properties of thermally-treated wheat gluten films

ELSEVIER Industrial Crops and Products 6 (1997) 177- 184

INDUSTRIAL CROPS AND PRODUCTS

AN INTERNATIONAL JOURNAL

Properties of thermally-treated wheat gluten films’

Yusuf Ali, Viswas M. Ghorpade, Milford A. Hanna *

Industrial Agricultural Products Center, Department of Biological Systems Engineering, University of Nebraska-Lincoln, Lincoln NE 68583-0730, USA

Received 8 October 1996; accepted 27 December 1996

Abstract

Effects of thermal treatments on selected properties of wheat gluten film were studied. Films were cast from heated alkaline aqueous solutions of wheat gluten, ethanol and glycerin and subsequently heat treated at 65, 80, or 95°C for 2, 4, 6, 12, 18, or 24 h. Water vapor permeability (WVP), Hunter L, a and b values, tensile strength (TS) and elongation at break (E) were determined and compared with untreated film (control). Significant reduction in WVP of film occurred with increasing curing temperature and exposure time. Hunter L value (whiteness) decreased, whereas a (redness) and b (yellowness) values increased with increasing heat treatment temperature and exposure time. Also, an increase in TS and a decrease in E were present with increasing treatment temperature and exposure time. 0 1997 Elsevier Science B.V.

Keywords: Wheat gluten film; Water vapor permeability; Film property

1. Introduction

Development of edible films from starch and protein, and non-edible biodegradable films from synthetic polymers have received increased inter- est in the past several years because of growing concern over solid waste management of non-

* Corresponding author. Fax: + 1 402 4726338; e-mail: [email protected]

’ Journal Series Number 11717 of the University of Ne- braska-Lincoln, Agricultural Research Division.

biodegradable plastics. Edible biopolymer films

offer a number of advantages over conventional

synthetic packaging materials including their re-

newable and biodegradable nature, nutritional

supplementation of packaged food, application in

the interior of foods to control intercomponent

moisture and solute migration, individual packag-

ing of small-sized food products, use as an inter-

nal food contact layer in multi-layer food

packaging material, and potential for microencap- sulation and controlled release of food additives

such as preservatives and flavorings (Gennadios

and Weller, 1990).

0926-6690/97/$17.00 0 1997 Elsevier Science B.V. All rights reserved

PII SO926-6690(97)00216-l

178 Y. Ali et al. /Industrial Crops and Products 6 (1997) 177-184

Several studies have been reported on produc- tion and properties of films made from starch, protein and lipid substances. Potential applications and properties of edible films from starch and proteins have been reviewed by Kester and Fennema (1986); Guilbert (1986); Gennadios and Weller (1990, 1991). They reported that edible films can affect migration of water, oxygen, lipid and flavor in many food systems. Preparation and properties of edible films from soy protein and wheat gluten have been reported by Gontard et al. (1993); Gennadios et al. (1993a,b); Ghorpade et al. (1995, 1996), from sodium caseinate and whey protein isolate by McHugh et al. (1994), and cellulose-based films by Park et al. (1993). Glyc- erin is used as a plasticizer in film-forming solutions to make films more flexible (Brandenburg and Weller, 1993). Gontard et al. (1993) reported that glycerin and water plasticized with cast wheat gluten films decreased puncture strength, im- proved elasticity, and increased extensibility and water vapor transmission rate. Without plasticizer the films are too brittle to handle.

Water vapor permeability (WVP) of hydrophilic films is affected by film thickness and partial pressure of water vapor at the film inter- face (McHugh et al., 1993; Gennadios et al., 1994). Most edible films are hydrophilic in nature, which results in characteristic positive slope rela- tionships between thickness and WVP (McHugh et al., 1993). Several explanations have been provided for this anomalous effect of thickness. Hauser and McLaren (1948) attributed the thickness effect in regenerated cellulose films to different structures being formed at different thicknesses. Banker et al. (1966) explained that film thickness may have been affected by development of attractive forces between fihns and water, causing film swelling and consequently varying film structures. Hagenmaier and Shaw (1990) also reported thickness effects in hydroxy propyl methylcellulose films. McHugh et al. (1993) suggested that WVP values could be determined by a WVP correction method and these values, as well as film thickness, should be reported in edible film studies. Gennadios et al. (1994) suggested WVP corrections to account for resistance of the air layer underneath tested hydrophilic film specimens.

