EFFECT OF GIBBERELLIC ACID (GA3) ON ENHANCING...

307 Annals

Agric. Sci., Ain Shams Univ.,

Cairo, 54(2), 307-321, 2009

(Received August 23, 2009) (Accepted September 6, 2009)

DEXTRAN PRODUCTION BY SOME LOCAL Leuconostoc mesenteroides STRAINS

[24] Abdel-Azeem, Hoda1 H.M.; Gehan F. Galal2 and Enas A. Hassan2

1- Desert Research Center Unit of Microbiology, Matariya, Cairo, Egypt

2- Department of Agricultural Microbiology, Faculty of Agriculture, Ain Shams University,

Shoubra El-Kheima, Cairo, Egypt

Keywords: Dextran, Leuconostoc mesenteroides,

Viscosity, Nutritional requirement, Environmental

factor, Static batch culture

ABSTRACT

Eighty dextran-producing bacteria were ob-

tained from different food stuffes and tested for

dextran production. These isolates were classified

into four categories namely, high, moderate, weak

and inactive according to their efficiency to pro-

duce dextran. The most efficient dextran-producing

isolates (first category) were presented in the per-

centage of 7.5% and isolated from sugar cane

juice, tomato juice, milk and infected lettuce.

McCleskey’s medium was superior for dextran

production by these isolates than other tested me-

dia. The production of dextran was obtained during

the end of logarithmic phase till the start of statio-

nary phase (10-16 hr). Only one isolate (No. IL1)

recorded the highest dextran yield as well as con-

version coefficient on the natural medium (WBE3).

This isolate was completely identified as strain of

Leuconostoc mesenteroides subsp. mesente-

roides. The effect of some nutritional and environ-

mental requirements of the dextran production by

L. mesenteroides (IL1) using surface culture tech-

nique were studied. The results indicated that the

maximum dextran production was obtained by

growing the isolates on wheat bran medium con-

taining 100 ml L-1

wheat bran extract (5%), sucrose

10%, yeast extract 0.5%, tryptone 0.25%, K2HPO4

0.5% and pH = 8.0 then incubated at 25ºC for 16

hrs.

INTRODUCTION

Dextran is a polysaccharide consisting of glu-

cose monomers linked mainly (95%) by (1-6)

bonds. Commercial application for dextran is gen-

erally in the pharmaceutical industry, but new ap-

plications are being considered in the food and

textile industries (Shamala and Prasad, 1995).

The effect of nutritional requirements and environ-

mental factors on the production of dextran by

Leuconostoc mesenteroides growing in different

media as a batch culture was studied by many

investigators (Santos et al 2005, Kim et al 2000

and Son et al 2008). The bacterium is grown in a

sucrose-rich media releasing an enzyme, dextran-

sucrase, which converts excess sucrose to dextran

and fructose, when high sucrose concentration is

used to produce dextran and fructose, broth vis-

cosity becomes high and process control becomes

more difficult. Acceptor molecules, such as mal-

tose present in the culture media can influence

dextran molecular weight by allowing the growing

chain to be separated from the enzyme active site

and transfer to the acceptor (Dols et al 1998).

Santos et al (2005) found that dextran molecular

weight decreases when complementary sugar

(maltose, lactose and galactose) together with su-

crose were used. Whereas, Kim and Robyt (1995)

get four mutants produced extracellular dextransu-

crases on glucose with higher activities (2.5-11

times) than the parent strain on sucrose. Differ-

ences in physical appearance, solubility and sus-

ceptibility to endo-dextranase hydrolysis of the

Abdel-Azeem, Hoda; Gehan Galal and Enas Hassan

Annals Agric. Sci., 54(2), 2009

308

dextrans prepared by different mutants grown on

glucose and sucrose were found.

In fermentation by Leuconostoc sp., the factors

that affect dextran production include the type and

concentration of salts, temperature, pH and su-

crose concentration (Kim et al 2000). Also, Son et

al (2008) stated that the type and production of

dextran were greatly affected by the sucrose con-

centration, food ingredients and fermentation time.

