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An investigation on pretreatments for inactivation of lipase in naked oat kernels using microwave heating
Journal of Food Engineering 95 (2009) 280–284
Contents lists available at ScienceDirect
Journal of Food Engineering journal homepage: www.elsevier.com/locate/jfoodeng
An investigation on pretreatments for inactivation of lipase in naked oat kernels using microwave heating Qian Keying a,1, Ren Changzhong a,b,1, Li Zaigui a,* a b
Lab. of Cereal Science, China Agricultural University, Beijing 100083, PR China Engineering and Technology Research Center of Oats, Baicheng Academy of Agricultural Sciences, Baicheng City, Jilin 137000, PR China
a r t i c l e
i n f o
Article history: Received 26 July 2008 Received in revised form 11 March 2009 Accepted 3 May 2009 Available online 8 May 2009 Keywords: Naked oat kernels Microwave heating Lipase activity Inactivation Steam quantity
a b s t r a c t To investigate the primary factors affecting the efficiency of microwave heating pretreatments in inactivating lipase in naked oat kernels, the effects of temperature, duration of heating, moisture content of oat kernels, tempering time and packaging conditions on the inactivation of lipase activities were examined. Microwave heat treatment for 45 s was found to inactivate 98–99% lipase in oat kernels (20 g in polyethylene package) with 11.1% moisture content, thereby stabilizing oat kernels against triglyceride lipolysis. The steam quantity played a crucial role in lipase inactivation. A trend was found: the higher the quantity of steam evaporated, the lower the residual lipase activities remained in whole kernels. Pretreatments, such as vacuum packaging, increasing the moisture content of oat kernels to a range of 17.0–25.0% from 11.1%, and conducting heat treatments as soon as the kernel moisture contents were adjusted, were found to be beneficial in accelerating steam evaporation; the efficiency of microwave heating on lipase inactivation was thereby increased. Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction Oats originated in China, of which 95% are naked oats (Avena nuda L.), the other 5% are hulled oats. They have a similar nutritional component as hulled oats and also contain higher fat content than other cereal grains. The fat content in 90% of 4000 entries in the world oat collection is 5.0–9.0% (Youngs, 1978), while that of 710 entries of Chinese oat kernels is 3.7–9.4%, with most of them being above 5.0% (Gong et al., 1999). Oats also have a higher lipase activity than any other cereals (O’Connor et al., 1992). Since triglycerides in whole kernels comprise about 78% of total lipids, the increase of free fatty acids at the expense of triglycerides catalyzed by lipase is the primary difficulty in the application of oat in new food. Prevention of lipid hydrolysis is the main goal in the manufacturing of oat products (Liukkonen et al., 1992). In effect, lipid content in intact oat kernels stored for 1 year at room temperature is quite stable (Hutchinson and Martin, 1951), for the protection of endogenous antioxidants, such as tocopherols, Lascorbic acid, thiol, phenolic amino acid and phenol compounds. Though some of these chemicals are heat sensitive under commercial heat treatment (Bryngelsson et al., 2002), some research also showed that heat treatment at 100 °C for 30 min did not significantly reduce the phenolic content in oats (Zadernowski et al., 1999). Even microwave heating at 150 °C for 15 min significantly * Corresponding author. Tel.: +86 10 6273 7293; fax: +86 10 6273 7392. E-mail address: [email protected] (L. Zaigui). 1 Authors with equal contributions. 0260-8774/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.jfoodeng.2009.05.002
