Heterogeneous Interesterification of Triacylglycerols Catalyzed by

Article pubs.acs.org/JAFC

Heterogeneous Interesterification of Triacylglycerols Catalyzed by Using Potassium-Doped Alumina as a Solid Catalyst Wenlei Xie* and Jing Chen School of Chemistry and Chemical Engineering, Henan University of Technology, Zhengzhou 450001, People’s Republic of China ABSTRACT: Heterogeneous interesterification of vegetable oils offers an environmentally more attractive option for the modification of edible oils to meet the specifications for certain food applications. In this work, potassium-doped alumina (KNO3/Al2O3) was prepared using an impregnation method, followed by calcinations at a temperature of 700 °C, and was then employed as heterogeneous catalysts for the interesterification of triacylglycerols. The solid catalyst was characterized by means of Hammett titration method, power X-ray diffraction, scanning electron microscopy, and nitrogen adsorption−desorption techniques. It was determined that the catalyst with KNO3 loading of 35% on alumina support and calcined at 700 °C exhibited the best catalytic activities toward the interesterification between soybean oil and methyl stearate under solvent-free conditions. Also, the solid base catalyst was successfully applied to the interesterification of soybean oil and lard blends in a heterogeneous manner. The physicochemical properties of the interesterified products were investigated using gas chromatography, highperformance liquid chromatography, and confocal laser scanning microscopy. It was found that the slip melting point and crystal morphology had a significant variation after the interesterification reaction as a result of the modification in the TAG profile. With the solid base catalyst, an environmentally friendly approach for the interesterification of triacylglycerols in a heterogeneous manner was developed. KEYWORDS: heterogeneous catalyst, interesterification, triacylglycerol, soybean oil, lard

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INTRODUCTION The physicochemical and functional properties of triacylglycerols are highly dependent on the nature of fatty acids (FAs) on the glycerol backbone. Native vegetable oils have limited applications in the edible oil industry in their original form.1,2 However, they can be modified in order to enhance their functional performance to meet the specifications for certain food applications.3 The modifications such as hydrogenation and interesterification, are the commonly used methods to change the physicochemical characteristics of food oils. Unfortunately, the hydrogenated oils always contain larger amounts of trans FAs, which are known to be detrimental to human health since the intake of trans FAs can contribute the increased risk factors for cardiovascular disease.4 Interesterification of vegetable oils has been considered as an alternative approach to conventional hydrogenation for obtaining zero trans fats with great potential applications.5 This interesterification process has been widely used to formulate margarine and shortening fats with zero trans fat and to improve functional and nutritional properties. The interesterified product exhibits beneficial effects in comparison with the physical mixtures of oils having similar FA compositions.6−8 In general, the interesterification reaction can be carried out chemically or enzymatically, effectively altering the FAs composition or their positional distribution in triacylglycerol (TAG) molecules. Enzymatic interesterification (EIE) processes have several benefits, such as reaction specificity, milder reaction conditions, and fewer side-reactions.9,10 However, the practical application of EIE reactions for the production of modified lipids still has some limitations associated with the high cost of the lipase used.11 From the industrial application point of view, homogeneous base catalysts (usually sodium © XXXX American Chemical Society

alkoxide and sodium hydroxide) have been traditionally employed as catalysts to perform the chemical interesterification (CIE) reaction in the edible oil industry, mainly thanks to their low cost and high efficiency at relatively low temperatures.12−14 This homogeneous process can result in a random distribution of FAs in the TAG molecules, since it always shows no selectivity for FA positions on the glycerol backbone. Although these homogeneous alkaline catalysts offer high catalytic activities, they suffer from the difficulty of the catalyst separating from reaction mixture, thus rendering them nonrecoverable and nonreusable.15 To overcome these difficulties, heterogeneous catalysts have become the focus of recent research for practical application from both a commercial and an environmental point of view. The merits of heterogeneous catalysts are their fast and easy separation from the reaction mixture without requiring aqueous quench and neutralization steps which themselves generate undesired wastewater.16,17 In heterogeneous catalysis, the solid catalyst is neither consumed nor dissolved in the reaction, and therefore it can be regenerated and reused. Recently, several investigators have tested the use of heterogeneous base catalysts such as alkaline metal-doped alumina as a catalyst for a variety of different organic reactions, including Knoevenagel condensation, transesterification, and coal char combustion.18−20 However, to my knowledge, there has been only a limited number of studies that have dealt with the solid base catalysts for the CIE reaction so far.21 Accordingly, the fabrication and Received: August 2, 2014 Revised: October 5, 2014 Accepted: October 9, 2014

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dx.doi.org/10.1021/jf503726a | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry

