**4. Bioconversions**

Lipids require modification in order to be used for special purposes or production of value added products. Lipids can be modified by hydrogenation, blending, fractionation and chemical or enzymatic reactions such as hydrolysis, direct esterification and interesterification.


**5. Synthesis of 3,4-epoxy-1-butene in liquid phase**

Two competitive methods for direct epoxidation of olefins are gas-phase oxidation with oxygen over silver catalyst and liquid-phase reactions with organic hydroperoxides or hydrogen peroxide in the presence of soluble or supported W, Mo, Ti complexes. The gas-phase epoxidation is typical for obtaining light epoxides, whereas epoxidation with peroxide compounds in liquid is applicable for a wide range of substrates containing double bonds. Both type reactions are based on interaction of olefin with electrophilic oxygen species. Under liquid-phase epoxidation, catalytically active metal complexes react with peroxides to attach the reactive oxygen as ligand which attack the double bond of olefin. Hydrogen peroxide is effective oxygen donor and has an advantage of low-temperature reaction giving environmentally benign water as a by-product [61, 62].

diluted aqueous H2

**Catalyst1 H2**

O)2 ]

OH solvent, room temperature.

CN solvent, room temperature.

TBA4[γ-SiW10O34(H2

(3 μmol)3

Catalyst, H2

O2

CN solvent, 60 °C.

CN solvent, 50 °C.

**Table 5.** Epoxidation of BD with H2

1

2 CH3

3 CH3

4 CH3

5 CH3

**O2 1 (mmol)** **BD (bar)**

As a result, the efficiency of H2

dation of olefins [67].

O2

O2

in solvents.

tungstates exhibit equally effective catalysis.

cursor of tungsten-depleted anions [PW4

demanded and valuable chemicals.

in acetonitrile solution. Epoxidation of BD has been shown to pro-

consumption for producing 3,4-epoxy-1-butene is extremely

O2

O2

], which operate as the most effective

[WO(O2 )2 ]4

O2

is minimal or

, thus increasing

consumption.

}3− in epoxi-

ceed with high selectivity for 3,4-epoxy-1-butene. Appearance of small admixtures of furan, 3-butene-1,2-diol, and 2-butene-1,4-oxygenates is associated with isomerization and hydro-

**Time (h) Epoxide (mmol)**

0.3 2.5 9 0.30 99 99 [64]

Reactivity of a Simplest Conjugated Diolefin in Liquid-Phase Oxidation: Mechanisms and Products

TS-1(6 mg)2 0.5 1.5 1 0.25 n.d. 52 [63]

TBA-PW11(10 μmol)4 0.9 1 5.5 0.51 88 88 [65] EMIm-PW11(10 μmol)4 0.9 1 5 0.65 91 90 [65] EMIm-PW11(2.3 μmol)5 1.0 1 5 0.20 97 100 [65]

and epoxide produced were normalized to 2 mL of the reaction mixture.

**Sel.BD (%) H2**

**O2**

**(%)**

 **efficiency**

http://dx.doi.org/10.5772/intechopen.71259

**Reference**

111

absent when the reaction is carried out at a low temperature and at a low concentration of hydrogen peroxide. This is favorable for maintaining high selectivity for 3,4-epoxy-1-butene, because the secondary oxidation of 3,4-epoxy-1-butene by radical intermediates is prevented. As a result, only negligible amount of acrolein appears in the product. Moreover, small addi-

high, it approaches to 100% under favorable conditions. Both Si- and P-centered heteropoly-

Under reaction conditions, the catalytically active anions are generated from starting lacunary polyoxotungstate anion. It has been shown by NMR that [HPW11O39]6− anion is a pre-

] and [PW2

activators of hydrogen peroxide and are responsible for epoxidation (**Scheme 9**) [65]. This is

Despite the limited use of 3,4-epoxy-1-butene itself, it is nevertheless a raw material for the synthesis of various C4-oxygenates such as 1,4-butanediol [68], 3-butene-1,2-diol and 2-butene-1,4-diol [69–71], and 2,5-dihydrofuran [72]. Therefore, low-temperature and selective epoxidation of BD can be considered as a principal stage of alternative synthesis of

lysis of 3,4-epoxy-1-butene. The unproductive radical decomposition of H2

tives of EMImBr have been found to inhibit radical decomposition of H2

confirmed by the high reactivity of a specially synthesized anion {PO4

O2

the selectivity of BD to 3,4-epoxy-1-butene conversion and efficiency of H2

The liquid-phase epoxidation of BD with H2 O2 is known to occur over titanium silicates in CH3 OH [63] and in CH3 CN solution of heteropoly compounds [64, 65]. The data for these reactions are given in **Table 5**.

Both catalysts are activators of hydrogen peroxide, capable of forming peroxide complexes. Thoroughly investigated for various olefins, the mechanism of epoxidation is realized for the conversion of BD to 3,4-epoxy-1-butene. Coordinated on metal ion, the electrophilic oxygen interacts with one of the equivalent double bonds of BD leaving intact the second C═C bond. Oxygen transfer from peroxide ligand to double bond of olefin has been proved using isotopic reagents [64]. The addition of oxygen to the second bond of BD is more difficult; therefore, the formation of a diepoxide is not detected in reactions with hydrogen peroxide.

Lacunary polyoxotungstates are effective catalysts for epoxidation of olefins with H2 O2 [66]. Besides olefins, [HPW11O39]6−and [γ-SiW10O34(H2 O)2 ]4− anions catalyze epoxidation of BD with

Reactivity of a Simplest Conjugated Diolefin in Liquid-Phase Oxidation: Mechanisms and Products http://dx.doi.org/10.5772/intechopen.71259 111

1 Catalyst, H2 O2 and epoxide produced were normalized to 2 mL of the reaction mixture.

2 CH3 OH solvent, room temperature.

3 CH3 CN solvent, room temperature.

4 CH3 CN solvent, 60 °C.

**5. Synthesis of 3,4-epoxy-1-butene in liquid phase**

**Catalyst, conditions Products (mmol)**

The liquid-phase epoxidation of BD with H2

Besides olefins, [HPW11O39]6−and [γ-SiW10O34(H2

OH [63] and in CH3

reactions are given in **Table 5**.