Studies on heat treatment of film forming solutions for whey protein films (McHugh et al., 1994), as well as heat treatment of dried soy protein films (Gennadios et al., 1996) have shown that heating at elevated temperatures increased the film strength and water resistant properties. Heat-treated protein film had increased stress and decreased strain, water solubility and water vapor permeability (Gennadios et al., 1996).

The objective of this study was to determine the effects of thermal treatments of wheat gluten films on water vapor permeability (WVP), Hunter L, u and b values, tensile strength (TS) and elongation at break (E) and to determine the trends of these properties following different temperature and exposure time combinations.

2. Materials and methods

2.1. Materials

Whetpro- wheat gluten with an approximate 80% (db) protein content was provided by Ogilvie Mills Ltd., Quebec, Canada. Glycerin (ACS grade) and ammonium hydroxide were procured from Fisher Scientific, Pittsburgh, PA. Ethanol (200 proof) was obtained from Baxter Diagnos- tics, Inc., McGraw Park, IL. Ethanol was diluted to 95% by adding 5% (v/v) distilled water.

2.2. Film preparation

A film-forming solution was prepared by mix- ing 10 g of wheat gluten in a premixed solution of 48 ml of 95% ethanol and 3.36 g glycerin. Glyc- erin was added as a plasticizer. The solution was warmed and stirred on a magnetic stirrer/hot plate, and 32 ml of distilled water and 8 ml of 5 M ammonium hydroxide were simultaneously added. The mixture was heated on the hot plate until the gluten dispersed. At this point (75 + 2°C) the viscosity of the solution decreased significantly. Once this stage was reached, the film- forming solution was filtered through cheese cloth to remove any lumps of wheat gluten.

The filtered film-forming solution, while still warm, was poured onto a clean, flat glass surface,

Y. Ali et al. /Industrial Crops and Products 6 (1997) 177-184 179

preheated to 80°C. A thin-layer chromatography spreader bar (Brinkman Co., NY) was used to cast the film. Gennadios et al. (1993a) observed that this film casting technique provided a limited degree of monoxial molecular orientation parallel to the direction in which the spreader was moved as opposed to the random orientation of molecules occurring upon pouring and spreading the solution on the substrate surface. Molecular orientation is generally desirable in film produc- tion since it improves barrier properties (Pascat, 1986).

Cast films on glass plates were dried overnight at room temperature and were peeled from the glass surface. Films were then conditioned for 48 h at a relative humidity (RH) of 50 f 5% and a temperature of 25 f 2°C (ASTM D 618-61, ASTM, 1989) in an environmental chamber (Model 3 17322, Hotpack Corp., Philadelphia, PA).

2.3. Heat treatment

The conditioned films were mounted on glass plates by applying masking tape around the film edges and heated at 65, 80 or 95°C in a forced convection oven for 2, 4, 6, 12, 18 or 24 h. Masking tape was used to prevent curling and rippling of films during heating. More severe heat treatments were not used to avoid evaporative losses of glycerin. The boiling point of pure glycerin is 290°C and drops substantially in the pres- ence of water (Gregory, 1991). Following heat treatments, the films were again conditioned in the environmental chamber for 48 h at 50 f 5% RH and 25 rf 2°C before film properties were evaluated.

2.4. Water vapor permeability

Water vapor permeability (WVP) of all heat- treated and conditioned wheat gluten films was determined by the method described by Genna- dios et al. (1994). Film specimens (7 x 7 cm) were cut and thickness measurements of each film were taken using a hand held micrometer (B.C. Ames Co., Waltham, MA) to the nearest 2.54 pm (0.1 ml). Five thickness measurements were taken on

each film sample, one at the center and four around the perimeter, and the mean was used in WVP calculations.