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10 mL of methanol and allowed to equilibrate for 2 h. The basic strength is quoted as being stronger than the weakest indictor that shows a color change, and weaker than the strongest indicator, which exhibits no color change. The basicity of the solid catalyst was measured by the method of Hammett indicator involving the benzene carboxylic acid (0.02 mol/L methanol solution).17 Scanning electron microscope (SEM) measurements were performed with a field-emission microscope (JEOL, JSM-6390LV) using an accelerating voltage of 15 kV. The nitrogen adsorption− desorption isotherms at −196 °C were measured using a Quantachrome NOVA 1000e instrument. The Brunauer−Emmett− Teller (BET) method was used to calculate the specific surface area. The total pore volume and pore size distribution were estimated according to the Barrett−Joyner−Halenda (BJH) method based on the adsorption isotherm. Interesterification Procedures. The interesterification of soybean oil and methyl stearate was carried out in a 50 mL round-bottom flask. Prior to each reaction, a mixture of 20 g of soybean oil and 41 g of methyl stearate was dried in the reactor under reduced pressure at 80 °C. By adding 10 wt % of the solid catalyst, the heterogeneous reaction mixture was allowed to proceed under reduced pressure at 200 °C. Constant stirring at 750 rpm was used to minimize mass transfer limitations. After 4 h of reaction time, the reactor was cooled to room temperature. Thereafter, the catalyst was recovered from the reaction mixture by simple filtration, and the filtrate was then used for subsequent analysis. Soybean oil and lard were blended in different proportions of 80:20, 60:40, 40:60, 20:80 ratios (w/w), and then the binary blends were interesterified in batch reactors. In order to avoid oil oxidation, the interesterifications were carried out in a nitrogen atmosphere. Portions (100 g) of the blends were initially heated under vacuum at 80 °C to remove air and moisture. When the reaction mixture had reached a reaction temperature of 100 °C, 5 wt % of the solid catalyst was added in a batch reactor, and the blends were stirred vigorously. The interesterification reaction of different blends of soybean oil and lard was performed under reduced pressure at 100 °C in a 150 mL stoppered flask with magnetic stirring (∼750 rpm) for a time frame of 5 h. At the end of the reaction, the interesterified product was filtered, and then employed for subsequent analysis. Analytical Methods. The FA compositions of triacylglycerols were analyzed by a gas chromatography (GC) according to AOAC method.24 After complete conversion of FA residues into fatty acid methyl esters (FAMEs), the methylated FA residues were determined using an Agilent 6890 N gas chromatograph (Santa Clara, CA. U.S.A.) equipped with a flame-ionization detector, an auto injector and a fused silica capillary column (60 m × 0.25 mm) coated with 0.25 μm of BPX-70 (SGE, Australia). Nitrogen was used as a carrier gas at flow rate of 1.2 mL/min with a split ratio of 1:20. The temperatures of injector and detector were set at 260 and 300 °C, respectively. The initial temperature of the program was increased to 160 °C and then held for 5 min. This temperature was finally increased to 200 °C at a rate of 5 °C/min, and held isothermally for another 42 min at the final temperature. The FAME composition was identified by comparison with the relative retention time of FAME standards. The FAME content was obtained by area normalization and expressed as mass percentage.25 For the interesterification of soybean oil with methyl stearate, the triacylglycerol was separated from the reaction mixtures by thin-layer chromatography (TLC) on a silica gel 60G plate. The interesterified blends were spotted on TLC silica gel plates, and then developed with petroleum ether/ethyl ether/acetic acid (90:10:1,v/v/v). The developed TLC plates were sprayed with 2% 2,7-dichlorofluorescein in methanol. Bands corresponding to triacylglycerols, which were identified using triolein as the standard, were scraped from the silica plates, extracted with n-hexane, and subsequently analyzed for the FA profiles. The progress of the interesterification reaction was followed by the determination of stearoyl incorporation into the TAGs. The FA profiles at the sn-2 position of triacylglycerols were measured by using pancreatic lipase according to the method described in the literature.25 The pancreatic lipase can be applied to

development of heterogeneous catalysts for the interesterification reaction in an environmentally friendly manner has become an area of great importance particularly from the green chemistry point of view. The interesterification reaction involves as least two oils with different FA compositions. Due to the complex compositions in the oils, it is hard to evaluate quantitatively the interesterification process. For the optimization of the catalyst preparation conditions, in the current study, soybean oil and methyl stearate were used as reactants, and the incorporation of stearoyl groups into TAGs was determined by gas chromatography (GC) techniques.22 Besides, the interesterification reaction between soybean oil and lard was also carried out to investigate the properties of the interesterified products. Alumina has attracted much interest as a heterogeneous catalyst support, due to its high surface area, simple structure and availability.23 Thus, in the present study, KNO3/Al2O3 catalysts with different KNO3 loadings were prepared using alumina as a catalyst support, and then were tested in the interesterification reaction. The prepared solid catalysts were characterized by using Hammett titration method, power X-ray diffraction (XRD), scanning electron microscopy (SEM), and nitrogen adsorption−desorption techniques. The effect of calcination temperature and KNO3 loading was investigated with the interesterification of soybean oil with methyl stearate in terms of stearoyl incorporation into the TAGs. By using this solid base catalyst, the interesterification reactions of soybean oil and lard blends were carried out in a batch reactor, and the properties of the interesterified products were investigated by gas chromatography (GC), high-performance liquid chromatography (HPLC), and confocal laser scanning microscopy.