The culture broth with the highest consistency, was

obtained from the defined medium containing 20%

sucrose, 1.5% skim milk and 0.5% potato powder

and it showed typical pseudoplastic behavior with

a pH of 4.07.

Santos et al (2000) stated that the optimum

conditions for dextran and fructose production

were T = 35ºC and pH 6.5. Whereas, Kim et al

(2003) reported that the temperature had very little

effect on the dextran molecular weight but it has a

significant effect on the degree of branching, at

4ºC and increased to 14.7% at 45ºC. Both the mo-

lecular weight (Mw) and the degree of branching

not significantly affected by different pH values

between 4.5 and 6.0. Shamala and Prasad (1995)

noted that temperature of 23-26ºC unaerated fer-

mentation of 10-20 % sucrose by L. dextranicum

FRW-10 increased culture viscosity.

The aim of the present study is to evaluate the

potential of using local bacterial strains for dextran

production. The study focused on isolation of dex-

tran producing bacteria from different foodstuffs

and identified as wells as, optimization conditions

for dextran production using static, batch culture

technique.

MATERIALS AND METHODS

1. Samples

Sample of sugar cane, Apple, Tomato juices as

well as cucumber, lettuce, milk and pickles (lemon

& carrot) were collected from local markets of Cai-

ro, Egypt. The samples were directly transferred

into the laboratory for the isolation of dextran pro-

ducing bacteria.

2. Media used

- McCleskey’s medium (McCleskey et al 1947)

was used for isolation and dextran production from

dextran- producing bacteria.

- Kurt’s medium (Kurt and Curt, 1983) was

used for dextran production.

- Wheat bran extract media (Shamala and

Prasad, 1995).

Three media containing wheat bran extract

(WBE1, WBE2 & WBE3 media) were used for dex-

tran production.

- WBE1 medium containing sucrose, 100 g;

Yeast extract, 5 g and WBE, 100 ml/L.

- WBE2 medium containing sucrose, 100 g;

Tryptone, 1.5 g; and WBE 100 ml/L.

- WBE3 medium containing sucrose, 100g; Y.

extract, 5 g; Tryptone, 2.5 g; K2HPO4, 5 g; WBE,

100 ml; Distilled water, 900 ml and pH 6.7.

Wheat bran extract (WBE) was prepared by

soaking 10 g of wheat bran in 80 ml distilled water

under stirring for 1 h. The soaked slurries were

filtered and the residues were washed and filtered

again to collect 80 ml filtrate, the pH of extract was

adjusted to 5.5, heated till boiling ,cooled and cen-

trifuged at 8000 rpm.

- Preservation medium (Shamala and Prasad,

1995) was used for propagation and preservation

of different dextran producing cultures.

3. Isolation of dextran producing bacteria

The representative samples of infected cu-

cumber and infected lettuce (each weighing 10 g)

or juices, milk and pickles lemon (10 ml each) were

mixed or diluted by 100 ml sterile tap water and

thoroughly shaken for 10 minutes. Streak and pour

plate methods were used for isolation of dextran-

producing bacteria from these samples on

McCleskey’s medium .The developed colonies on

the plates (incubated at 25ºC for 1-3 days) were

picked under aseptic conditions, purified and mi-

croscopically examined. Isolates were maintained

on slants of the preservation medium at 5ºC.

4. Maintenance of cultures

Stock culture slants were maintained at 5ºC on

preservation medium after incubation at 25ºC for

24 hr.

5. Standard inoculum

Standard inoculum was prepared by inoculation

of conical flasks (250 ml in volume) containing 50

ml of preservation medium with a loop of tested

culture. The inoculated flask was incubated at

25ºC for 24 hrs. The content of this flask was used

as a standard inoculum (O.D. ranged from 0.2 x 10

to 0.3 x 10) for production experiments.

