**3.3 Lipid profile**

The lipid profile of wheat flour and soybean flour obtained by HPLC analysis and based on these results the lipid profile of wheat-soybean flour mixtures, is presented in Table 3. The content of components was determined by measuring the peak area at 1.76 min for free fatty acids, peak area at 2.15 min for methyl esters, peaks area in the range of 3.44-4.58 min for

The Main Components Content,

content was 3.7.

**3.4 Statistical analysis data** 

Rheology Properties and Lipid Profile of Wheat-Soybean Flour 89

The lipid from soybean flour had higher free fatty acids content (25.9 g/100g lipid) and triacylglycerols content (69.4 g/100g lipid) than lipid from wheat flour (11.9 and 50.2 g/100g lipid, respectively), while the contents of methyl esters, monoacylglycerols and diacylglycerols were lower than appropriate contents in wheat flour, even 14.5, 3.1 and 5.3 times, respectively. The content of free fatty acids and triacylglycerols in mixtures had the same changing tendency as soybean flour portion: it increased when the soybean flour portion in flour mixtures increased, while the contents of monoacylglycerols and diacylglycerols had opposite changing tendency: they decreased when the soybean flour portion in flour mixture increased. The properties of dough such as water absorption, development time, dough stability and gelatinization temperature had the same dependency on soybean flour portion as the content of free fatty acids and triacylglycerols and these components seemed to have a proper influence on the mentioned dough properties. Further, the content of monoacylglycerols and diacylglycerols had a proper influence on dough energy, resistance, extensibility and gelatization maximum. Some of

these dependences were confirmed by determination of correlations coefficients.

The fatty acids composition of flours obtained by GC analysis and based on these results, the fatty acids composition in flour mixtures is presented in Table 4. The lipid contained: linoleic (C18:2), -linolenic (C18:3), oleic (C18:1), palmitic (C16:0), stearic (C18:0), arachidonic (C20:4), behenic (C22:0), nonadecanoic (C19:0), -linolenic, lignocerinic, myristic (C14:0) and several non-determined components (ND). GC analysis showed the lipid from wheat flour contained 78.14 g/100g of the total unsaturated fatty acids, consisting of linoleic (57.91 g/100g) and oleic acid (20.23 g/100g). The total polyunsaturated fatty acids content in lipid was 57.91 g/100g and it was from linoleic acid. The monounsaturated fatty acids content was 20.23 g/100g, from oleic acid, while the total saturated fatty acids content was 21.07 g/100g where the main fatty acids were palmitic and stearic acid with the content of 19.45 and 1.36 g/100g, respectively. The ratio of total unsaturated to total saturated fatty acids

The lipid from soybean flour contained 81.46 g/100g of the total unsaturated fatty acids, composed of linoleic, oleic and linolenic acid with content of 55.23, 13.07 and 13.46 g/100g of lipid, respectively. The palmitic and stearic acid were the main saturated fatty acids with content of 10.35 and 5.23 g/100g, respectively, while the content of other detected saturated fatty acid was 0.69 g/100g. The myristic, linolenic, nonadecanoic, arachidic and lignocerinic fatty acids were detected only in soybean flour, while phthalic acid was detected only in wheat flour. Based on this composition and content of fatty acids, the ratio of total unsaturated to total saturated content in soybean flour was 4.58 and it was higher than ratio in wheat flour. Such fatty acids composition had influence on fatty acids composition in wheat-soybean flour mixtures: the content of total saturated fatty acids decreased when soybean flour portion in flour mixture increased, and all values were lower than the content of total saturated fatty acids in wheat flour; the content of total unsaturated fatty acids increased when soybean flour portion in flour mixture increased and all values (except in mixture with portion of soybean flour of 3%) were higher than the content of total saturated fatty acids in wheat flour. Finally, the ratio of total unsaturated to total saturated fatty acids content was higher in flour mixtures

The correlation coefficients between the rheological properties (water absorption, dough stability, Ex, gelatinization temperature and gelatization maximum) and the content of some lipid components (free fatty acids, monoacylglycerols, diacylglycerols, triacylglycerols,

(except in mixture with portion of soybean flour of 3%) than in wheat flour.


monoacylglycerols, peaks area in the range of 5.28-8.68 min for diacylglycerols and peaks area in the range of 10.91-15.81 min for triacylglycerols (Holčapek et al., 1999).