To determine WVP of films, water vapor transmission rate (WVTR) was determined gravimetri- tally using a modified ASTM method E 96-80 (ASTM, 1989). Film specimens were mounted on poly(methy1 methacrylate) cups (Gennadios et al., 1994) filled with 18 ml of distilled water. The cups were placed in an environmental chamber at 25°C and 50% RH. A fan was operated within the chamber to provide an air velocity of 198 m/min over the surface of the cups for removing the water vapor. Weights of the cups were recorded first after 2 h and then five more times at 1 h intervals. Regression analysis of weight loss as a function of time was performed to insure that accurate steady state slopes were obtained. Re- gression coefficients were greater than 0.998 at Pr > F < 0.001. WVTR was determined as

WVTR = Slope/Film Area (1)

Where slope, weight loss per unit time, g/h.Using WVTR values from Eq. (l), WVP of the films were calculated as McHugh et al. 1993

WVP = (WVTR x L)/Ap (2)

where L, mean film thickness, mm and Ap, partial water vapor pressure difference (Pa) across the two sides of the film.

Because of the low water vapor resistance of protein-based films, the actual RH values at the film undersides during testing were lower than 100%. Actual RH values at the film undersides and film WVP values were corrected after ac- counting for the resistance of the stagnant air layer between the film undersides and the water surface in the cups (McHugh et al., 1993; Genna- dios et al., 1994). The mean of the initial and final stagnant air gap heights was used in the calculations.

2.5. Color coordinates

Color coordinates of all wheat gluten film samples, treated with different time and temperature combinations, were measured with a CR-300 Mi- nolta Chroma Meter (Minolta Camera Co., Ltd.,


Osaka, Japan). This instrument is a tristimulus color analyzer with an 8 mm diameter measur- ing area. The instrument was first calibrated with a white standard plate (Calibration Plate CR-A43). Film samples were placed on the white plate and Hunter L, a, and b color values were measured. The three color coordinates ranged from L = 0 (black) to L = 100 (white), - a (greenness) to + a (redness), and - b (blue- ness) to + b (yellowness) (Francis and Clydes- dale, 1975). Total color difference (A\E) was defined as

+ (bfi, - LndarcJ210.5 (3)

Standard values for the white calibration plate were L = 96.86, a = - 0.02, and b = 1.99. Mea- surements were taken at five places on each film sample and mean values were used. Color values measurement test was replicated three times.

2.6. Tensile strength and elongation at break

Film tensile strength (TS) and elongation at break (E) were determined using an Instron Universal Testing Machine, Model 5566 (Instron Corp., Canton, MA). ASTM test procedure D 882-88 (ASTM, 1989) was used. Film samples were conditioned as described in section 2.2. Film samples were cut into strips 2.54 cm wide and 15 cm long. Five thickness measurements were taken along the length of each sample with a hand held micrometer (B.C. Ames Co., Waltham, MA) and the mean was used in TS calculation. The initial grip separation and cross- head speed of the Instron were set at 10 cm and 5 cm/min, respectively.

Peak loads and elongation at break point were recorded. TS was calculated by dividing peak load by initial cross sectional area of the film. E was calculated by the dividing total elongation of film at the point of rupture by initial film length (10 cm). Elongation of film at break was reported in percentage. Both TS and E values for each temperature and exposure time combinations were replicated six times.

2.7. Experimental design and data analysis

The experimental design used in evaluating the effects of heat treatment on the properties of film was a randomized complete block split plot design with three temperatures as the main plot treatments, seven exposure times as the split plot treatments and two replications as blocks. There were two types of experimental units: (1) main plot units associated with each temperature in each block and (2) split plot units associated with exposure time within one temperature in each block. The means of each response variable for each split plot unit were used for statistical analysis.

Statistical analysis (SAS, 1989) were performed in three steps using analysis of variance (ANOVA) and regression. First, an ANOVA model was fitted on the split plot within three temperatures as main plot treatment and seven exposure times as subplot treatments, to estimate main plot and split plot mean square errors. In the second step, the control sample with no treatment was dropped out of analysis and the main effects (temperature and exposure time) and interactions were broken into one degree of freedom polynomial contrasts. The mean square for each polynomial contrast was compared with the respective mean square error determined in step 1, to find the significant terms. Significance was evaluated at a 95% confidence level for all statistical analyses. In the third step, significant contrasts from the second step were used to identify polynomial terms that were fitted in a regression model (Milliken and Johnson, 1984). The estimated regression model was then used to generate predicted points for measured properties on a response surface, which was plotted against exposure time and temperature.