6. Screening for efficient dextran producing

bacteria

All isolates were tested for their culture viscosi-

ties to select the most efficient dextran-producing

Dextran production


309

isolates.Conical flasks 250 ml containing 100 ml

McCleskey’s broth were inoculated with the tested

isolate and incubated at 25ºC for 72 hrs as a static

batch culture, 10 ml representative sample was

withdrawn every day for testing viscosity by visco-

meter. Efficient isolates were selected for further

studies.

7. Selection of suitable medium for dextran

production

Five media for dextran production being

McClesky’s medium (McCleskey et al 1947),

Kurt’s medium (Kurt and Curt, 1983) and WBE,

media No. 1,2 & 3 (Shamala and Prasad, 1995)

were used in this experiment for dextran produc-

tion in order to select the most suitable media for

screening high dextran production. The fermenta-

tion and determination of culture viscosity were

done as mentioned before.

8. Biological activity of dextran producing bac-

teria

In this investigation, the most efficient dextran-

producing isolates were grown in Erlenmeyer

flasks (500 ml in volume) containing 100 ml suita-

ble medium and incubated at 25ºC for 24 hrs as a

static batch culture. Samples (10 ml) were taken

from the growing culture periodically every 2 hrs

under aseptic conditions to determine the optical

density of growth, culture viscosity and pH. The

relation between time and optical density (growth

curve) was plotted on semi-log paper. Growth pa-

rameters such as specific growth rate, hourly

growth rate, generation time, multiplication rate

and number of generation were calculated from the

exponential phase.

At the end of fermentation period (24 hr), 10 ml

sample of culture fluid were diluted with distilled

water to 50 ml and mixed to reduce the viscosity

and then centrifuged at 14000 rpm for 30 minutes,

the sediment (bacterial cells) was washed twice

with distilled water, then dried at 70ºC to a con-

stant weight. The residual sugar was determined in

the supernatant. Dextran produced was precipi-

tated by ethanol from supernatant, dried and de-

termined as dry weight. Dextran yield, conversion

coefficient, productivity, specific viscosity rate,

hourly viscosity rate, sugar utilization efficiency and

yield factor were calculated.

9. Ultraviolet irradiation

According to the method of Carlton and

Brown (1981), vegetative cells (in phosphate buf-

fer suspension) were dispensed into 5 ml aliquots

in a sterile – glass Petri dishes (80 mm in diame-

ter) then exposed to a prewarmed (15-20 minutes

before use) short-wave ultraviolet lamp (254 nm) at

20 cm distance.

The bacterial suspensions in Petri dishes were

stirred with vigorous magnetic stirring for 1,2,3,4

and 6 minutes. Immediately after irradiation, 1 ml

sample of each treatment, suspended into 9 ml

nutrient broth and incubated for two hours, under

dim light to avoid photoreactivation, at 32ºC to al-

low segregation of the newly mutated chromosome

from the nonmutated one in the same cell. Three

replicates for each exposure period were carried

out. After 2 hours, each irradiated bacterial culture

was serially diluted and plated onto nutrient agar.

10. Factors affecting dextran production

The propagation was carried out in Erlenmyer

flask (250 ml in volume) containing 100 ml me-

dium. These flasks were inoculated with 5 ml stan-

dard inoculum and incubated for 16 hrs at 25ºC. At

the end of incubation period, viscosities of cultures,

growth density as optical density and final pH val-

ues were determined.

10.1. Sucrose concentrations

Three trial of sucrose concentrations, i.e 10, 20

& 30% were used to study their effect on dextran

production by L. mesenteroides subsp. mesente-

roides (IL1) and its mutants.

10.2. Nitrogen source

The nitrogen sources applied were yeast ex-

tract, tryptone, peptone, casein, beef extract, am-

monium phosphate, ammonium sulphate, ammo-

nium nitrate, ammonium chloride and a mixture of

yeast extract and tryptone. WBE3 medium without

the nitrogen source was used as a basal medium.