Table 3. The lipid profile of wheat and soybean flour and their mixtures obtained by HPLC


Table 4. Fatty acids composition of wheat and soybean flour and their mixtures obtained by GC

monoacylglycerols, peaks area in the range of 5.28-8.68 min for diacylglycerols and peaks

Wheat 11.90.4 23.21.3 2.50.2 12.20.9 50.21.5 Soybean 25.91.2 1.60.3 0.80.1 2.30.4 69.41.6 3% 12.3 22.5 2.4 11.9 50.8 5 % 12.6 22.1 2.4 11.7 51.2 10 % 13.3 21.0 2.3 11.2 52.1 20 % 14.7 18.9 2.2 10.2 54.0 30 % 16.1 16.7 2.0 9.2 55.9 Table 3. The lipid profile of wheat and soybean flour and their mixtures obtained by HPLC

MAG (g/100g)

5

3

1

2

6

9

TU 78.14 81.46 77.43 78.31 78.49 78.85 79.23 TMU 20.23 13.07 19.20 19.87 19.52 18.79 18.08 TPU 57.91 68.39 58.23 58.44 58.97 60.06 61.15 TS 21.07 17.84 20.92 20.82 20.58 20.44 20.13 TU/TS 3.71 4.58 3.70 3.76 3.81 3.86 3.94

Table 4. Fatty acids composition of wheat and soybean flour and their mixtures obtained by

Palmitic (C16:0) 19.450.45 10.350.6 19.17 18.99 18.54 17.63 16.71 Linoleic (C18:2) 57.910.72 55.231.2 57.83 57.77 57.63 57.37 57.11 Oleic (C18:1) 20.230.21 13.070.8 19.20 19.87 19.52 18.79 18.08 Linolenic (C18:3) 0 13.460.9 0.40 0.67 1.34 2.69 4.04 Stearic (C16:0) 1.360.14 5.230.8 1.48 1.56 1.76 2.18 2.58

Behenic (C22:0) 0.260.06 0.480.06 0.27 0.27 0.28 0.30 0.33

DAG (g/100g)

flour 3% 5% 10% 20% 30%

0.03 5

0.05

0.06

0.05 2

0.05

0.06

0.03 5

<sup>3</sup>0.10 0.16

<sup>1</sup>0.12 0.18

0.05 2

<sup>6</sup>0.11 0.17

<sup>9</sup>0.14 0.20

0.03 5

0.05 2

0.03 5

0.05 3

0.06 1

0.05 2

0.05 6

0.06 9

TAG (g/100g)

area in the range of 10.91-15.81 min for triacylglycerols (Holčapek et al., 1999).

ME (g/100g)

Soybean

Flour FFA

**Fatty Acid content in g/100g**

Lignocerinic

GC

(g/100g)