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MATERIALS AND METHODS

Materials. Refined soybean oil and lard used in this study were purchased from the local supermarket. Methyl stearate (≥98%) was obtained from Sinopharm Chemical Reagent Corporation (Shanghai, China). The alumina used as a support, with a surface area of 123 m3/ g, was obtained from Tianjin Chemical Reagent Factory. Porcine pancreatic lipase (EC 3.1.1.3, Type II; PPL) used for the analysis of FA profiles at the sn-2 position of the interesterified TAGs, was purchased from Sigma-Aldrich Corporation (St. Louis, U.S.A.). All the other materials used were of analytical or chromatographical grades. Catalyst Preparation. The potassium-doped alumina catalysts (KNO3/Al2O3), with different loadings of KNO3 over the alumina support, were prepared by the conventional incipient-wetness impregnation method with an aqueous solution of KNO3 using commercial alumina as a support. In a typical preparation, the required amount of alumina was placed into the aqueous KNO3 solution. The resulting suspensions were stirred for 24 h at room temperature, and water was then evaporated at 80 °C under reduced pressure. The concentration of initial KNO3 solution was changed to prepare the solid catalyst with different KNO3 loadings. After impregnation the white solid obtained hereby was dried overnight and was finally calcined under an air flow at 500−900 °C for 6 h prior to the catalytic tests. The solid catalysts prepared with various KNO3 loadings of 30, 35, 40, and 45 wt %, were expressed as 30%KNO3/Al2O3,35%KNO3/ Al2O3, 40%KNO3/Al2O3, and 45%KNO3/Al2O3, respectively. Catalyst Characterization. Powder X-ray diffraction (XRD) patterns of selected samples were obtained with a Bruker-AXS NanoSTAR instrument. Nickel filtered Kα radiation of copper was used at 40 kV and 30 mA. The data were analyzed with the DiffracPlus software, and the phases were identified using the powder diffraction file database (JCPDS, International Centre for Diffraction Data) Basic strengths of the solid catalyst (H_) were determined by using various Hammett indicators. For this purpose, about 300 mg of the sample was placed into 1 mL Hammett indicator solution diluted with B



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hydrolyze selectively FAs at sn-1,3 positions of TAGs, producing 2monoacylglycerol and FAs. The formed 2-monoacylglycerol was separated by TLC plates, and the sn-2 positional analysis of FA residues in the TAGs was carried out by GC techniques after 2monoacylglycerol was totally converted to FAMEs as described previously. For the interesterification of soybean oil with lard, the variation of TAG compositions occurred during the reaction was assessed using high-performance liquid chromatography (HPLC) fitted with commercially packed Genesis C18 column (150 mm × 4.6 mm2).25 Each sample was dissolved in chloroform (10 mg/mL), and 20 μL of the dissolved sample was injected into the HPLC system. The elution solvent consisted of dichloromethane and acetonitrile (HPLC grade) at a gradient composition from 35% acetonitrile increasing to 55% dichloromethane in 45 min. The column temperature was held at 40 °C. The TAG species were detected by an evaporative light scattering detector (Alltech 500). Individual TAG species were identified tentatively by comparing the relative retention time with those of TAG standards and according to the literature.26,27 The iodine value (IV) for the interesterified product was measured according to the AOCS Official Method Cd 1c-85 (AOCS, 2009).25 Slip melting point (SMP) was determined according to the AOCS Official Method Cc 3−25 (AOCS, 2009) using the open capillary tube method,25 The capillary tubes filled with 1 cm high column of the samples were stored in a refrigerator for 16 h prior to the measurements in a beaker of cold water. Thereafter, the sample tube was heated gradually, and the temperature at which the sold fat in the tube began to rise, was considered to be the SMP. This measurement was performed in triplicate, and the reported SMPs were the average of the determinations. The crystal morphology of the interesterified product was observed using a XP-203 polarized light microscopy fitted with a digital video camera. The samples were first heated at 70 °C in a water bath and maintained at this temperature for 30 min to eliminate all crystal nuclei. Afterward, about 10 mg of melted sample was placed on a preheated microscope slide, cooled to 25 °C and then held for 20 h. The crystal morphology was taken at room temperature.