CH3

5%Pd0.5%Te/C, H2

110 Alkenes

5%Pd2.7Te/C, H2

5%Pd2.7Te/C, H2

SO4

SO4

SO4

**Table 4.** Products of BD oxidation in solvent CH3

Two competitive methods for direct epoxidation of olefins are gas-phase oxidation with oxygen over silver catalyst and liquid-phase reactions with organic hydroperoxides or hydrogen peroxide in the presence of soluble or supported W, Mo, Ti complexes. The gas-phase epoxidation is typical for obtaining light epoxides, whereas epoxidation with peroxide compounds in liquid is applicable for a wide range of substrates containing double bonds. Both type reactions are based on interaction of olefin with electrophilic oxygen species. Under liquid-phase epoxidation, catalytically active metal complexes react with peroxides to attach the reactive oxygen as ligand which attack the double bond of olefin. Hydrogen peroxide is effective oxygen donor and has an advantage of low-temperature reaction giving environmentally benign water as a by-product [61, 62].

5%Pd0.5%Te/C, 10 mmol BD, 100°C, 3 h 0.54 1.16 0.35 1.58 0.17 1.48 0 0.73

OH (10% H2

, 10 mmol BD, 100°C, 3 h 0.45 2.02 0.08 0.12 1.42 2.30 0 1.81

, 10 mmol BD, 120°C, 2 h 0.24 0.06 0 0.06 0.66 2.80 2.0 0

, 40 mmol BD, 120°C, 2 h 0.23 0.06 0 0.07 0.78 6.70 3.90 0

O) (30 mL), H2

SO4

O2

Both catalysts are activators of hydrogen peroxide, capable of forming peroxide complexes. Thoroughly investigated for various olefins, the mechanism of epoxidation is realized for the conversion of BD to 3,4-epoxy-1-butene. Coordinated on metal ion, the electrophilic oxygen interacts with one of the equivalent double bonds of BD leaving intact the second C═C bond. Oxygen transfer from peroxide ligand to double bond of olefin has been proved using isotopic reagents [64]. The addition of oxygen to the second bond of BD is more difficult; therefore,

the formation of a diepoxide is not detected in reactions with hydrogen peroxide.

Lacunary polyoxotungstates are effective catalysts for epoxidation of olefins with H2

O)2

is known to occur over titanium silicates in

(0.1 mmol where indicated).

**1 2 3 4 5 6 7 8**

]4− anions catalyze epoxidation of BD with

O2 [66].

CN solution of heteropoly compounds [64, 65]. The data for these

5 CH3 CN solvent, 50 °C.

**Table 5.** Epoxidation of BD with H2 O2 in solvents.

diluted aqueous H2 O2 in acetonitrile solution. Epoxidation of BD has been shown to proceed with high selectivity for 3,4-epoxy-1-butene. Appearance of small admixtures of furan, 3-butene-1,2-diol, and 2-butene-1,4-oxygenates is associated with isomerization and hydrolysis of 3,4-epoxy-1-butene. The unproductive radical decomposition of H2 O2 is minimal or absent when the reaction is carried out at a low temperature and at a low concentration of hydrogen peroxide. This is favorable for maintaining high selectivity for 3,4-epoxy-1-butene, because the secondary oxidation of 3,4-epoxy-1-butene by radical intermediates is prevented. As a result, only negligible amount of acrolein appears in the product. Moreover, small additives of EMImBr have been found to inhibit radical decomposition of H2 O2 , thus increasing the selectivity of BD to 3,4-epoxy-1-butene conversion and efficiency of H2 O2 consumption. As a result, the efficiency of H2 O2 consumption for producing 3,4-epoxy-1-butene is extremely high, it approaches to 100% under favorable conditions. Both Si- and P-centered heteropolytungstates exhibit equally effective catalysis.

Under reaction conditions, the catalytically active anions are generated from starting lacunary polyoxotungstate anion. It has been shown by NMR that [HPW11O39]6− anion is a precursor of tungsten-depleted anions [PW4 ] and [PW2 ], which operate as the most effective activators of hydrogen peroxide and are responsible for epoxidation (**Scheme 9**) [65]. This is confirmed by the high reactivity of a specially synthesized anion {PO4 [WO(O2 )2 ]4 }3− in epoxidation of olefins [67].

Despite the limited use of 3,4-epoxy-1-butene itself, it is nevertheless a raw material for the synthesis of various C4-oxygenates such as 1,4-butanediol [68], 3-butene-1,2-diol and 2-butene-1,4-diol [69–71], and 2,5-dihydrofuran [72]. Therefore, low-temperature and selective epoxidation of BD can be considered as a principal stage of alternative synthesis of demanded and valuable chemicals.

**Acknowledgements**

**Author details**

**References**

Boreskov Institute of Catalysis.

\*Address all correspondence to<EMAIL_ADDRESS>
(metaphosphate) catalyst. US4171316; 1979

[5] Hönicke D. Partial oxidation of 1,3-butadiene on V2

Products and reaction routes. Journal of Catalysis. 1987;**105**:10-18

[7] Monnier JR, Muehlbauer PJ. Epoxidation catalyst. US5081096; 1992

Development. 1983;**22**(4):570-577

olefins. US5362890; 1999

Catalysis A: General. 2001;**221**:73-91

tion on performance of Ag–MoO3

This work was conducted within the framework of budget project No 0303-2016-0006 for

Reactivity of a Simplest Conjugated Diolefin in Liquid-Phase Oxidation: Mechanisms and Products

http://dx.doi.org/10.5772/intechopen.71259

113

Nina I. Kuznetsova\*, Lidia I. Kuznetsova, Olga A. Yakovina and Bair S. Bal'zhinimaev

[1] Pedersen SE. Preparation of maleic anhydride using a crystalline vanadium(IV)bis

[2] Cavani F, Centi G, Trifiro F. Oxidation of I-butene and butadiene to maleic anhydride. 2. Kinetics and mechanism. Industrial and Engineering Chemistry Product Research and

[3] Centi G, Trifiro F. Furan production by oxygen insertion in the 1-4 position of butadiene

[4] Trifirò F, Jiru P.About some possibilities and causes of changes in selectivity of vanadium containing zeolitic catalysts in oxidation reactions. Catalysis Today. 1988;**3**(5):519-524

[6] Schroeder WD, Fontenot CJ, Schrader GL. 1,3-Butadiene selective oxidation over VMoO catalysts: New insights into the reaction pathway. Journal of Catalysis. 2001;**203**:382-392

[8] Barnicki SD, Monnier JR, Peters KT. Gas phase process for the epoxidation of non-allylic

[9] Monnier JR, Peters KT. Selective epoxidation of conjugated diolefins. US6388106; 2002 [10] Monnier JR. The direct epoxidation of higher olefins using molecular oxygen. Applied

[11] Jin GJ, GZ L, Guo YL, Guo Y, Wang JS, Kong WY, Liu XH. Effect of preparation condi-

molecular oxygen. Journal of Molecular Catalysis A: Chemical. 2005;**232**(1-2):165