Wheat flour

Myristic (C14:0) 0 0.110.01 0.03

Nonadecanoic (C19:0) 0 0.520.09 0.05

Arachidic (C20:0) 0 0.600.1 0.06

ND RT 25.96 0 0.170.04 0.05

Phthalic acid 0.680.08 0 0.06

(C24:0) 0 0.550.01 0.05

The lipid from soybean flour had higher free fatty acids content (25.9 g/100g lipid) and triacylglycerols content (69.4 g/100g lipid) than lipid from wheat flour (11.9 and 50.2 g/100g lipid, respectively), while the contents of methyl esters, monoacylglycerols and diacylglycerols were lower than appropriate contents in wheat flour, even 14.5, 3.1 and 5.3 times, respectively. The content of free fatty acids and triacylglycerols in mixtures had the same changing tendency as soybean flour portion: it increased when the soybean flour portion in flour mixtures increased, while the contents of monoacylglycerols and diacylglycerols had opposite changing tendency: they decreased when the soybean flour portion in flour mixture increased. The properties of dough such as water absorption, development time, dough stability and gelatinization temperature had the same dependency on soybean flour portion as the content of free fatty acids and triacylglycerols and these components seemed to have a proper influence on the mentioned dough properties. Further, the content of monoacylglycerols and diacylglycerols had a proper influence on dough energy, resistance, extensibility and gelatization maximum. Some of these dependences were confirmed by determination of correlations coefficients.

The fatty acids composition of flours obtained by GC analysis and based on these results, the fatty acids composition in flour mixtures is presented in Table 4. The lipid contained: linoleic (C18:2), -linolenic (C18:3), oleic (C18:1), palmitic (C16:0), stearic (C18:0), arachidonic (C20:4), behenic (C22:0), nonadecanoic (C19:0), -linolenic, lignocerinic, myristic (C14:0) and several non-determined components (ND). GC analysis showed the lipid from wheat flour contained 78.14 g/100g of the total unsaturated fatty acids, consisting of linoleic (57.91 g/100g) and oleic acid (20.23 g/100g). The total polyunsaturated fatty acids content in lipid was 57.91 g/100g and it was from linoleic acid. The monounsaturated fatty acids content was 20.23 g/100g, from oleic acid, while the total saturated fatty acids content was 21.07 g/100g where the main fatty acids were palmitic and stearic acid with the content of 19.45 and 1.36 g/100g, respectively. The ratio of total unsaturated to total saturated fatty acids content was 3.7.

The lipid from soybean flour contained 81.46 g/100g of the total unsaturated fatty acids, composed of linoleic, oleic and linolenic acid with content of 55.23, 13.07 and 13.46 g/100g of lipid, respectively. The palmitic and stearic acid were the main saturated fatty acids with content of 10.35 and 5.23 g/100g, respectively, while the content of other detected saturated fatty acid was 0.69 g/100g. The myristic, linolenic, nonadecanoic, arachidic and lignocerinic fatty acids were detected only in soybean flour, while phthalic acid was detected only in wheat flour. Based on this composition and content of fatty acids, the ratio of total unsaturated to total saturated content in soybean flour was 4.58 and it was higher than ratio in wheat flour. Such fatty acids composition had influence on fatty acids composition in wheat-soybean flour mixtures: the content of total saturated fatty acids decreased when soybean flour portion in flour mixture increased, and all values were lower than the content of total saturated fatty acids in wheat flour; the content of total unsaturated fatty acids increased when soybean flour portion in flour mixture increased and all values (except in mixture with portion of soybean flour of 3%) were higher than the content of total saturated fatty acids in wheat flour. Finally, the ratio of total unsaturated to total saturated fatty acids content was higher in flour mixtures (except in mixture with portion of soybean flour of 3%) than in wheat flour.

#### **3.4 Statistical analysis data**

The correlation coefficients between the rheological properties (water absorption, dough stability, Ex, gelatinization temperature and gelatization maximum) and the content of some lipid components (free fatty acids, monoacylglycerols, diacylglycerols, triacylglycerols,

The Main Components Content,

Linkage Distance

**4. Conclusion** 

and presented by dendrogram in Fig. 1.