Figure 1. Power X-ray diffraction patterns for samples: (a) uncalcined 35%KNO3/Al2O3; (b) 30%KNO3/Al2O3 calcined at 700 °C; (c) 35% KNO3/Al2O3 calcined at 500 °C; (d) 35%KNO3/Al2O3 calcined at 700 °C; (e) 35%KNO3/Al2O3 calcined at 800 °C; (f) 40%KNO3/ Al2O3 calcined at 700 °C; and (g) 45%KNO3/Al2O3 calcined at 700 °C. (◇),KNO3; (▲), Al2O3.

in number when compared with the uncalcined sample. With increasing the KNO3 loading beyond 35%, the diffraction peaks attributed to KNO3 phases were observed to increase both in intensity and in number, mostly due to the overloading of KNO3 that saturated the alumina surface. At these KNO3 loadings, the dispersion of excess potassium compound could mask the active base sites that serve as active sites for the interesterification reaction. However, no obvious peak for K2O phase was clearly registered in the XRD patterns. The catalytically active phase such as K2O might have been present in an amorphous state, which was not detected by XRD techniques. By drawing on the results, it is reasonable to state that the surface basicity and the subsequent catalytic activity are resulted from the decomposition of impregnated KNO3 and the formation of K2O and AlOK groups that can act as active sites for the interesterification reactions.18,20 SEM techniques can be used to investigate the morphology feature of the solid catalyst. The typical SEM micrograph of alumina support and solid catalyst is illustrated in Figure 2. As can be seen, the support was presented as irregular shaped agglomerations of alumina particles with clear edges, with particle size ranging from approximately 2−5 μm. As evidenced in Figure 2, no significant morphological change was found between the alumina support and the solid base catalyst. After loading with the potassium compounds, the alumina cluster or agglomeration with clear demarcation into regions could also be observed. This observation suggested that the alumina retained its structure during the catalyst preparation process, and there was a homogeneous distribution of potassium compounds on alumina supports, which were important for catalysis. The nitrogen adsorption−desorption isotherms of the alumina support and the prepared catalyst are presented in Figure 3. The two samples exhibited the type IV isotherm according the IUPAC classification with a obvious hysteresis loop of type H4, implying the existence of micropores with narrow pore size distributions. Clearly, a significant reduction in the adsorbed volume was observed after impregnation of alumina support with potassium compounds, mostly likely owing to the pores obstruction of the active sites in the support, evidencing the incorporation of potassium compounds into the alumina support.

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RESULTS AND DISCUSSION Catalyst Characterization. A Hammett indicator method was employed to evaluate the basic properties.17 The alumina support was acidic and could change the color of dimethylaminoazobenzene (H_= 3.3) from yellow to red, and in consistent with this, the transesterification did not proceed when the alumina support was acted as a catalyst. However, as the KNO3 was loaded and subsequently calcined at 700 °C, the thus-obtained solid base catalyst showed the basic strength values (H_) of 15 < H_18.4, and the basicity of 35% KNO3/Al2O3 catalyst was determined to be 6.97 mmol/g. Owing to the basicity, the solid catalyst displayed comparable activities to the reaction as expected. Figure 1 gives the XRD patterns of the KNO3/Al2O3 catalysts prepared at different conditions. As can be seen, the XRD pattern of uncalcinated KNO3/Al2O3 sample exhibited the characteristic peaks of Al2O3 and KNO3 (curve a in Figure 1).28 Besides, there were other minor peaks, which could not be unequivocally identified. In the XRD pattern of 35%KNO3/ Al2O3 catalyst calcined at 500 °C, in addition to the XRD peaks associated with alumina, only three peaks of KNO3 was observed (curve c in Figure 1). As the calcination temperature increased beyond 700 °C, the number of the XRD peak corresponding to KNO3 phase was found to further decrease, indicating the decomposition of loaded KNO3 during the calcination processes. In the case of 30% KNO3/Al2O3 catalysts calcined at 700 °C (curve b in Figure 1), the diffraction lines of Al2O3 were visible; however the KNO3 phases were decreased C



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Figure 2. Scanning electron micrographs of samples: (a) Al2O3 and (b) 35%KNO3/Al2O3 catalyst.

the surface area occurred after KNO3 was impregnated and calcined at a higher temperature. Moreover, a decrease in the pore size and pore volume was also observed. Such results further demonstrated the potassium compounds on the alumina support were formed by the decomposition of potassium nitrates at a higher calcination temperature. Influence of Catalyst Preparation Conditions on the Catalytic Activities. In the current study, the interesterification of soybean oil and methyl stearate was chosen to investigate the effect of catalyst preparation conditions on the catalytic activity. To optimize the KNO3 loading, different loading amounts of KNO3 were employed for the preparation of the catalyst. Table 2 shows the evolution of stearoyl Table 2. Different Catalyst Preparation Conditions and the Results of the Interesterification between Soybean Oil and Methyl Stearate Catalyzed by the Heterogeneous Catalyst

Figure 3. Nitrogen adsorption/desorption isotherms and pore size distribution profiles of samples: (a) Al2O3 and (b) 35%KNO3/Al2O3 catalyst.