The amount of nitrogen compound added to the

media was calculated to give a final concentration

of 1.02 g nitrogen/L.

10.3. Wheat bran concentrations

Different wheat bran concentrations ranging

from 5 to 20% were used to study their effect on

dextran production.

10.4. Wheat bran extract’s volume

Different wheat bran extract volumes ranged

from 2.5 to 20% were added to WBE3 medium in

order to select the best treatment for dextran pro-

duction by L. mesenteroides subsp. mesente-

roides.



310

10.5. Initial pH

Eleven levels of initial pH (ranging from 4.5 to

9.5) were chosen for studying their effect on dex-

tran production.

10.6. Incubation temperature

Six different degree temperature, i.e. 20, 22,

24, 26, 28 and 30ºC were investigated to detect

the optimum temperature for dextran production.

10.7. Inoculum size

Different volumes of standard inoculum ranging

from 1 to 6 ml (O.D ranged from 0.2 x 10 to 0.3 x

10) were used to inoculate 100 ml WBE3 medium

to detect the suitable inoculum size. Then followed

the previous procedures till the end of the incuba-

tion period in order to determine viscosity, optical

density and pH of cultures.

11. Chemical determination

- Dextran was determined either viscometrically

or by dry weight culture. Viscosity as an index of

dextran production was measured at 25ºC with a

(Cole-C Parmer) rotational viscometer at a con-

stant speed of 0.6 rpm using spindle number 5.

- The method of Bailey and Oxford (1958)

was followed for precipitation and purification of

dextran.

- Total sugar was determined using Anthrone

method according to Trevelyan and Harrison

(1956).

12. Calculation

The specific growth rate (µ) and doubling time

(td) were cauculated from the exponential phase

according to Painter & Marr (1963).

Yield factor, carbon utilization efficiency and

dextran parameters (conversion coefficient, yield

and productivity) were also calculated.

RESULTS AND DISCUSSION

1. Isolation and selection of dextran-producing

bacteria

In this study, eighty dextran producing bacteria

were isolated from different foodstuffs and tested

on McCleskey’s medium for dextran production.

These isolates were characterized by forming

round, convex and slimy colonies with smooth

margins. Cells were gram-positive cocci, non-

motile and occurred in pairs or chains.

Table (1) shows that the highest percentage of

bacterial isolates was obtained from sugar cane

juice followed by pickles lemon being 43.75% and

18.75%, respectively. 10% of the total isolates

were isolated from tomato juice or infected cu-

cumber. The lowest percentage of isolates was

obtained from apple juice or infected lettuce being

5%. The culture viscosity of these isolates varied

from one isolate to another as well as fermentation

times (from one to three days). So, these isolates

were classified into four categories according to

their culture viscosity produced. These groups

namely, high, moderate, weak and inactive dex-

tran-producing isolates which gave culture viscosi-

ty ranged from 4000 to > 8000 cP, 1000 to > 4000

cP, 100 to >1000 cP and >100 cP, with the inci-

dence percentage of 7.5, 15, 46.25 and 31.25%,

respectively. Also, it could be noticed that most

efficient dextran-producing isolates were obtained

from sugar cane juice, tomato juice, milk and in-

fected lettuce which represent 3.75% (No. SCJ7,

SCJ25 & SCJ28), 1.25% (No. TJ2), 1.25% (No.

M3) and 1.25% (No. IL1) of the total isolates, so,

these isolates were selected for further studies.