Rheology Properties and Lipid Profile of Wheat-Soybean Flour 91

triacylglycerols, palmitic, linolenic and oleic acid content. Linkage distances were obtained

Tree Diagram for 6 Variables Single Linkage Euclidean distances

WS30 WSF20 WS10 WSF5 WSF3 WF

The cluster analysis based on flour rheological and lipid characteristics, shows the linkage distance between wheat flour and flour mixtures increases when soybean flour portion in mixtures increases. The mixtures with soybean flour portion of 3 and 5% (w/w) are joined with wheat flour at the same distance level of 1.6 and make the first group. The mixtures with soybean flour portion of 10 % (w/w) are joined with wheat flour at the distance level of 1.7 and could be added to the first group. The mixture with soybean flour portion of 20 and 30% (w/w) is joined with wheat flour at distance level of 8.6 and make the second group. This means that soybean flour portion of 30% could be used to more enrich dough with soybean protein and the main rheological properties remain satisfactory as at portion of 20%. This provides a possibility of soybean flour being included in portions even higher than 30%, so future work could include this investigations as well as investigations to examine what happens with lipid components during dough mixing and baking: does their

Fig. 1. Dendrogram obtained for wheat flour and wheat-soybean flour mixtures

content and composition stay the same as in flour mixture or there occur changes.

The soybean flour addition increased the protein content up to 20.0%, the ash content up to 2.08% and lipid content up to 7.2%, while decreased starch, gluten and carbohydrates content for 19.8, 7.2 and 16 g/100 g flour mixture, respectively. Dough rheological properties and lipid profile depend on soybean flour portion. The soybean flour addition had positive influence on the quality number and group and extended duration of dough stability. The dough water absorption and the degree of softening increased with increasing soybean flour portion. The dough with soybean flour portion of 20 and 30% had lower values for energy in comparison to dough made of wheat flour only and it could be economically important. Values of gelatinization temperature for dough with soybean flour were higher than dough with wheat flour, maybe due to a specific behaviour of soy starch in


palmitic (Pal), linoleic (Lin) and oleic (Ole) acid) in wheat flour and wheat-soybean flour mixtures are presented in Table 5.

Table 5. Correlation coefficients between rheological properties and lipid components content (correlations are significant at p 0.05, N=12)

The sample size was twelve (N=12 (6x2): wheat flour and five wheat-soybean flour mixtures with minimal and maximal obtained value). As there were many correlations, only the one which had absolute value 0.85 and above 0.85 were taken into consideration. There were 54.5% of correlations, among which 30.3% were proper, and 24.2% were opposite correlations. The correlations can be divided into three groups: correlations between rheological properties, between rheological properties and lipid components content and between lipid components content. In the first group, there are the correlations where high water absorption value is associated with high dough stability and gelatinization temperature and low extensibility, high dough stability is associated with high gelatinization temperature and low gelatization maximum, and correlation between low gelatization maximum and high extensibility. In the second group of correlations, the high content of free fatty acids caused high water absorption, dough stability, and gelatinization temperature but low extensibility as well as the high monoacylglycerols and diacylglycerols content was proper correlated with extensibility. Also, when triacylglycerols content was higher, the water absorption, dough stability and gelatinization temperature were higher and extensibility was lower. The palmitic and oleic acid had opposite effect on the rheological properties, such as water absorption, dough stability and gelatinization temperature and proper effect on extensibility. The linolenic acid content was not associated with any rheological properties. Among lipid components content, there were proper correlations between free fatty acids and triacylglycerols content, between monoacylglycerols and diacylglycerols contents on one side and palmitic, linoleic and oleic acid content on the other side. It can mean that monoacylglycerols and diacylglycerols are mainly consists of palmitic, linoleic and oleic acids. The higher oleic acid content was associated with the higher palmitic acid content and it was only correlation among fatty acids.

By cluster analysis, based on multiple variables, wheat and wheat-soybean flour mixtures were classified into groups. Number of variables was six: wheat and five wheat-soybean flour mixtures (3, 5, 10, 20 and 30% w/w of soybean flour portions); number of cases i.e. parameters were eight: water absorption, dough stability, free fatty acids, diacylglycerols,

palmitic (Pal), linoleic (Lin) and oleic (Ole) acid) in wheat flour and wheat-soybean flour

WA DSt Ex Tmax max FFA MAG DAG TAG Pal Lin

mixtures are presented in Table 5.

DSt **0.88** 

acids.