The textural properties of the solid base catalyst are shown in Table 1. The alumina support gave a surface area of 123 m2/g, and an mean pore size of 5.8 nm, In the case of the solid catalyst, the surface area calculated from the N2 adsorption− desorption isotherm was decreased to 19 m2/g, while the pore size was decreased to 3.8 nm. Anyway, an obvious decrease of

SBETa (m2/g)

Vpb (cm3/g)

DBJHc (nm)

Al2O3 KNO3/Al2O3

123 19

0.17 0.07

5.8 3.8

loading amount of KNO3 (h)

calcination temperature (°C)

stearoyl incorporation (%)

1 2 3 4 5 6 7 8 9 10 11

35 35 35 35 35 35 20 25 30 40 45

500 600 650 700 750 800 700 700 700 700 700

0 0 0 50.6 46.1 38.2 30.8 37.6 46.7 48.2 45.3

incorporation on the solid catalyst with different KNO3 loadings. As can be seen, the stearoyl incorporation achieved at the KNO3 loading of 20% was 30.8%, and then increased to the maximum value of 50.6% with increasing the KNO3 loading up to 35%. However, with subsequent increase in the KNO3 loading, the stearoyl incorporation remained nearly unchanged. In light of the results, the suitable KNO3 loading is determined to be 35%. In order to assess the effect of calcination temperature on the catalytic activity of the solid catalyst, the calcination temperature was set from 500 to 800 °C. The experimental results are also included in Table 2. As observed, the catalyst did not exhibit any activity in the transesterification reaction as the calcination temperature was below 700 °C. The maximum

Table 1. Textual Characteristics of Al2O3 and 35%KNO3/ Al2O3 Catalyst sample

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a

BET surface area. bPore volume. cAverage pore diameter from BJH desorption. D



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Table 3. Fatty Acid Composition (Percent) and Iodine Values of Lard, Soybean Oil and Their Blends in Different Ratios before and after Interesterification Reaction LA/SO ratio LA 80:20 80:20 60:40 60:40 40:60 40:60 20:80 20:80 SO

before after before after before after before after

myristic 14:0 1.8 1.3 1.4 1.4 1.2 1.0 1.1 0.4 0.5 0.0

± ± ± ± ± ± ± ± ± ±

palmitic 16:0 26.5 23.8 25.1 23.9 23.6 19.6 20.4 14.8 15.8 11.9

0.1 0.2 0.0 0.1 0.0 0.2 0.2 0.1 0.1 0.0

± ± ± ± ± ± ± ± ± ±

0.1 0.1 0.2 0.2 0.1 0.0 0.5 0.1 0.4 0.2

palmtoleic 16:1 2.5 2.1 2.6 2.0 1.7 1.5 1.4 0.6 0.8 0.0

± ± ± ± ± ± ± ± ± ±

stearic 18:0 12.3 11.7 12.2 8.9 11.2 7.3 7.8 6.3 7.0 3.4

0.2 0.2 0.2 0.1 0.2 0.0 0.0 0.1 0.2 0.0

± ± ± ± ± ± ± ± ± ±

0.2 0.1 0.1 0.5 0.2 0.2 0.1 0.2 0.0 0.2

oleic 18:1 38.7 39.6 39.6 33.4 33.9 36.7 35.4 28.1 28.1 22.2

± ± ± ± ± ± ± ± ± ±

linoleic 18:2 17.2 17.5 17.5 27.5 27.3 29.8 29.8 44.2 42.3 55.0

0.2 0.1 0.2 0.1 0.2 0.2 0.1 0.0 0.0 0.5

± ± ± ± ± ± ± ± ± ±

0.1 0.2 0.2 0.1 0.0 0.1 0.0 0.3 0.2 0.4

linolenic 18:3 0.7 1.6 1.6 2.8 2.1 3.9 3.9 5.6 5.0 7.6

± ± ± ± ± ± ± ± ± ±

0.1 0.1 0.5 0.1 0.1 0.1 0.0 0.1 0.2 0.1

IV 67.2 42.8 41.9 57.1 58.0 74.1 74.8 84.0 83.7 134.5

Table 4. Triacylglycerol Composition (Percent) of Lard, Soybean Oil, and Their Blends in Different Ratios before and after Interesterification Reaction TAG LLnLn LLLn LLL 0LL OOL OOO StOO StOL PLnLn PLLn POL PLL POO PStO PStSt MOL PPL PPO PPSt MPO