3. Results and discussion

3.1. Water vapor permeability

The mean water vapor permeability @VVP) values along with actual RH test conditions for all wheat gluten films treated at different temperature


and exposure time combinations are presented in Table 1 and the trends in Fig. 1. No interaction was observed between temperature and exposure time on WVP. The regression analysis indicated significant linear effects of temperature and exposure time on the WVP of the films. At a given temperature, the WVP decreased as the exposure time increased. The control film sample, with no thermal treatment had a WVP of 1.93 x 10 - ’ g/m per s per Pa, which reduced to 1.91 x 10 -9, 1.77 x 10e9 and 1.62 x lop9 g/m per s per Pa after treating the film for 2 h at temperatures of 65, 80 and 95”C, respectively. It further decreased to 1.60 x 10e9, 1.46 x lop9 and 1.32 x 10e9 g/m per s per Pa at the same temperatures, respectively, after 24 h exposure time. Similarly, at a given exposure time, the WVP decreased as the temperature increased. As heating temperature and exposure time increased, the WVP of the films was reduced.

The decrease in WVP of the heat treated films may be attributed to the formation of covalent links within the films during heat treatment and to a decrease in hydrophilicity of the films. Genna- dios et al. (1996) observed similar trends for soy protein films heat-treated at 80 or 95°C for 3, 6, 14 or 24 h.

3.2. Color coordinates

The Hunter L, a, and b color values of all heat-treated films and the total color difference

2.5

p” 3 E 3I

2

m- ‘0 x 1.5

1 1. I a, I. I

0 5 10 15 20 25

Treatment Time, h

Fig. 1. Effects of treatment time and temperature on water vapor permeability of wheat gluten films, RZ = 0.742. WVP = 2.56 - 9.56 x 10-3t-0.014h, where t = temp (“C) and h =

time (h).

50’ I 0 5 10 15 20 25

Treatment Time, h

Fig. 2. Effects of treatment time and temperature on Hunter L value of wheat gluten films, R2 = 0.964. L = 113.03 - 0.062t +

3.93 x 10-3t2 - 11.13h + 0.52h2 + 0.32th - 2.1 x 10-3t2h

- O.O14th* + 9.1 x 10V5t2h2, where t = temp (“C) and h =

time (h).

(AE) values were determined. The mean values are presented in Table 2 and their trends also are given in Figs. 2-4.

Statistical analysis conducted on effects of treatment temperature and exposure time on Hunter L value of the films indicated a significant quadratic interaction between temperature and exposure time. Exposure time and treatment temperature also had a significant quadratic main effect on the L value. At 65”C, the exposure time did not affect the film color markedly, but at higher temperatures the L values of the film were reduced significantly (Fig. 2).

-2 ’ 0 5 10 15 20

Treatment Time, h

Fig. 3. Effects of treatment time and temperature on Hunter a

value of wheat gluten films, R2 = 0.992. a = 15.51 - 0.43t +

2.72 x 10-3t2 + 2.03h - O.llh* - 0.06th + 3.91 x lo-“t’h

+ 0.003th2- 1.88 x lo- ‘t*h*, where t = temp (“C) and h =

time (h).

Tab

le

1 E

ffec

t of

hea

ting

whe

at

glut

en

film

s (1

14

+ 10

pm

th

ick)

at

di

ffer

ent

tem

pera

ture

s an

d ex

posu

re

times

on

w

ater

va

por

perm

eabi

lity

at

25°C

an

d 50

%

RH

en

viro

nmen

ta

Hea

ting

time

(h)

65°C

80

°C

95°C

WV

P (x

lo

-’

g/m

pe

r s

per

Pa)

0 1.

93

( &

0.0

8)

2 2.

00

( *

0.13

)

4 1.

97

(kO

.13)

6 1.

69

( f

0.01

)

12

1.66

(

f 0.

01)

18

1.61

(+

0.

01)

24

1.61

(k

O.0

1)

RH

in

side

cu

p W

VP

(x

IO-’

g/

m

per

s

(“/)

pe

r Pa

)

RH

in

side

cu

p

(“J)

WV

P (x

lo

-’

g/m

pe

r s

per

Pa)

RH

in

side

cu

p

(“!)

-

77.9

0 (

+ 1.

36)

76.5

8 (

f 1.

84)

76.1

8 (f

1.

52)

75.7

1 (

f 0.

74)

76.2

0 (k

2.

71)

79.2

0 (

f 1.

49)

75.6

0 (+

3.

59)

1.93

(

+ 0.

01)

1.90

(

f 0.