/ZrO2

O5 /A12 O3

catalyst for direct epoxidation of propylene by

/Al-coated catalysts:

on V-P-O-based catalysts. Journal of Molecular Catalysis. 1986;**35**:255-265

Boreskov Institute of Catalysis SB RAS, Novosibirsk, Russian Federation

**Scheme 9.** Transformations of heteropolytungstates in oxidation of BD to 3,4-epoxy-1-butene (EpB) [65].


**3. Result and Discussion**

### *3.1. Determination of Steady Shear Rheological Properties of Ice Cream Mix for the Formulation Optimization*

In this study, the data of the steady shear rheological properties were used for the formulation optimization of ice cream mixtures. The flow curves of the ice cream mix obtained from 17 different trial points are shown in Figure 1. There was a decrease in the slope of the shear rate versus shear stress graphs of the ice cream mixes, indicating that the viscosity of all samples decreased with increasing shear rate. The reduction in viscosity can be explained by the structural breakdown of the intermolecular interaction [35]. The ice cream mixes showed shear-thinning flow characteristics, which is typical flow behavior for an ice cream mix. The shear-thinning behavior is an important factor in choosing the pump size for mixing [35].

**Figure 1.** Steady shear rheological behavior of the ice cream mixes.

Power law model parameters ( *K* and *n* values) and determination coefficient ( *R*2) calculated for 17 different points created with the trial design are shown in Table 1. *R*2 values of the power law model were higher than 0.99. This shows that the power law model is suitable for determining the flow behavior properties of ice cream mixes. According to the ice cream formulations, *K* and *n* values differed and were found as 4.01–26.05 Pasn and 0.23–0.38, respectively. *n* values lower than 1 indicated that all ice cream mixtures exhibited the non-Newtonian pseudoplastic flow behavior (Table 1). Dairy products generally exhibit shear-thinning (pseudoplastic) behavior with flow behavior indexes of 0 < *n* <1[36]. As seen in Table 1, the sample −10 (0.3% XG, 2.5% fat, 1% CSOB) showed the lowest *K* value (4.01 Pasn), while the sample-11 (0.4% XG, 12.5% fat, 3% CSOB) exhibited the highest *K* value (26.05 Pasn). The *K* value of the samples increased with increasing CSOB, gum, and fat content. These results showed that increasing the *K* value with the increase in CSOB can improve the shear-thinning properties of ice cream mixes. The desired consistency values can be obtained by using CSOB even at low-fat and gum content. Thus, CSOB can be used for improving rheological properties in low-fat ice cream.

### *3.2. The Effect of Model Parameters on K and n Value and Determination Optimum Formulation*

Figure 2 presents the impact of gum, fat, and CSOB in the formulation on *K* value. The increase in fat, gum, and CSOB, as shown in Figure 2, resulted in an increase in the *K* value of the ice cream mixes. The structure of CSOB, which contains polysaccharides with high water-holding ability, can explain this phenomenon. This polysaccharide structure offers excellent water retention and stabilizing capabilities. CSOB can be adsorbed at the interface area and has surface-active qualities, in addition to its stabilizer action. Because of these CSOB characteristics, the *K* value of the ice cream mixes increased. Furthermore, increasing the amount of gum resulted in a considerable rise in the *K* value, particularly in formulations including xanthan gum at a specific level. The increase in *K* value in both increases in fat, CSOB, and gum content can be explained by the synergistic impact of the ice cream mix's components.

**Figure 2.** Response surface plot showing the effect of model parameters on the *K* value of ice cream mixes. (**a**): XG-CSOB, (**b**): XG-Fat, (**c**): CSOB-Fat (A: XG (xanthan gum), B: CSOB (chia seed oil by-product), C: fat (milk fat), *K*: consistency coefficient).

Table 2 showed that the response surface method (RSM) and the quadratic model were used to describe the influence of formulation components on the flow behavior parameters (*K*, *n*) of ice-cream mixtures. As seen in Table 2, the ANOVA analysis of variance was used to statistically evaluate the influence of dependent variables on the *K* value of the ice cream mixes.

**Table 2.** Quadratic model parameter's corresponding *K* value.

XG: xanthan gum, CSOB: chia seed oil by-product, *K*: consistency coefficient.

The *R*2 and Adjusted *R*2 values of the model used were determined as 0.9864 and 0.9618, respectively. The differences between Adjusted *R*2 and predicted *R*2 was lower than 0.2, and the lack of fit value was found as insignificant. These results indicated that the quadratic model could be successfully used to describe the effect of formulation on the *K* value of the samples. The *p*-value of the model was lower than 0.05, indicating that the model terms significantly affected the *K* value. In this model, A, B, C, AB, C2 are significant model terms. The linear effect of all independent variables was significant. The interaction and model terms A and B, and the quadratic effect of C was also be found as significant.

Steady shear rheological analyzes showed that the increase in the amount of CSOB in the formulation resulted in a significant increase in the *K* values of the samples. Chia seeds affect fat binding and gel-forming properties due to the functional properties of dietary fiber. Olivos-Lugo et al. [37] reported that chia seeds are high in dietary fiber (34.6%), oil contents (32.2%), and protein (24.6%). Therewith, Akcicek and Karasu [20] suggested that CSOB could be used as a fat replacer in a low-fat salad dressing. Considering the properties of CSOB in this study, it is understood that it has a stabilizer feature due to its polysaccharide content and that the proteins it contains can be adsorbed in the interface area and have emulsifier quality. Proteins show surface-active properties and decrease the interfacial tension, which is predicted to cause an increase in consistency coefficient. The stabilizer feature comes from the branched polysaccharide structure in its content, and these polysaccharides can hold water [12]. Based on these properties, CSOB causes an increase in consistency coefficient (*K*) and can be used in the production of low-fat ice cream. With the increase in the *K* value of the mixtures, the *n* value decreases and shows pseudoplastic behavior, which is the typical flow behavior characteristic of ice cream. Due to the solid particles in CSOB, it significantly increases the *K* value by affecting the viscosity of the ice cream mixture. The fat, emulsifier, stabilizer, and CSOB used in ice cream mixtures significantly increase the *K* consistency coefficient of the mixtures. The main reason for this is the synergistic interaction between the components. The *K* value of the full-fat control sample was used to determine the optimum formulation. The sample with the highest desirability value and minimum fat content was selected as the optimum formulation. The optimum formulation was determined as 2.5% fat, 0.29% XG, 2.51% CSOB.