LA 0.0 0.0 0.0 0.7 1.8 3.3 3.6 4.7 0.0 0.4 8.9 13.7 22.8 12.8 3.2 1.5 8.6 9.7 1.8 1.3

80:20

60:40

40:60

20:80

SO

before

after

before

after

before

after

before

after

0.0 0.0 3.8 5.2 3.7 2.6 2.8 3.7 0.3 0.5 9.6 13.5 19.7 11.7 2.8 1.3 6.8 8.7 1.3 1.2

0.0 0.0 3.2 4.3 5.5 1.8 2.3 2.5 0.2 0.4 6.7 16.5 15.4 14.8 1.9 1.6 7.8 9.8 2.8 1.8

0.3 0.5 7.6 8.7 6.6 1.7 2.6 3.5 0.5 0.4 10.5 15.3 16.5 9.5 1.6 0.7 5.2 6.3 0.8 0.7

0.7 0.3 6.4 5.6 4.8 1.5 1.9 2.8 0.4 0.3 7.6 18.7 13.2 12.8 1.3 0.9 7.8 8.7 1.8 1.6

0.2 0.5 10.4 11.8 10.9 0.9 2.8 3.1 0.7 0.2 10.8 15.8 12.4 7.2 0.7 0.7 4.2 4.4 0.8 0.5

0.6 0.2 9.4 7.9 8.6 0.8 2.5 2.7 0.6 0.6 8.6 19.2 10.6 10.5 0.4 0.9 6.7 6.8 1.2 0.8

0.5 0.8 15.3 14.1 12.3 0.3 2.4 2.7 0.9 1.6 11.7 16.1 9.2 4.8 0.0 0.2 3.1 3.2 0.5 0.0

0.8 0.5 12.5 12.8 10.7 0.2 2.1 1.8 0.8 1.5 9.2 21.6 8.2 6.1 0.0 0.3 4.6 4.8 0.7 0.0

1.2 1.1 17.2 16.3 14.5 0.0 2.1 1.2 1.1 1.7 12.4 20.8 4.7 1.2 0.0 0.0 2.4 1.5 0.0 0.0

IV of the soybean oil was determined to be 134.5. Owing to large amounts of mono- and polyunsaturated FAs, soybean oil always exhibited a liquid state at room temperatures. The total FA profile and IV of lard and soybean oil are in agreement with those reported in the literature.29 As indicated in Table 3, there was no significant change in the total FA profile to be observed for the different blends of lard and soybean oil after the interesterification reaction. Moreover, the IV of the binary blends before and after the CIE reaction remained almost constant, implying that the interesterification did not affect the degree of saturation. Besides, the trans FAs were not detected in the interesterified blends catalyzed by the solid base catalyst, and the interesterification did not cause conjugation during the interesterification process. These results revealed that the total FA compositions of the binary blends were not significantly altered between the initial blend and the interesterified product. In spite of this, at the present study, the distribution pattern of FAs in the TAGs can influence the physicochemical and nutritional properties.

stearoyl incorporation of 50.6% was achieved at a calcination temperature of 700 °C. However, when the calcination temperature was further increased beyond 700 °C, there was an obvious decline trend in stearoyl incorporation to be observed. Accordingly, the best calcination temperature is 700 °C. Fatty Acid Composition and Iodine Value. The interesterification of lard and soybean oil was carried out using the solid catalyst prepared at a calcination temperature of 700 °C with a KNO3 loading of 35%. Table 3 shows the total FA compositions and the iodine values (IV) of lard, soybean oil and their binary blends before and after the interesterification. The total FA profile of lard was oleic acid (38.7%), palmitic acid (26.5%), linoleic acid (17.2%), stearic acid (12.3%) and palmitoleic acid (2.5%), with the IV of 67.2. The predominant FAs in the lard were accounted to be about 40% of long-chain saturated FAs, and accordingly at room temperature the lard was usually in solid state. As shown in Table 3, soybean oil had predominantly the following FA profiles: linoleic acid (55%), oleic acid (22.2%) and palmitic acid (11.9%), with small amounts of linolenic acid (7.6%) and stearic acid (3.4%). The E