01)

1.65

( f

0.05

)

1.74

(

* 0.

09)

1.66

(

* 0.

07)

1.72

( & 0

.08)

1.54

(k

0.

16)

77.9

0 (

f 0.

92)

74.6

5 (f

3.

61)

74.0

1 (

& 1

.42)

77.1

0 (

f 0.

68)

77.7

4 (+

0.

94)

77.0

9 (

k 2.

69)

79.3

2 (k

0.

17)

1.93

(

f 0.

08)

1.65

(+

0.

12)

1.56

(k

0.

12)

1.43

(

+ 0.

09)

1.53

(

f 0.

49)

1.43

( f

0.38

)

1.22

(

f 0.

04)

77.9

0 (

* 0.

68)

79.6

8 (

k 1.

40)

80.5

2 (

f 0.

37)

80.2

3 (

k 0.

30)

79.1

1 (

+ 2.

95)

79.9

7 (

* 1.

55)

81.1

5 (k

0.

39)

a Sp

ecif

ic

regr

essi

on

mod

el

on

a re

spon

se

surf

ace

for

wat

er

vapo

r pe

rmea

bilit

y m

eans

ar

e pr

esen

ted

in

Fig.

1.

Tab

le

2

Ave

rage

H

un

ter

colo

r co

ordi

nat

es

(L,

0,

and

b)

and

tota

l co

lor

diff

eren

ce

(AE

) fo

r w

hea

t gl

ute

n f

ilm

s tr

eate

d at

dif

fere

nt

tem

pera

ture

s an

d ex

posu

re

tim

es=

.

Hea

tin

g ti

me

(h)

65°C

80

°C

95°C

L

a b

AE

L

n

b

AE

L

(1

b

Aa5

0

88.8

0 (+

0.

59)

-0.9

4 (

f 0.

06)

9.56

( f

0.

87)

11.2

6 88

.80

( f

0.59

) -0

.94

( f

0.06

) 9.

56 (

+

0.87

) II

.26

88.8

0 (

f 0.

59)

-0.9

4 (

+ 0

.06)

9.

56

( f

0.87

) 11

.26

2 89

.56

( f

0.35

) -0

.92

( +

0.0

5)

8.90

(k

0.67

) 10

.09

89.0

7 (+

0.

32)

- 1.

50 (

+ 0

.07)

12

.89

( f

0.63

) 13

.52

89.2

3 (k

O.2

1)

-0.3

9 (

f 0.

06)

8.28

( +

0.4

5)

9.90

4 89

.41

( f

0.33

) -1

.03

(kO

.10)

9.

61 (

k 0.

74)

10.7

0 89

.77

( f

0.33

) -

1.37

( ?

r 0.

02)

11.9

1 (k

O.9

1)

12.2

7 88

.24

(+

0.46

) -0

.29

( f

0.08

) 12

.67

(f

1.53

) 13

.73

6 89

.48

(f

0.14

) -0

.99

( f

0.00

) 8.

25 (

+

0.97

) 9.

73

89.4

0 (

f 0.

62)

- 1.

40 (

+ 0

.02)

13

.51

( f

0.45

) 13

.79

85.9

7 (+

0.

76)

0.21

( f

0.

24)

19.7

5 (

f 2.

00)

20.8

3

12

89.8

2 (

f 0.

26)

- 1 .

Ol

( f

0.03

) 9.

45 (

k 0.

47)

10.3

1 88

.90

( f

0.44

) -

1.29

( f

0.

07)

16.1

7 (k

O.7

4)

16.3

1 84

.22

(k

0.14

) 0.

59 (

kO

.12)

24

.97

(k

2.22

) 26

.24

18

89.1

0 (f

0.

49)

- 1.

20 (

f

0.07

) 11

.33

(f

1.05

) 12

.20

88.1

3 (f

0.79

) -1

.09

(+o.

Iz)

17.2

5 (

f 1.

52)

17.6

1 84

.91

( f

0.53

) 0.

43 (

f

0.02

) 23

.93

( f

1.08

) 24

.99

24

88.9

0 (+

0.

97)

-1.2

8 (f

0.12

) 12

.54

(k

2.27

) 13

.27

84.8

4 (

f 2.

14)

-0.4

0 (

f 0.

60)

26.6

3 (

+ 4

.93)

27

.42

83.8

2 (

f 1.