### *3.3. Rheological Properties of Optimum and Control Ice Cream Mixtures*

The steady shear, frequency sweep, and thixotropic properties of the samples produced based on optimum formulation were compared with the values of the full-fat and low-fat control sample. The flow curves of optimum and control ice cream samples were given in Figure 3. The ice cream mixtures performed a pseudoplastic (shear-thinning) rheological behavior; that is, the shear stress increased with increasing shear rate (Figure 3). Pseudoplastic rheological behavior is the typical behavior to characterize the flow properties of most ice cream mixtures [38,39]. The optimum and high-fat control samples showed similar viscosity curves. The power law model was used to determine the consistency coefficient (*K*) and flow behavior index (*n*) values of the optimal and control ice cream mixtures. Table 3 represents the power law model parameters (*K* and *n* values) as well as the determination coefficient (*R*2). *K* values varied from 3.46 to 5.66 Pasn, *n* values from 0.30 to 0.33, and *R*2 values higher than 0.99. While the *K* value of the full-fat control samples and the *K* value of the sample containing CSOB were found to be statistically insignificant, the *K* value of both samples was found to be significantly higher than the low-fat control. This result showed that an ice cream mix similar to the consistency properties of full-fat ice cream could be produced with the use of CSOB.

**Figure 3.** Steady shear rheological behavior of ice cream mixes. FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product.

The dynamic rheological properties of ice creams produced from optimum and highfat and low-fat control ice cream formulations were investigated. The frequency sweep test can simulate the liquid behavior of ice cream samples during chewing in the mouth [40], which helps to comprehensively evaluate the impact of CSOB addition on ice cream quality. Increasing *G* and *G* values of samples with increasing frequency is evidence of gel-like behavior in ice cream samples [41]. As seen in Figure 4, the value of *G* of all samples was higher than the value of *G*, indicating that the solid character of all ice cream samples is more dominant than the liquid character. As seen in Figure 4, the *G* value of FF-IC (12.5% fat) and CBLF-IC (2.5% fat) samples were almost at the same level, which explained that 10% fat can be compensated with 2.51% CSOB. As can be seen from the graph, the LF-IC instance has the lowest *G* and the lowest *G* . The *G* and *G* values of the LF-IC (2.5% fat, 0.35% XG) were lower than the *G* and *G* values of the CBLF-IC (2.5% fat, 0.29% XG, 2.51% CSOB). As it can also be seen in Table 3, the CBLF-IC has an elastic modulus value similar to FF-IC with full-fat content (12.5%). Aziz et al. [42] investigated the effect of

adding okra gum on the rheological, textural, and melting properties of low-fat ice cream samples. It was stated that *G* values were higher than *G* values for all ice cream samples. Substituting the fat content in ice cream with okra gum increased the viscous modulus (*G*) values of the samples. Previous studies on viscoelastic properties were consistent with the results of our study in terms of *G* and *G* values. Synergetic interactions between CSOB and ice cream ingredients can lead to improved food quality and expanded food applications due to enhanced functional properties. It may also have commercial potential for cost reduction.

**Table 3.** Steady shear, dynamic, and 3-ITT rheological model parameters of FF-IC, LF-IC, and CBLF-IC.

FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product. Different lowercase letters in the same line indicate a statistical difference (*p* < 0.05).

> The dynamic rheological parameters (*K*, *K*, *n*, and *n* values) were also calculated by using the power law model (Table 3). The *R*2 values of the model were found in the range of 0.97–0.98. As can be seen in Table 3, the *K* and *K* values of the samples were in the range of 3.09–23.41 and 2.46–8.46, respectively; the values of *n* and *n* were found in the range of 0.268–0.535 and 0.212–0.288, respectively. The *K* values were higher than the *K* values for all samples. Accordingly, all of the ice cream samples showed a viscoelastic solid character. When CSOB was added to low-fat ice cream, the *K* and *K* values increased when compared to the low-fat ice cream sample.

> All ice cream samples in the third interval exhibited thixotropic behavior (Figure 5). This result indicated that all ice cream samples could recover their viscoelastic character after high sudden deformation during food processing, such as homogenization or pumping. For ice cream combinations, this flow behavior is desirable. Akcicek and Karasu [20] investigated the thixotropic behavior of salad dressing samples stabilized by chia seed oil by-products and found that recoverable characteristics in the third interval are similar to our findings. The current investigation found that CSOB enhanced the thixotropic behavior of ice cream samples following rapid deformation.

**Figure 4.** Dynamic rheological properties of the ice cream mixes. (**A**): Storage modulus (*G*) vs. ω, (**B**): loss modulus (*G* ) vs. ω. FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product.

Parameters (G0, Ge, *k*, G0/Ge) were obtained with the second-order structural kinetic model. G0, Ge, G0/Ge, *k* × 1000, and *R*2 values were 8.42–21.12, 14.81–38.73, 1.760–1.834, 11.10–27.00, and 0.995–0.996, respectively. FF-IC showed the highest G0, Ge, G0/Ge, and k × 1000 values, indicating that the full-fat control sample (FF-IC) showed the highest thixotropic behavior. The amount of fat in an O/W emulsion has a significant impact on its rheological characteristics. Therefore, the decrease in the fat content of the ice cream samples causes weak rheological properties, especially the recoverable character, as the fat content of the ice cream samples decreases. Although CBLF-IC has low-fat content (2.5%), FF-IC and CBLF-IC showed similar thixotropic behavior and viscoelastic solid character so the higher recoverable behavior obtained with CSOB addition can be explained by more intermolecular interactions by the formation of small aggregates of hydrocolloid. These rheological properties indicate that CSOB can be utilized to enhance the rheological properties and thixotropy of low-fat ice cream samples.

**Figure 5.** 3-ITT rheological properties of ice cream mixes. FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product.

### *3.4. Emulsion Stability and Microstructure Properties of Ice Cream Mixes*

The emulsion stability of the ice cream mixes was determined by the thermal loop test. The thermal loop test could be used as a fast method to measure emulsion stability by subjecting to different numbers of thermal cycles. The temperature fluctuations during processing, production, storage, and transportation stages were simulated in this test [31]. The structural or morphological changes due to the applied thermal stress are determined by the change of modules (*G\**, *G*) from cycle to cycle. Figure 6 shows the change in the *G\** value after applied thermal stress. As can be seen, after 10 applied thermal cycles, a slight increase in the *G\** value of all samples was observed. This insignificant change indicates that all samples are resistant to thermal stress and have high emulsion stability. The percent change (Δ*G\**) in the *G\** values of the samples was calculated as 14.37, 10.10, 8.37 for FF-IC, LF-IC, and CBLF-IC, respectively. This may indicate that the sample containing CSOB may show higher stability than other samples.

