-840 nm was 0.2461 and 0.2415 nm, respectively. Our result further shows that at wavelengths 700-840 nm, absorbance value of white wine (control) and that of date palm wine clarified using commercial pectinase were significantly different. The average absorbance value of white wine (control) and date palm wine clarified using commercial pectinase was 0.6753 and 0.624 nm, respectively. Different substrate used for wine production and method of wine clarification could be the reason behind significant differences in absorbance value of white wine, date must and date wine separately clarified using Aspergillus tamarii TN275 and A. tamarii strain DTO:129-ES immobilized on a biomatrix.

Depectinisation of must through the use of pectinases has been reported as an efficient alternative to reduce turbidity of the product [7, 8]. Pectinases degrade pectin, thereby resulting in viscosity reduction and cluster formation which facilitates separation through centrifugation or filtration. These processes improve wine clarity [18, 19]. This justifies our results on clarification of date wine using commercial pectinases. Absorbance value of date wine clarified using Aspergillus tamarii isolate TN-

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

Control (White wine) Clarified with commercial pectinase

Clarified with immobilized RF10

Clarified with immobilized RF11

Av.

ab

sorb

ance

(n

m)

Product

11. S. P. J. App Bio Res 275 and Aspergillus tamarii strain DTO:129-ES immobilized on papaya trunk is an indication that it could be used for wine clarification. Date wine clarified using Aspergillus tarmarii isolate TN-275 showed slightly higher absorbance value when compared with that of Aspergillus tamari strain DTO:129-ES. It has been established that the use of natural polymeric gel for immobilization is more advantageous than synthetic gels because of reusability, freedom from toxicity problems, mechanical strength for necessary support and open spaces within the matrix for growing cells thus avoiding rupture and diffusion problems associated with polymeric gels and chemicals [18, 20].

Findings from this study showed that steady reduction in viscosity, sugar content, total dissolved solid, pH and reducing sugar occurred as the fermentation period of date must increased from Day 0-27 with few exceptions. Ogodo et al. [21] and Awe and Nnadoze [2] also reported similar trend of results from a related study. This could be as a result of fermentation of date must by Saccharomyces cerevisiae involve continuous utilization of solid substrates and release of end products. Low pH of wine is beneficial because it inhibit the growth of spoilage bacteria and favours the growth of desirable organisms.

In the course of the fermentation of date must, there was reduction in total soluble solids although the values were fluctuating. This could be attributed to breakdown of substrates by the organisms involved in the fermentation process [20]. During the fermentation period, total soluble solids reduced from 1810 - 1615 mg/l. Similarly, there was steady reduction in quantity of reducing sugar from 42 - 1.2 % within the fermentation period. Continuous utilization of reducing sugar by the fermenting organism could be responsible for steady reduction in quantity of reducing sugar available in the fermenting substrate.

Notably observed in this study was steady increase in alcohol content of date must within the period of fermentation from 0.5 - 7.0 %. This result trend is in agreement with a similar study carried out by Awe and Nnadoze [2]. Ogodo et al. [21] reported a similar increase in alcohol content during primary fermentation of fruit wine from mango. This could be attributed to ethanol, esters, carbonyl compounds, acids and acetyls being the products of alcoholic fermentation. These products are associated with pleasant aromas. Generally, alcohol content of alcoholic beverages influences level of consumer acceptability of the product. According to Awe and Nnadoze [2], a good wine is supposed to have 8 - 14 % alcohol content.

Also observed in this study was increase in total titratable acidity (TTA) of date must which could be attributed to slightly acidic nature of some fermentation end products. This could also be the reason for reduction in pH of date must from 6.1-3.9 within its fermentation period. According to Ogodo et al. [21], lowering of pH and concomitant increase in total titratable acidity during fermentation of must could be attributed to presence of organic acids such as acetic, lactic, formic etc. The release of phenolic compounds, carbon dioxide and esters could also influence lowering the pH of the fermenting must. In a related study, Awe and Nnadoze [2] also reported increase in TTA during fermentation of date must by Saccharomyces cerevisiae.

In the course of fermentation of date must, there was fluctuation in temperature similar to what was reported by Ogodo et al. [21] from a related study. Despite the fluctuations, the temperature of the fermenting substrate decreased from 29.0 o C at the beginning of fermentation to 25.8 oC at Day 27 when the process was discontinued. Biochemical changes which result from activities of organisms metabolizing the substrate could be responsible for fluctuation in temperature of the substrate from 29.0 - 25.8 oC.

Although there was reduction in specific gravity of date must within the fermentation period, the values were relatively stable which range from 1.055 - 1.00. The values obtained were within the range suitable for wine. Our results is in agreement with a related study carried out by Awe and Nnadoze [2] which reported that during fermentation of date must by Saccharomyces cerevisiae, its specific gravity dropped from 1.070 - 1.026.

Maduka et al 12. CONCLUSION Commercial pectinase was a more effective clarifying agent than Aspergillus strains immobilized on papaya trunk. However, white wine had higher clarity than date wine clarified using commercial pectinase or Aspergillus strains immobilized on a biomatrix. Findings from this study further revealed that average absorbance of date wine separately clarified using Aspergillus tamarii isolate TN – 275 and Aspergillus tamarii strain DTO:129-ES immobilized on a biomatrix were almost of the same value. Since it had previously been reported that the use of biostructures in wine clarification does not pose toxicity problems among other advantages over synthetic polymeric materials, this study have demonstrated the potential of using Aspergillus tamarii isolate TN – 275 or/and Aspergillus tamarii strain DTO:129-ES immobilized on papaya trunk for wine clarification as a strategy to boost local wine production.

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Cite this article as:

Maduka et al (2019). Assessment of Date Wine Clarified Using Commercial Pectinase and

Aspergillus tamarii Strains Immobilized on Biomatrix

SPJABR 2(2)019-031

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