82)

0.71

(k

O.1

8)

27.0

7 (+

2.

91)

28.2

8

a S

peci

fic

regr

essi

on

mod

el

on

a re

spon

se

surf

ace

for

L,

a,

and

b av

erag

es

are

pres

ente

d in

Fig

s.

2-4


5

I

“0 5 10 15 2-J 25

Treatment Time, h

Fig. 4. Effects of treatment time and temperature on Hunter b

value of wheat gluten films, R* = 0.987. b = 123.22 - 3.71t - 0.03t2 + 22.39h - 0.62h2 - 0.0h3 - 0.72th + 5.62 x lo-‘t’h + 0.03th2 - 2.36 x lo-“t*h* + 2.48 x 10V4th3 , where t = temp (“C) and h = time (h).

Hunter a value also had a significant quadratic interaction between temperature and exposure time (Fig. 3). Both temperature and time were significant quadratic main effects. At lower temperature (65°C) the film color was more toward the green end of the ‘a’ scale and further decreased as the exposure time increased. However, at higher temperatures (80 and 95”(Z), the Hunter a value changed from green to red as the exposure time increased.

A significant interaction between temperature (quadratic) and exposure time (cubic) was indicated for film b values. There was a quadratic main effect of temperature and cubic main effect of heat treatment. As the exposure time increased the yellowness of the films increased for all temperatures (Fig. 4).

As temperature and exposure time increased, the whiteness decreased and redness and yellowness increased (Table 2, Figs. 2 and 4). The com- bined effects of Hunter L, a, and b values resulted in increased total color difference (AE). Genna- dios et al. (1996) reported that film color also was affected by the moisture content and thickness of the films.

3.3. Tensile strength and elongation at break

The effect of temperature and exposure time on the tensile strength (TS) and elongation at break (E) of wheat gluten films are presented in Figs. 5

5 10 15 20

Treatment Time, h

Fig. 5. Effects of treatment time and temperature on tensile strength of wheat gluten films, R* = 0.961. TS= - 1.86 + 0.072t - 0.15h - 0.05h2 + 1.06 x 10W3h3 + O.O12th, where t = temp (“C) and h = time (h).

and 6, respectively. There existed a significant interaction between temperature and exposure time on TS. Temperature had a linear main effect, whereas exposure time had a cubic main effect on TS. Untreated (control) film had a mean TS of 3.78 MPA, which increased significantly with increases in temperature and exposure time (Fig. 5).

The effects of temperature and exposure time on E, a measure of a film’s ability to stretch, were inverse to that of TS. An interaction existed between temperature (quadratic) and exposure time (linear). Both temperature and exposure time had a quadratic main effects on E. The untreated film sample had a mean E value of 187.5% which decreased at a very rapid rate with increases in temperature and exposure time. Temperature and exposure times tended to reduce E (Fig. 6). Gen-

200 .

8

5 150

t m

iii 100

s

% z 50

P w

0 1 0

Treatment Time, h

Fig. 6. Effects of treatment time and temperature on elongation at break of wheat gluten films, R2 = 0 977. E = 475.58 - 6.27t + 0.03tZ + 44.57h +0.13/r* - 1.19th + 6.78 x lo-%*h,

where t = temp (“C) and h = time (h).


nadios et al. (1996) reported an increase in TS and decrease in E with temperature and time for wheat gluten and soy protein films, respectively. However, TS and E for soy protein films were markedly less than those for wheat gluten films. They suggested that the effects of heating on film TS and E were partially attributed to the development of heat-induced cross linkages within the film structure.

4. Conclusions

Tensile, water vapor barrier and color properties of wheat gluten films were affected by thermal treatment generated by combinations of temperatures and exposure times. Water vapor permeability was reduced with increases in treatment temperature as well as with increases in exposure time at a particular temperature. The Hunter L, a, and b color coordinates also were affected. An increase in tensile strength with increasing temperature and exposure time was observed, but this was at the cost of reduced elongation properties of the films.

References

ASTM, 1989. Annual Book of ASTM Standards. American Society for Testing Materials, Philadelphia, PA.

Banker, G.S., Gore, A.Y. and Swarbrick, J., 1966. Water vapor transmission properties of free polymer films. J. Pharm. Pharmac., 18: 457-466.

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