The fat particle size (d32), PDI value, and zeta potential values of the samples were found as 0.494–1.305 μm, (−39.13)–(−28.90) mV and 0.493–0.741, respectively (Table 3). As can be seen, the sample containing CSOB exhibited lower particle size and PDI value. These results are consistent with the thermal loop test. A decrease was observed in the zeta potential values of the samples as the water ratio increased. However, all samples have a sufficient zeta potential value. Figure 7 shows the milk fat particle distribution. The high-fat control sample and the sample containing CSOB have homogeneous droplet distribution and low-fat droplet size. These results indicated that the use of CSOB could have a positive effect on the fat droplet size and distribution, and emulsion stability in ice cream.

**Figure 6.** The change in the *G*\* value after applying the thermal loop. (**a**) CBLF-IC: low-fat ice cream with chia seed oil by-product, (**b**) FF-IC: full-fat ice cream, (**c**) LF-IC: low-fat ice cream.

**Figure 7.** Light microscopy pictures of the ice cream mixtures. FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product. Samples were characterized at room temperature at 20× magnification.

### *3.5. Quality Parameters of Ice Cream*

### 3.5.1. Thermal Properties of Ice Cream

The thermal properties of the ice cream samples are presented in Table 4. The freezing points (Tf) of FF-IC, LF-IC, and CBLF-IC were obtained as −3.66, −3.35, and −3.99 ◦C, respectively. The freezing points of the ice cream samples were significantly differed (*p* < 0.05). The freezing point temperature of ice cream is closely related to the serum phase concentration and the soluble biopolymer concentration. Generally, the Tf value decreases as the serum phase concentration increases or the molecular weight of the soluble biopolymers decreases [43,44]. Soukoulis, Lebesi and Tzia [33] investigated the effect of different fiber contents on the thermal properties of ice cream. Similar to our study, a significant decrease was observed in the Tf values of ice creams when 2% wheat and oat fiber were added. Researchers explained this result by increasing the serum phase concentration due to the high water holding capacity of wheat and oat fiber. They reported an increase in the Tf value with the addition of apple fiber and inulin, which have higher soluble fiber content. The high water holding capacity of the insoluble fibers in the CSBO content may have caused a decrease in the Tf value. In addition, the use of less XG in the sample containing CSOB may have caused a decrease in the less soluble biopolymer substance in the serum phase, thus significantly reducing the Tf value. The higher Tf value obtained from LF-IC could be due to a decrease in solid concentration by reducing fat contents.

**Table 4.** Thermal properties of the control ice cream mixtures and optimum ice cream mixture contained CSOB.

FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product. Different letters in the same column indicate a statistical difference (*p* < 0.05).

The melting resistance of ice cream represents the ability of ice cream to resist melting when exposed to high temperatures. The heating system in the DSC provides the temperature that causes the formation of an endothermic peak. The melting enthalpy ( ΔH) is calculated by integrating with the area of the endothermic peak and is the amount of energy leaving the system. It occurs as a change in the overall internal energy of food [45]. In our study, the melting enthalpy values of the samples were found as 166.0, 199.3, and 146.8 J/g for the FF-IC, LF-IC, and CBLF-IC, respectively. The freezable water amount is a critical parameter affecting ice melting enthalpy in ice cream. Melting enthalpy is less in ice creams with lower freezable water content [46]. The enthalpy value of the sample CBLF-IC (containing CSOB) was found to be lower than that of FF-IC and LF-IC (the full-fat and low-fat control samples, respectively). These results can be explained by the interaction between water and CSOB. The addition of CSOB to ice creams mix could increase the amount of bound water and reduced the amount of freezable water thanks to its water-binding capacity, which may lead to a decrease in enthalpy. In addition, as the fat reduction in ice cream samples was balanced by adding water, it caused high ice formation in ice cream. However, the CSOB produced a balanced effect by chemically binding the free water, preventing excessive ice crystal formation. The lower enthalpy value of ice cream containing CSOB than FF-IC despite higher water content could be explained by the higher water retention capacity and lowering amount of freezable water.

ΔT values were found as 14.16, 15.95, and 17.05 for the samples CBLF-IC, LF-IC, and FF-IC, respectively. The sample containing CSOB had the lowest ΔT value than control samples. The temperature range ( ΔT) could be used as an indicator of the uniformity of the size distribution of ice crystals. Therefore, a narrow melting temperature range indicates a more homogeneous distribution of ice crystals melting in a smaller temperature range [47]. The enrichment of ice creams in terms of polysaccharides and protein with CSOB facilitates the formation of tiny ice crystals, contributing to the improvement of texture perception and stability of ice crystals during cold storage.

### 3.5.2. Overrun Properties of Ice Cream

The overrun values are shown in Table 4. There was a significant difference between the overrun values of the samples. The sample containing CSOB exhibited the highest overrun value, while the low-fat control sample exhibited the lowest overrun value. The increase in the overrun value with the addition of CSOB may be due to the protein and high molecular carbohydrates in the CSOB content. Proteins play an important role in increasing foam stability thanks to their emulsification properties [48]. With the addition of CSOB to the formulation, both the protein content is gained, and the high molecular carbohydrate ratio is increased. Thus, an increase in the overrun value was observed with the addition of CSOB. Akalın et al. [49] reported that dietary fibers obtained from orange, apple, and wheat provided a significant increase in overrun values compared to the control sample. On the other hand, Mansour et al. [50] reported that the addition of datary fiber powder significantly reduced the overrun value of the ice cream samples. Some researchers have suggested that there is a relationship between the overrun value and the rheological properties [6,26]. Samakradhamrongthai et al. [51] reported that the overrun value increased with increased ice cream mix viscosity. The higher overrun with increasing viscosity could be explained by a more efficient breakdown of the incorporated air cells to a smaller air cell size of ice cream mix during freezing [51,52]. Similarly, the increase in the overrun value with the addition of CSOB in our study can be explained by the increase in the consistency index of the ice cream mix.