2. Selection of suitable medium for dextran-

production

Data presented in Table (2) clearly show that

the culture viscosities obtained by different cultures

grown on McClesky’s medium during 3 days fer-

mentation period at 25ºC were higher than the

other four tested media except isolate IL1 which

gave approximately the same values of viscosity

on wheat bran extract medium No. 3. The viscosity

of McCleskey’s medium varied greatly from one

isolate to an other. The isolates No. IL1, TJ2, M3

and SCJ28 gave the highest culture viscosity being

7790, 7785, 7400 and 7300 cP after the first day of

incubation. Also, SCJ25 recorded the highest vis-

cosity being 7792 cP after 3 days of incubation

whereas SCJ7 gave the lowest figure on the same

medium. So, it could be stated that McClskey’s

medium was chosen as preferable medium for

dextran production by all isolates and also WBE3

medium was chosen for isolate No. IL1. Therefore

these isolates were tested for their biological activi-

ty expressed as growth parameters and dextran

parameters on their preferable media.

3. Biological activity of dextran-producing iso-

lates

Results illustrated by Fig. (1) clearly show that

isolates grew exponentially during the first 2-10 hrs

Dextran production


311

Table 1. The percentage of active, moderate, weak and inactive dextran producing isolates from

different foodstuffs

Foodstuffs

Total

No.

Isolates

% High Moderate Weak Inactive

No. % No. % No. % No. %

Sugarcane juice 35 43.75 3 3.75 4 5.0 13 16.25 15 18.78

Tomato juice 8 10 1 1.25 1 1.25 6 7.0 0.0 0.0

Apple juice 4 5 0.0 0.0 0.0 0.0 1 1.25 3 3.75

Milk 6 7.5 1 1.25 0.0 0.0 5 6.25 0.0 0.00

Pickles lemon 15 18.75 0.0 0.0 7 8.75 5 6.25 3 3.75

Infected cucumber 8 10 0.0 0.0 0.0 0.0 6 7.5 2 2.5

Infected lettuce 4 5 1 1.25 0.0 0.0 1 1.25 2 2.5

Total 80 6 7.5 12 15 37 46.25 25 31.25

* High = Isolates producing culture viscosity ranged from 4000 - >8000.

** Moderate = Isolates producing culture viscosity ranged from 1000 = > 4000 cP

* Weak = Isolates producing culture viscosity ranged from 100 to > 1000 cP.

** Inactive = isolates producing culture viscosity ranged from 15 > 100 cP

cP = Centipoises

incubation period on McCleskey’s medium. The-

reafter the growth rate decreased gradually (phase

of decleraing growth) to be more constant (statio-

nary phase) at the last ten hours of incubation. The

highest figures of growth (expressed as optical

density) were recorded for IL1, M3 and SCJ28

isolates being 2.6, 2.21 and 2.0, respectively after

10 hrs of incubation at 25ºC on McClesky’s me-

dium. Also, these isolates recorded the highest

figures of specific growth rate, hourly growth rate,

number of generation and multiplication rate as

shown in Table (3).

On WBE3 medium, isolate No. IL1 grew expo-

nentially during the first 2-24 hr to reach the maxi-

mum growth (as optical density) being 2.35 after

24 hr incubation at 25ºC. The growth parameters

obtained by this treatment were 0.303 h-1

, 1.354,

2.29, 3.49 h and 0.44 for specific growth rate,

hourly growth rate, doubling time, number of gen-

eration and multiplication rate, respectively. Slight

increase in culture viscosity of all the tested iso-

lates was observed during the first 8 hrs of incuba-

tion, while the rate of increase was higher at the

end of exponential phase and during stationary

phase to reach the maximum value after 16 hrs.

The highest range of culture viscosity was ob-

served on McCleskey’s medium by IL1 and TJ2

being 7783-7790 cP and followed by M3 & SCJ28

being 7437-7490 cP, whereas IL.1 gave 7100 cP

on WNE3 medium.

Regarding to the relation between consumed

sugar and growth or dextran production after 24

hrs on McCleskey’s or WBE3 med., data presented

in Table (4) clearly show that all the tested isolates

consumed high amount of sucrose ranged be-

tween 60.0 to 74.0 gL-1

and recorded high percen-

tage of sugar utilization efficiency. The lowest con-

sumed sucrose was observed by IL.1 on WBE3

medium being 60 gL-1

. After 24 hrs fermentation

period, all the tested isolates gave biomass dry

weight ranged from 2.4-2.54 gL-1

whereas the

amount of dextran dry weight ranged from 10.25-

14.15 gL-1

.