Article

The FA compositions at sn-2 position for the 60:40 blend, before and after the CIE catalyzed using the solid catalyst, were investigated. The interesterification reaction did not occur without catalyst under the reaction conditions employed here. It was shown that palmitic acid, stearic acid, oleic acid, linoleic acid at sn-2 position for the 60:40 blend of lard and soybean oil were varied from 29.5%, 21.6%, 25.5% and 22.8% to 33.9%, 16.1%, 20.3% and 29.1%, respectively, after the CIE reaction. As a consequence, the FA compositions on the glycerol backbone were changed obviously after the interesterification reaction, and thus the solid catalyst was demonstrated to be active for the interesterification of soybean oil with lard. Triacylglycerol Composition. The TAG compositions of lard, soybean oil as well as their blends before and after the interesterification, are presented in Table 4. As observed, the main TAG compositions of lard were POO (22.8%), PLL (13.7%), PStO (12.8%), PPO (9.7%), POL(8.9%), PPL (8.6%), and StOL (4.7%). The soybean oil contained a wide range of unsaturated TAG species such as PLL (20.8%), LLL (17.2%), OLL (16.3%), OOL (14.5%), POL (12.4%), and POO (4.7%). As illustrated in Table 4, for different mass ratios of lard to soybean oil, different TAG species of the reaction mixtures were obtained. Moreover, the interesterification of the blends could result in a noticeable change in the TAG species as compared to their physical blends. For example, when the 60:40 blend of lard and soybean oil was interesterified, the interesterified product showed a reduced amount on POO, POL, OLL, LLL, OOL, and StOL from 16.5%, 10.5%, 8.7%, 7.6%, 6.6%, and 3.5% to 13.2%,7.6%,5.6%,6.4%, 4.8%, and 2.8%, respectively, but an increased amount of PLL, PStO, PPO,PPL, StOL, and PPSt from 15.3%, 9.5%, 6.3%,5.2%, and 0.8% to 18.7%,12.8%,8.7%,7.8%, and 1.8% was observed as a result of the CIE reaction. Similar phenomena were also reported previously.27 Indeed, it is difficult to evaluate quantitatively the progression of the interesterification process due to the complex compositions in the reaction mixture. According to the method described by De Clercq et al.,30 the degree of interesterification (DI) could be evaluated on the basis of TAG composition in the interesterified product. For the 80:20, 60:40, 40:60, and 20:80 blends of lard and soybean oil, the DI for the interesterification reaction was calculated to be 47.4%,76.8%, 81.4% and 43.6%, respectively. These obtained results suggested that the solid catalyst could efficiently catalyze the interesterification reaction. In the present investigation, to avoid hydrolysis reaction, free water was totally removed from the reaction mixture under reduced pressure prior to the interesterification process. Moreover, the acid value (AV) of the feedstocks was lowered below 0.1 mg KOH/g, since the free fatty acid could result in the deactivation of the solid base catalyst. Therefore, after the interesterification reaction, the undesirable byproducts, such as free fatty acids, monoglycerides, and diglycerides formed by the hydrolysis reaction, were not detected by TLC techniques. However, in the enzymatic interesterification catalyzed by immobilized lipase, such byproducts were formed during the reaction as water is essential to maintain the activity of enzyme.31 Slip Melting Point. In general, the melting properties vary with the TAG composition in the interesterified product, and the slip melting points (SMPs) of the samples are associated with their TAG compositions. The interesterification reactions between solid fat and liquid oil were conducted to obtain fat mixtures with better melting behavior. The SMP was

determined by the open capillary tube method, and the results are presented in Table 5. It was shown that the SMPs of the Table 5. Slip Melting Point of Binary Soybean Oil and Lard Blends before and after Interesterification Reaction LA/SO

SMP (°C) before CIE

80:20 60:40 40:60 20:80

44.1 39.3 33.4 26.2

± ± ± ±

0.1 0.1 0.1 0.0

after CIE 38.6 34.5 30.4 23.1

± ± ± ±

0.1 0.1 0.0 0.1

physical blends and the interesterified blends were significantly different, and the proportion of lard to soybean oil had significant effect on the SMP. As observed, these physical blends showed SMPs ranging from 26.2 to 44.1 °C, and the SMP was decreased with an increase in the proportion of soybean oil in the blends mostly due to the increase of the unsaturated FAs. Notably, the interesterified blends possessed lower SMPs for all samples when compared with their physical blends, which could be regarded as another evidence for the catalyst activity. The interesterified fat had an SMP below 37 °C, showing that it could melt almost completely at body temperature. This observation has demonstrated that the heterogeneous interesterification could give an attractive method for producing new tailored-fats with desired melting profiles that meet consumer preferences. Crystal Structure and Crystal Morphology. Crystal properties can greatly influence the consistency and acceptability of the final product. Small crystals result in firmer products, whereas large crystals yield a sandy mouthfeel.32 Figure 4 shows the crystal microstructures of the selected samples. As can be seen, soybean oil displayed no any distinct crystal at a temperature of 25 °C, while lard contained a dense network of nearly globular collections of clustered crystals. For the physical blend of lard and soybean oil, the sample exhibited small spherulite and needle-shaped like structure with no evident regular pattern, which was probably resulted from a dilution of the lard microstructure. Compared with the physical blend, the interesterified sample showed a very small crystal size, less aggregation, no large crystals, and a more homogeneous distribution of the crystals. As a result, the interesterification of soybean oil with lard led to not only a decrease in the size of the crystal, but also a change in the morphology with a network of small crystal. Small crystals are necessary for margarine fats and shortenings, owing to their stabilization of air bubbles to food products. The change in textural properties can be explained by the variation in TAG compositions observed for the interesterification. The TAG composition can be responsible for arrangement of different kinds of crystals, which strongly influences the product texture. In conclusion, an environmentally friendly catalyst based on alumina impregnated with potassium was prepared by the conventional incipient-wetness impregnation method. The prepared solid catalyst was demonstrated to be an efficient solid base catalyst for the interesterification reactions. The HPLC analysis of the TAGs and analysis of the sn-2 positional FA compositions showed that the CIE reaction could lead to the rearrangement of FAs in the TAGs. After the interesterification, the SMPs and crystal morphology of the interesterificated products were varied substantially. The solid catalyst has several advantages of green or sustainable chemistry for F