### 3.5.3. Sensory and Color Quality of Ice Cream

Sensory scores of ice cream samples are shown in Table 5. As can be seen, a significant difference was observed between the sensory scores of FF-IC and LF-IC. Whole-fat ice cream showed the highest value in all criteria. Low-fat ice cream containing CSOB (CBLF-IC), on the other hand, showed similar sensory properties to the full-fat sample (FF-IC),

except for the color and appearance criteria. With the addition of CSOB, the change in color values compared to the full-fat sample (FF-IC) is expected. There was no significant difference in the overall acceptability criteria of the CSOB-containing sample (CBLF-IC) and the full-fat control sample (FF-IC). This indicated that the color difference observed with the addition of CSOB could not adversely affect the consumability of ice cream. Eskandari and Akbari [53] and Akın et al. [54] reported that the addition of dietary fiber and other fat replacers did not cause a negative change in sensory scores of ice cream, similar to our study. In another study [51], the reduced fat ice cream prepared with inulin showed an acceptable sensory score, similar to our study. These results indicated that with the addition of CSOB, the tested quality properties of ice cream could be improved without adversely affecting the sensory properties and that low-fat ice cream could be produced in a similar way to achieve the quality properties of full-fat ice cream. *L\**, *a\** and *b\** color values of the samples are presented in Table 6. *L\**, *a\** and *b\** values of the samples were found as 77.08–83.42, 4.23–4.93 and 13.63–15.41, respectively. As can be seen, a significant difference was detected between the color values of the samples. The highest *L\** value was measured from the FF-IC sample, while the lowest *L\** value was obtained from the CBLF-IC sample. The high *L\** value of the FF-IC sample can be explained by the higher fat content than the other two samples. In addition, with the introduction of CSOB into the ice cream formulation, a decrease in *L\**, *a\** and *b\** values was observed. This result shows that CBLF-IC will cause a significant change in the color values of the ice cream samples.

**Table 5.** Sensory analysis of ice cream samples.

FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product. Different letters in the same column indicate a statistical difference (*p* < 0.05).

FF-IC: full-fat ice cream, LF-IC: low-fat ice cream, CBLF-IC: low-fat ice cream with chia seed oil by-product. Different letters in the same column indicate a statistical difference (*p* < 0.05).


**3. Results**

#### **3.1. LMW activator is considered to be HATKTAK**

As it was shown in a previous study that both thrombin and trypsin could produce a substance with LMW activator activity from platelet lysate [27] and that incubation of Apo CIII with thrombin resulted in production of LMW activator, Apo CIII was treated with thrombin or trypsin and the LMW activator activities produced were compared to determine whether LMW activators produced by these enzymes are the same substance. Both thrombin and trypsin could produce LMW activator activity, but 1 unit/ml trypsin could produce LMW activator 100 to 1,000 times more effectively than 0.1 unit/ml thrombin (Figure 1A).

The LMW activator activities prepared using thrombin and trypsin appeared at the same NaCl concentration of the MONO S cation exchange chromatography (0.2 M NaCl, pH 8.0) (Figure 1B and Figure 1C, respectively), and at the same elution volume (16 ml) of the Superdex peptide gel filtration (Figure 1D and Figure 1E, respectively), which corresponds to the approximate molecular size of 800 Da (Figure 1F). These findings suggest that the two LMW activators are the same substance.

The fact that LMW activator can be produced by trypsin digestion of Apo CIII suggests that the C terminal of LMW activator consists of K or R [35]. LMW activator is suggested to be one of the basic peptides that would appear after trypsin digestion of Apo CIII (S1-K24, H18-R40, H18-K24, H18-K21, T22-K24) (see Figure 1G for the sequence of Apo CIII, distribution of basic and anionic amino acids and the sugar binding amino acid). These peptides were artificially produced and filtered on the Superdex peptide column to use as molecular size markers. The optical peaks of OPA fluorescence and LMW activator activity by H18-K24 (HATKTAK) appeared at the same elution volume of Superdex peptide gel filtration corresponding to Apo CIII-derived LMW activator (Figure 1D, 1E, 2A and 2B). In MONO S cation exchange chromatography, the action of HATKTAK as LMW activator was recovered at the fraction with the same NaCl concentration (0.2 M NaCl, pH 8.0) (Figure 2C) as the Apo CIII-derived LMW activator (Figure 1B and 1C).

#### **3.2. Only HATKTAK among examined peptides showed MAPP-forming activity**

Among the peptides used for calibration on Superdex peptide gel filtration (Figure 1F), H18-R40 and H18-K21 were separated in the fractions near to HATKTAK (LMW activator). The activity of these peptides as LMW activators were examined. Only HATKTAK showed LMW activator activity with a peak at 1 nM (Figure 2D). Then, some Apo CIII-derived peptides that contain part or all of H18-K24 were examined for their LMW activator activity. All of them were used at a concentration of 1 nM. Only HATKTAK showed MAPP-forming activity (Table 1).

**Figure 2.** HATKTAK functions as the LMW activator. **A.** Superdex peptide gel filtration of HATKTAK (200 g, 0.2 ml). The peptide concentration of each fraction was measured by the OPA fluorescent

method. The inset shows the standard graph for the assay of HATKTAK. **B.** MAPP formation using fractions of Superdex peptide gel filtration of HATKTAK (200 g, 0.2 ml) at a dilution of x106 by incubation with one of the precursors of MAPPs. **C.** MONO S cation exchange chromatography of 1 g/ml HATKTAK. MAPPs were generated by incubation of each fraction from the chromatography at a dilution of x104 and one of the precursors of MAPPs. **D.** MAPP formation using peptides containing part of the sequence of apolipoprotein CIII. Each peptide was serially diluted x10 and incubated with one of the precursors of MAPPs. B, **C** and **D** were performed using the precursor of s-MAPP.

728 Lipoproteins – Role in Health and Diseases

**Figure 2.** HATKTAK functions as the LMW activator. **A.** Superdex peptide gel filtration of HATKTAK (200 g, 0.2 ml). The peptide concentration of each fraction was measured by the OPA fluorescent

**Table 1.** MAPP formation from precursors of MAPPs with peptides derived from K16 to D25 of Apo CIII (KHATKTAKD)

#### **3.3. LMW activator activity in platelet release products and platelet lysate**

In the next part of the study, the LMW activator activities in variously prepared samples including platelet release products and platelet lysate were compared to determine whether platelets release LMW activator. Platelet release products were prepared using platelets from fresh and stored platelet-rich plasma. The reason why we examined platelet release products prepared from stored platelet-rich plasma along with those prepared from the fresh equivalent is that platelet releasate from stored platelet-rich plasma was expected to contain a higher concentration of LMW activator because platelets lose the precursors of MAPPs during storage [26]. From all of these samples, the low-molecular-weight fraction was separated using PD10 columns, and the activity corresponding to LMW activator was compared by the largest dilution from the original sample for which a phagocytic index higher than 150 was recorded (effective dilution).