Although the isolates No. TJ2 and IL.1 gave the

highest dextran yield and dextran productivity on

McClesky’s med., but isolate No. IL.1 gave the

highest yield factor and conversion coefficient be-

ing 4.15% & 21.6% respectively on WBE3 medium.

From the aforementioned results it could be no-

ticed that the dextran production were concurrent

with the growth during the end of logarithmic phase

till the start of the stationary phase (10-16 hr) for all

the tested isolates either on McCleskey’s medium

or on WBE3 medium. These results also indicate

that 16 hr fermentation period was the favourest

for dextran production by all the tested isolates.

Inspite of isolate No. IL1 gave slight decrease

in dextran production on WBR3 medium comparing

to McCleskey’s medium, but the former medium

should be taken in consideration due to the low



312

Dextran production


313

0

1000

2000

3000

4000

5000

6000

7000

8000

0 2 4 6 8 10 12 14 16 24 26 28 48

Incubation period (hours)

Cu

ltu

re v

isco

sity

(cP

)

IL 1 in WBE medium

0.01

0.1

1

10

0 2 4 6 8 10 12 14 16 24 26 28 48

Incubation time (hours)

Op

tica

l d

ensi

ty (

OD

)

IL 1 in WBE medium

In McClesky medium

0

1000

2000

3000

4000

5000

6000

7000

8000

0 2 4 6 8 10 12 14 16 24

Incubation period (hours)

Cu

ltu

re v

isco

sity

(cP

)

ASJ 25

SCJ 28

In McClesky medium

0.01

0.1

1

10

0 2 4 6 8 10 12 14 16 24


Op

tica

l d

ensi

ty (

OD

)ASJ 25

SCJ 28

In McClesky medium

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

0 2 4 6 8 10 12 14 16 24


Cu

ltu

re v

isco

sity

(cP

) M 3

TJ 2

IL 1

In McClesky medium

0.01

0.1

1

10

0 2 4 6 8 10 12 14 16 24


Op

tica

l d

ensi

ty (

OD

) M 3

TJ 2

IL 1

Fig. 1. Culture viscosity and growth curves of the most efficient dextran producing cultures

grown on McClesky medium and wheat bran extract medium (WBE) during 24 hours incubation at 25°C

Table 3. Specific growth rate (µ), hourly growth rate (HGR), doubling time (td) number

of generation (N) and multiplication rate (MR) of the most efficient dextran-

producing isolates grown on McClesky’s and WBE3 media at 25ºC

Bacterial isolates µ

(h-1

) HGR

td

(h) N MR

McClesky’s medium

SCJ25

SCJ28

M3

TJ2

IL1

WBE3 med.

IL.1

0.364

0.40

0.415

0.388

0.452

0.303

1.439

1.492

1.514

1.474

1.571

1.354

1.9

1.73

1.67

1.79

1.53

2.29

4.21

4.62

4.79

4.47

5.22

3.49

0.53

0.58

0.60

0.56

0.65

0.44



314

Dextran production


315

cost price of the constituent medium. Therefore,

the isolate No. IL.1 was identified according to

Bergey’s Manual (1994) which proved to be Leu-

conostoc mesenteroides subsp. mesenteroides.

4. Ultraviolet irradiation

Data presented in Table (5) show the survival

percentage of L. mesenteroides (IL.1) after expos-

ing at different times to UV (254 mm) at 20 cm

distance. The survival percentages were 86.4,

65.6, 50.0, 20.0 and 0.01 after exposing to UV for

1, 2, 3, 4 and 6 minute respectively. All the mutant

strains were tested for their ability to grow on dif-

ferent concentrations of sucrose namely 10, 20

and 30%. Five mutant strains varied in their beha-

viour.