Ex **-0.94** -0.84

Tmax **0.91 0.98 -0.90**  max -0.64 **-0.86** 0.67 -0.78

FFA **0.96 0.96 -0.93 0.98** -0.77

MAG -0.75 -0.78 **0.91** -0.83 0.71 -0.83

content (correlations are significant at p 0.05, N=12)

DAG -0.81 -0.81 **0.94 -0.86** 0.73 **-0.86 -0.97** 

TAG **0.94 0.94 -0.85 0.94** -0.72 **0.97** -0.69 -0.72

Pal **-0.89 -0.89 0.97 -0.93** 0.78 **-0.94 0.95 0.98** -0.84

Lin -0.45 -0.47 0.71 -0.53 0.51 -0.51 **0.85 0.88** -0.30 0.77 Ole **-0.85 -0.85 0.90 -0.85** 0.78 **-0.88 0.88** 0.84 -0.81 **0.92** 0.65

Table 5. Correlation coefficients between rheological properties and lipid components

The sample size was twelve (N=12 (6x2): wheat flour and five wheat-soybean flour mixtures with minimal and maximal obtained value). As there were many correlations, only the one which had absolute value 0.85 and above 0.85 were taken into consideration. There were 54.5% of correlations, among which 30.3% were proper, and 24.2% were opposite correlations. The correlations can be divided into three groups: correlations between rheological properties, between rheological properties and lipid components content and between lipid components content. In the first group, there are the correlations where high water absorption value is associated with high dough stability and gelatinization temperature and low extensibility, high dough stability is associated with high gelatinization temperature and low gelatization maximum, and correlation between low gelatization maximum and high extensibility. In the second group of correlations, the high content of free fatty acids caused high water absorption, dough stability, and gelatinization temperature but low extensibility as well as the high monoacylglycerols and diacylglycerols content was proper correlated with extensibility. Also, when triacylglycerols content was higher, the water absorption, dough stability and gelatinization temperature were higher and extensibility was lower. The palmitic and oleic acid had opposite effect on the rheological properties, such as water absorption, dough stability and gelatinization temperature and proper effect on extensibility. The linolenic acid content was not associated with any rheological properties. Among lipid components content, there were proper correlations between free fatty acids and triacylglycerols content, between monoacylglycerols and diacylglycerols contents on one side and palmitic, linoleic and oleic acid content on the other side. It can mean that monoacylglycerols and diacylglycerols are mainly consists of palmitic, linoleic and oleic acids. The higher oleic acid content was associated with the higher palmitic acid content and it was only correlation among fatty

By cluster analysis, based on multiple variables, wheat and wheat-soybean flour mixtures were classified into groups. Number of variables was six: wheat and five wheat-soybean flour mixtures (3, 5, 10, 20 and 30% w/w of soybean flour portions); number of cases i.e. parameters were eight: water absorption, dough stability, free fatty acids, diacylglycerols, triacylglycerols, palmitic, linolenic and oleic acid content. Linkage distances were obtained and presented by dendrogram in Fig. 1.

Fig. 1. Dendrogram obtained for wheat flour and wheat-soybean flour mixtures

The cluster analysis based on flour rheological and lipid characteristics, shows the linkage distance between wheat flour and flour mixtures increases when soybean flour portion in mixtures increases. The mixtures with soybean flour portion of 3 and 5% (w/w) are joined with wheat flour at the same distance level of 1.6 and make the first group. The mixtures with soybean flour portion of 10 % (w/w) are joined with wheat flour at the distance level of 1.7 and could be added to the first group. The mixture with soybean flour portion of 20 and 30% (w/w) is joined with wheat flour at distance level of 8.6 and make the second group. This means that soybean flour portion of 30% could be used to more enrich dough with soybean protein and the main rheological properties remain satisfactory as at portion of 20%. This provides a possibility of soybean flour being included in portions even higher than 30%, so future work could include this investigations as well as investigations to examine what happens with lipid components during dough mixing and baking: does their content and composition stay the same as in flour mixture or there occur changes.