1 ng/ml HATKTAK and platelet release products from fresh platelets stimulated with 4 mEq/l Ca++ and 0.1 unit/ml thrombin showed an effective dilution of 102 to 103, whereas fresh platelets stimulated only with 4 mEq/l Ca++ or 0.1 unit/ml thrombin for 1 minute showed far lower effective dilution, and platelet release products prepared from platelets after storage for 72 hours and 120 hours in the form of platelet-rich plasma showed extremely high effective dilution (103 to 105, platelets from platelet-rich plasma stored for 72 hours; 107 to 108, those stored for 120 hours). None of the platelet lysate produced from platelets in fresh and stored platelet-rich plasma showed LMW activator activity (Figure 3).

To confirm whether these LMW activator activities were by the same substance, Superdex peptide gel filtrations of the low-molecular-weight fractions of platelet release products prepared from fresh platelets (A), and those stored for 72 hours (B) and 120 hours (C), with stimulation with 0.1 unit/ml thrombin in the presence of 4 mEq/l Ca++ were performed. Fractions of the gel filtrations were diluted x102 (A), x104 (B) and x106 (C). All of these samples showed LMW activator activity at the fraction corresponding to HATKTAK (compare Figure 4A, 4B and 4C with Figure 2B).


**AMM Sharif Ullah**

Faculty of Engineering, Kitami Institute of Technology, 165 Koen-cho, Kitami 090-8507, Japan<EMAIL_ADDRESS>
Received: 27 July 2019; Accepted: 26 August 2019; Published: 29 August 2019

**Abstract:** This article addresses some fundamental issues of concept mapping relevant to discipline-based education. The focus is on manufacturing knowledge representation from the viewpoints of both human and machine learning. The concept of new-generation manufacturing (Industry 4.0, smart manufacturing, and connected factory) necessitates learning factory (human learning) and human-cyber-physical systems (machine learning). Both learning factory and human-cyber-physical systems require semantic web-embedded dynamic knowledge bases, which are subjected to syntax (machine-to-machine communication), semantics (the meaning of the contents), and pragmatics (the preferences of individuals involved). This article argues that knowledge-aware concept mapping is a solution to create and analyze the semantic web-embedded dynamic knowledge bases for both human and machine learning. Accordingly, this article defines five types of knowledge, namely, analytic a priori knowledge, synthetic a priori knowledge, synthetic a posteriori knowledge, meaningful knowledge, and skeptic knowledge. These types of knowledge help find some rules and guidelines to create and analyze concept maps for the purposes human and machine learning. The presence of these types of knowledge is elucidated using a real-life manufacturing knowledge representation case. Their implications in learning manufacturing knowledge are also described. The outcomes of this article help install knowledge-aware concept maps for discipline-based education.

**Keywords:** concept map; learning; semantic web; knowledge representation; epistemology


**3. Asthma Exacerbation and miRNAs**

MiRNA molecules perform various functions in the body. In addition to the diagnostic role described above, they can be used to predict exacerbation of the disease with greater accuracy. The risk of exacerbation of the disease increases with the lack of use of inhaled corticosteroids, lack of adequate disease control, spirometric indices indicating deepening of obstruction, signs of eosinophilic inflammation, or exposure to allergens or irritants. In a group of children diagnosed with asthma and using anti-inflammatory treatment (inhaled steroids) for a period of 12 months, 12 miRNA molecules associated with exacerbations during observation were isolated. In addition, each doubling of the concentration of these molecules was associated with an increase in the risk of exacerbations by 25–67%, respectively. MiRNA-146b-5p, miRNA-206, and miRNA-720 were identified, which, in combination with clinical symptoms, enabled a better prediction of exacerbation of the disease in patients with asthma using inhaled corticosteroids, compared to a model based only on a single component. Involvement of these miRNAs in pathways involved in the pathogenesis of asthma was discovered, e.g., NF-kβ and GSK3 (glycogen synthase kinase-3)/AKT, which are responsible for remodeling the airways and deepening the inflammatory process [2].

MiRNAs are biomolecules that also play a key role in other obstructive respiratory diseases, including chronic obstructive pulmonary disease (COPD). Researchers compared circulating serum miRNAs associated with exacerbation of the disease in the pediatric population diagnosed with asthma and adults diagnosed with COPD. A total of 20 miRNA molecules were associated with worsening of symptoms in children and 5 of them (451b; 7-5p; 532-3p; 296-5p, and 766-3p) in the adult population with severe chronic obstructive pulmonary disease. Participation of these molecules in the signaling pathways MAPK (mitogen activated protein kinases) and PI3K-Akt, which are responsible for the increased response of both eosinophils and neutrophils, the production of immunoglobulin IgE, and the activation of tumor necrosis factor alpha (TNF-a), was proven. This leads to excessive production of interleukins, cytokines, and intensification of inflammation [24]. In addition, the MAPK pathway is activated by tobacco smoke, indicating its important role in the pathogenesis of COPD and neutrophilic asthma. Thanks to the analysis of the concentrations of appropriate miRNAs, it is also possible to identify the pathomechanism that may lead to exacerbation of symptoms among children with asthma who are exposed to tobacco smoke in the home environment. This is undoubtedly a factor that stimulates an increased immune response and affects the disease symptom severity. However, anti-inflammatory molecules have been identified that are also related to the concentration of the relevant miRNAs. One of them is annexin (ANXA1), also known as lipocortin I. It is a protein with a significant content in the secretions of the respiratory tract. It was shown that an increase in ANXA1 concentration was associated with the decreased concentration of miRNA-196-a2. It is also interesting that in subjects with a moderate disease severity, the level of ANXA 1 was higher than in those with severe asthma. In addition, miRNA-196-2a is responsible for the production of key interleukins involved in the pathogenesis of asthma, IL-5 and IL-13, whose role is described above [25–27]. Many factors can exacerbate the course of asthma, such as tobacco smoke, infections (mainly viral), allergens, irritants (e.g., aerosols or household cleaners, paint fumes, other occupational exposures), physical exertion, air pollution, medications, and foods. Respiratory infections caused by viruses, including RSV, influenza, and rhinoviruses, are of particular importance among the group of the youngest patients. When these pathogens infect human bronchial epithelial cells (HBECs), it was demonstrated that the NF-κB pathway and interferon signaling are activated to stimulate cellular responses, reduce viral replication, and avoid tissue damage. It was suspected that the HBEC cells of asthmatics impair the above-described processes and, consequently, exacerbate symptoms. Understanding miRNAs targeting the NF-κB and interferon pathway will uncover modulators of cellular responses and prevent the development of adverse events [28]. MiRNA-146a and miRNA-146b (miRNA-146a/b) are anti-inflammatory molecules more heavily produced in response to rhinovirus (RV) infection, targeting the NF-κB pathway specifically. In experimental animal models of allergic asthma, deprivation of HBEC cells of these miRNAs led to an increased inflammatory process involving Th1, Th17 lymphocytes with a reduced participation of Th2 lymphocytes. These molecules are therefore responsible for alleviating RV-induced allergic airway inflammation and represent a potential future therapeutic target [29]. Influenza virus is another important infectious agent responsible for exacerbating asthma symptoms. After infection of bronchial epithelial cells with H1N1 influenza virus in the laboratory, miRNA-22 growth was demonstrated only in a sample obtained from healthy individuals. The molecule blocks the CD147 receptor which is a transmembrane glycoprotein involved in the invasion of viral and bacterial infections. This molecule also participates in the remodeling of the respiratory tract through the increased synthesis of matrix metalloproteinases (MMPs). Therefore, asthma patients with low levels of miRNA-22 due to the inability to block CD147 lose one of the important defense barriers against viral infection, becoming more susceptible to infection. In the era of the COVID-19 pandemic, it was shown that the CD147 receptor is also responsible for the penetration of the SARS-CoV-2 virus into the cell. This proves the role of miRNA-22 in the defense process against a wider spectrum of viral infections than just influenza [30].