5. Factors affecting the production of dextran

The natural medium (WBE3) was used to se-

lect the most suitable ingredients and environmen-

tal factors for securing high dextran production by

Leuconostoc mesenteroides (IL.1) and its mutants

No. 1,2,3 & 4.

5.1. Sucrose concentration

Data presented in Table (6) show that there

was a gradual decrease in culture viscosity of all

strains except mutant No. 4 (C6) and mutant No. 5

(A5) with the increase of sucrose concentration

from 10% to 30% except L. mesenteroides mu-

tants No. 4 (C6) and No. 5 (A5) which recorded the

highest culture viscosity at 20 and 30% sucrose,

respectively. The highest culture viscosity was

obtained by L. mesenteroides (IL.1) and mutant

No. 3 (C4) at 10% sucrose being 7213 and 6715

cP followed by mutant No. 4 (C6) and mutant No. 2

(B3) at 20% and 10% sucrose being 6698 and

6310 cP, respectively. The lowest culture viscosity

was observed by mutant No. 5 (A5) at all sucrose

concentrations. The growth of all strains gave the

same trend at different sucrose concentrations.

The highest growth which expressed as optical

density (O.D) was observed by L. mesenteroides

(IL.1) followed by mutant No. 3 (C4) and No. 4 (C6)

being 5.0 and 4.6 or 4.5, at 10% and 20% sucrose

respectively. The corresponding figures of final pH

values were 3.63, 3.79 and 4.08, respectively.

Generally, it could be concluded that 10% su-

crose was the best concentration for the growth

and dextran production by L. mesenteroides (IL.1)

and mutant No. 3 (C4), whereas 20% sucrose was

suitable for the growth and dextran production by

mutant No. 4 (C6). These results are in agreement

with those of Son et al (2008) who reported that

the highest dextran yield was obtained at 20% su-

crose.

5.2. Nitrogen source

Generally, it could be stated that all the tested

organic sources has a drastic effect on dextran

production (viscosity), by all the tested strains

comparing to the control treatment. The highest

value of culture viscosity, pH & optical density were

obtained by L. mesenteroides (IL.1) being 7200

cP, 3.91 and 4.5, respectively (Table 7).

Table 5. Survival percentage of Leuconostoc mesenteroides (IL.1) after exposing at differ-

ent time to UV (254 nm) at 20 cm distance

Time/min Dilution Replicate No.

Average Survival rate

(%) I II III

0

0

0

10-8

10-6

10-7

240

389

345

260

374

330

250

382

337

250

381

337

1 10-6

10-7

300

285

310

300

305

290

305

291

ــــــ

86.4

2 10-6

10-7

255

210

260

225

263

230

259

221

ـــــ

65.6

3 10-5

10-6

250

197

242

180

245

195

245

190

ــــ

50.0

4 10-5

10-6

100

83

90

75

85

70

91

76

ــــ

20.0

6 10-3

10-4

35

10

20

16

40

15

31

13

0.01

ــــ

Not calculated ــــ



316

Dextran production


317



318

Fig. 2. Culture viscosity, final pH and optical density of Leuconostoc mesenteroides (IL1) as in-

fluenced by some nutritional and environmental factors i.e. wheat bran extract concen-

trations (A), volumes ofwheat bran extract (B), initial pH (C), temperature (D) and inocu-

lum size (E) after 16 hours incubation period at25°C

B

0

1

2

3

4

5

6

7

25

50

75

10

0

12

5

15

0

17

5

20

0

Volume of wheat bran extract (5 %) ml

Op

tical

den

sity

an

d

fin

al

pH

0

2000

4000

6000

8000

10000

Cu

ltu

re v

isco

sity

(cP

)

Final pHOptical density (OD)Culture viscosity (cP)

C

0

2

4

6

8

10

4.5 5

5.5 6

6.5 7

7.5 8

8.5 9

9.5

Initial pH

Op

tica

l d

ensi

ty a

nd

fin

al

pH

0

2000

4000

6000

8000

10000

Cu

ltu

re v

isco

sity

(cP

)

Final pHOptical density (OD)Culture viscosity (cP)

Dextran production


319

5.3. Wheat bran extract concentration

Data illustrated by Fig. (2,A) show that no re-

markable differences could be detected between

culture viscosity and growth density values in

WBE3 medium supplemented with wheat bran

extract 5% or 10% (control on WBE3 medium).