Article

Figure 4. Crystal morphology of samples: (a) soybean oil; (b) lard; (c) physical 60:40 blend of lard and soybean oil; and (d) interesterificated 60:40 blend of lard and soybean oil. nutritive shortenings produced from regioselective hardening of soybean oil with Pt containing zeolite. J. Am. Oil Chem. Soc. 2011, 88, 2023−2034. (2) Xu, X. Production of specific-structured triacylglycerols by lipasecatalyzed reactions: A review. Eur. J. Lipid. Sci. Techol. 2000, 102, 287− 303. (3) Lee, J. H.; Akoh, C. C.; Himmelsbach, D. S.; Lee, K. T. Preparation of interesterified plastic fats from fats and oils free of trans fatty acid. J. Agric. Food Chem. 2008, 56, 4039−4046. (4) Etherton, P. K. Trans fatty acid and coronary heart diseases risk. Am. J. Clin. Nutr. 1995, 62, 655S−708S. (5) Dhaka, V.; Gulia, N.; Ahlawat, K. S.; Khatkar, B. S. Trans fatssources, health risks and alternative approach: a review. J. Food Sci. Technol. 2011, 48, 534−541. (6) Ferrari, R. A.; Esteves, W.; Mukherjee, K. D. Alteration of steroyl ester content and positional distribution of fatty acids in triacylglycerols by chemical and enzymatic interesterification of plant oils. J. Am. Oil Chem. Soc. 1997, 74, 93−96. (7) Pacheco, C.; Palla, C.; Crapiste, G. H.; Carrín, M. E. Optimization of reaction conditions in the enzymatic interesterification of soybean oil and fully hydrogenated soybean oil to produce plastic fats. J. Am. Oil Chem. Soc. 2013, 90, 391−400. (8) Ribeiro, A. P. B.; Grimaldi, R.; Gioielli, L. A.; Goncalves, L. A. G. Zero trans fats from soybean oil and fully hydrogenated soybean oil: physico-chemical properties and food applications. Food Res.Int. 2009, 42, 401−410. (9) Jeyarani, T.; Reddy, S. Y. Effect of enzymatic interesterification on physicochemical properties of mahua oil and kokum fat blend. Food Chem. 2010, 123, 249−253. (10) Xu, X.; Skands, A. R. H.; Høy, C. E.; Mu, H.; Balchen, S.; AdlerNissen, J. Production of specific-structured lipids by enzymatic interesterification: elucidation of acyl migration by response surface design. J. Am. Oil Chem. Soc. 1988, 75, 1179−1186. (11) Yang, T.; Fruekilde, M. B.; Xu, X. Applications of immobilized Thermomyces lanuginosa lipase in interesterification. J. Am. Oil Chem. Soc. 2003, 80, 881−887.

catalyst recovery, high catalytic activity, and recyclability of the solid catalyst.

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AUTHOR INFORMATION

Corresponding Author

*Tel.: +86 371 67756302; fax: +86 371 67756718; e-mail: [email protected]. Funding

This work was financially supported by research grants from the National Natural Science Foundation of China (Project No. 21276066, 21476062), the Plan for Scientific Innovation Talent of Henan Province (144200510006) and the Program for Innovative Research Team in Universities of Henan Province in China (2012IRTSTHN009). Notes

The authors declare no competing financial interest.

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ABBREVIATIONS USED AOCS, American Oil Chemists’ Society; BET, Brunauer− Emmett−Teller; BJH, Barrett−Joyner−Halenda; CIE, chemical interesterification; EIE, enzymatic interesterification; FA, fatty acid; FAME, fatty acid methyl ester; FT-IR, Fourier transform infrared; GC, gas chromatography; HPLC, high-performance liquid chromatography; IV, iodine value; L, linoleic acid; LA, lard; Ln, linolenic acid; 2-MAG, 2-monoacylglycerol; O, oleic acid; P, palmitic acid; SEM, scanning electron microscopy; PPL, porcine pancreatic lipase; SMP, slip melting point; SO, soybean oil; St, stearic acid; TAG, triacylglycerol; TLC, thinlayer chromatography; XRD, power X-ray diffraction

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Heterogeneous Interesterification of Triacylglycerols Catalyzed by

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