Unfortunately, not only infectious factors play a leading role in the pathogenesis of asthma in children, but also increasing air pollution and high concentrations of harmful substances in the environment. After measuring the content of PM 2.5 particles in the composition of the air at home, a significant correlation was shown between their concentration and the incidence of asthma. Moreover, greater exposure resulted in an increase in serum concentrations of miRNA-155, which is responsible for the enhancement of the type 2 immune response [31]. The relationship of this biomolecule was also confirmed with exposure to tobacco smoke, which is a special type of air pollution due to its local occurrence and relative ease of elimination from inhaled air compared to other factors. MiRNA-155-5p, miRNA-21-3p, and miRNA-18a-5 are a set of molecules that are overexpressed in the blood of mice exposed to cigarette smoke in utero, in which asthma was later induced with albumin. Additionally, their concentrations were positively correlated with proasthmatic Th2 cytokine levels in bronchoalveolar lavage fluid (BALF) samples [32]. Another aspect that was emphasized was the seasonal variability of miRNAs and its impact on the severity of the allergic process and asthma symptoms in the pediatric population, especially in the spring. It is at this time of the year patients are at risk of exacerbating asthmatic symptoms, especially those with hay fever. Of the 26 miRNAs studied, which were associated with a specific season and with the allergen causing the symptoms of this period, two miRNAs-328-3p and let-7d-3p were isolated. A decrease in the concentration of let-7d-3p was observed in spring and among those allergic to mulberry (spring blooming), while it increased after the allergen immunotherapy process (performed in patients allergic to wasp venom). The protective role of this molecule was proven, which is most likely achieved by reducing the concentration of IL-13. Its protective effect in relation to asthmatic patients is also supported by its abundant presence in the lung tissue. The role of miRNA-328-3p is more complex. It was observed that its concentration decreases in autumn. On the other hand, an increase was noted in patients with a concomitant allergy to aspergillus, which resulted in an exacerbation of symptoms. MiRNA-328-3p was shown to be involved in wound healing of the bronchial epithelium, but also facilitates the spread of bacterial infection in the lungs [33].

MiRNAs may participate in the process of airway remodeling, leading to airway obstruction and affecting airway function parameters. MiRNAs involved in a number of pathophysiological processes affect smooth muscle cells, the epithelium, and goblet cells and intensify the inflammatory process, causing narrowing of the bronchial lumen [34].

The relationship between 22 miRNAs and lung function parameters in the pediatric population was demonstrated. An example may be the following molecules: miRNA-186-5p, which participate in the release of acetylcholine and modulation of airway tone through the cholinergic pathway [35,36]; miRNA-203, which is associated with an increase in IgE concentration and intensifies the inflammatory process in the airways, leading to their obstruction [35,37]; and miRNA-26, which is released by bronchial smooth muscles after their physical stretching, causing cell hypertrophy. Over the course of many years of observation, it has also been proven that miRNA-145-5p is associated with an early decrease in FEV1 in children with asthma, leading to the development of COPD. It is also responsible for the increased proliferation of airway smooth muscle cells [38].

The results of the studies on miRNAs in pediatric asthma are summarized in Table 1.

**Table 1.** The relevance of miRNA in asthma based on studies conducted in the pediatric population.

**Table 1.** *Cont.*

receptor subfamily three group C member 1 gene; PTEN—phosphatase and tensin homolog gene; GSK3—glycogen synthase kinase-3; AKT—protein kinase *B*; NF-kB—nuclear factor kappa-light-chain-enhancer of activated B cells; MAPK—mitogen-activated protein kinases, PI3K—the phosphatidylinositol 3-kinase; Ige—immunoglobulin E; GATA3—GATA binding protein three; Th1—Type 1 T helper; Th2—Type 2 T helper.


**CFD Simulations of Marine Hydrodynamics**

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**Yigit Kemal Demirel**

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**5. Conclusions**

Based on the importance of the MYB family in plants and the strong adaptability of *H. hamabo* to stressed environments, genome-wide identification and analysis of the MYB transcription factor family in H. hamabo were performed for the first time in this study. The search for MYB family members in the *H. hamabo* genome led to the identification of 204 putative HhMYB family members. The HhMYB family was clustered into 28 subfamilies with AtMYBs according to phylogenetics and their motif patterns. In addition, we found that the 204 *HhMYB* genes were unevenly distributed across the 46 chromosomes of *H. hamabo*. Expression analysis with transcriptomic data and qRT-PCR showed that the candidate HhMYB family displays a wide range of sensitivities to abiotic stress and hormones. Several *HhMYB*s were involved in the plant responses to both saline and drought stresses to varying degrees, indicating that these *HhMYB*s may have regulatory functions in tolerance to saline and drought stress and deserve further investigation. Our results provide a foundation for further researching MYB gene functions in *H. hamabo.*

**Supplementary Materials:** The following are available online at https://www.mdpi.com/article/ 10.3390/plants12071429/s1. Figure S1: Phylogeny (a), conserved motif (b), and gene structure (c) analyses of HhMYBs; Figure S2: Chromosome distribution of *HhMYB*s; Table S1: Primer sequences for qRT-PCR; Table S2: List of the MYB family genes in *H. hamabo.*

**Author Contributions:** Conceptualization, C.G. and Z.W.; investigation, D.L. and Z.F.; writing original draft preparation, D.L. and Z.W.; writing—review and editing, Z.W. and C.G. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by Six Talent Peaks Project of Jiangsu Province [NY-042].

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** No new data were created or analyzed in this study. Data sharing is not applicable to this article.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**

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