Increasing wheat bran extract more than 10% re-

sulted in decrease in the yield of dextran and

growth density values. So, 5% wheat bran will be

added to WBE3 medium instead of 10% in the

further studies.

The aforementioned results are nearly similar

to those obtained by Shamala and Prasad (1995)

who stated that the maximum viscosity achieved

by L. dextranicum FPW-10 was 6900 Cp in 5%

wheat bran extract (WBE), then decreased to 2800

cP at 20% WBE.

5.4. Volume of wheat bran extract

The effect of volume of 5% wheat bran extract

on the growth and dextran production by L. mesen-

teroides (IL.1) was studied by increasing WBE

volume from 25 to 200 ml/L. Data in Fig. (2,B)

show that the culture viscosity of L. mesenteroides

(IL.1) increased with increasing WBE volume till

reaching the maximum value being 7647 cP in

WBE3 medium containing 100 ml/L 5% wheat bran

extract. Increasing the volume of WBE from 100 to

175 ml/L didn't increase the culture viscosity but

200 ml/L decreased the culture viscosity about

11.63%. Data also revealed that the highest

growth density and pH values being 5.8 and 4.43

were recorded at higher volume of WBE being 200

and 150 ml/L, respectively.

5.5. Initial pH

Results in Fig. (2,C) indicate that pH 8.0 was

the most favourable one for dextran production by

L. mesenteroides (IL.1). Moreover, bacterial growth

was higher when initial pH ranged from 6.5 to 9. It

seems therefore that the initial pH ranged between

7 to 8.5 was suitable for growth and dextran pro-

duction. At initial pH ranged between 4.5 to 6.0 or

9.5 no viscosity was produced and lower bacterial

growth was recorded. Veljkovic et al (1992) rec-

orded that optimum pH for cell growth and dextran

production ranged between 6.0 to 6.9.

5.6. Incubation temperature

Data illustrated by Fig. (2,D) clearly show that

the culture viscosity and growth density of L. me-

senteroides (IL.1) were increased by increasing

incubation temperature till reached the maximum

being 7739 cP at 25ºC. It is interesting to notice

that increasing the temperature from 25ºC to 26ºC

decrease the yield of dextran about 14.2%. Gener-

ally, it could be noticed that the high efficiency of L.

mesenteroides to produce dextran were attained at

the low tested temperature which ranged between

20 to 25ºC. This means that high level of dextran-

sucrase which converted sucrose to dextran was

produced at lower temperature than was needed

for maximal bacterial growth. These results are not

in accordance with those of Kim et al (2003) who

stated that biosynthetic efficiency of dextran was

attained at the same temperature of bacterial

growth of Leuconostoc strains being 28ºC.

Final pH values showed a sharp drop than ini-

tial pH (8.5) ranging between 3.65-4.25. Finally, it

could be recommended to incubate the culture of

L. mesenteroides (IL.1) for 16 hours at 25ºC in

static culture for optimum dextran production.

5.7. Inoculum size

Results illustrated by Fig. (2,E) show that in-

oculation with 3-5% of standard inoculum gave the

highest culture viscosity being 7767 cP by L. me-

senteroides (IL.1) after 16 hours incubation period

at 25ºC. The corresponding figures for growth den-

sity and final pH were 4.9-5.2 and 4.0- 3.6 respec-

tively. The variation in size of inoculum had a neg-

ative effect on pH values where the pH decreased

by increasing the size of inoculum.

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