increased the phenolic content and antioxidant activity of oat bran extracts (Stevenson et al., 2008). However, rather than the antioxidants, any damage brought about by processing to the oat intact kernels will instantly start the hydrolysis, due to a highly increased exposure of lipid to the lipase. In this way, any residual lipase can affect the storage quality of the ground oat meal and can cause offflavors such as soapiness and bitterness in oatcakes and biscuits made from oatmeal dough (Ekstrand et al., 1992). It is therefore generally considered necessary to inactivate the lipases completely. Up to date, however, steaming and/or kilning are still the typical industrial pretreatments, which require temperature up to 95–100 °C for 1.5–2.0 h per batch and pose difficult working conditions (Ganbmann and Vorwerck, 1995). One of the research focuses is to seek for an alternative, efficient and energy-saving technique of lipase inactivations. Microwave heat treatment varying in severity has been able to inactivate lipase and lipoxygenase in cereal bran, germ, soybean (Vetrimani et al., 1992; Jiaxun et al., 1993), groundnuts (Ramesh et al., 1995), rapeseeds (Ponne et al., 1996) and olive oil (Farag et al., 1997). Heating with domestic microwave oven has also been reported to be effective in extending the shelf-life of rice bran by reducing about 90% of increasing rate of free fatty acids (FFAs) in 3-month (Ramezanzadeh et al., 1999) and 4-month (Faiyaz Ahmed, 2007) storage. Further, it has also been reported that the inactivation of rice bran lipase depends upon temperature, duration of heat treatment, and moisture content, the latter being critical (Jiaxun et al., 1993). Earlier research (Hutchinson and Martin, 1951) has also shown that to inactivate 97–98% of lipase, oat kernels with a
Q. Keying et al. / Journal of Food Engineering 95 (2009) 280–284
moisture content of 6, 8, 10, 12, 14, 16, 18 or 20% should be roasted for 1 h under 120, 104, 92, 83, 76, 71, 67 or 64 °C, respectively. Although microwaving has been applied to many cereals and other food materials, and does serve rapid and efficient heating for it requires no heat transmission or loss of heat medium (e.g., steam in steaming), it has seldom been applied to oat processing. In Grant’s experimental setup, 130 g of whole oats was heated in a 22.5 cm tall, 5 cm wide glass cylinder with four 0.3 cm holes randomly placed around the cylinder and covered with a glass lid. Samples were treated for 180 s to a final temperature of 108 °C using a microwave oven (1500 W, 2450 MHz) and the lipase activity was not significantly inactivated (Grant, 1999). This was attributed to the continuous moisture loss during microwave treatment. However, the lipase inactivation by microwave heat treatment, as well as the role of moisture in the process, has not been investigated in previous researches. This work examines the feasibility of microwave heating for inactivating lipase in oat kernels. The effects of moisture content of oat kernels, heating time, temperature, tempering time and packaging on the extent of inactivation are investigated.
281
ene bag (6.5 5.5 1.3 cm3). Sample bags were placed in the center of the revolving glass tray and heated by a domestic microwave oven (Glanz, 2450 MHz, 800 W, Foshan Shunde Galanz Microwave Oven Electrical Appliance Ltd.) for a period of 10– 45 s, respectively, with a starting temperature of 19–22 °C. The center temperature of the packages (with oat kernels sealed inside) by the end of heating was immediately measured in triplicates using a non-contact infrared thermometer (DT-8811H, CEM Shenzhen Ever-best Machinery Industry Co., Ltd.). Treated oat kernels were cooled for 1 min at room temperature (19–22 °C), then poured out from each bag onto a paper tray and air-dried at 40 °C for 1 h. Dried kernels were then ground using a high-speed pulverizer (HY-04 B, Beijing Huanya Tianyuan Machinery technology Co., Ltd.) until over 90% of the oat flour had a particle size of less than 0.425 mm (>90% could pass through a 40mesh sieve). Oat flour obtained was taken out (3 g), packed with a filter paper and defatted by Soxhlet extraction for 6 h using diethyl ether. Defatted flour was dried at 40 °C for 1 h and then stored at 18 °C for the determination of lipase activity. 2.5. Effect of moisture content and tempering time on lipase inactivation
2. Materials and methods 2.1. Materials Oats (Huawan No. 6) cultivated in Zhangjiakou, Hebei province, China, were harvested in September 2007. The whole oat kernels were stored at 10 °C in a sealed polyethylene bag after threshing and removing husked kernels (as contaminants in naked oat, about 5%), fragmentary kernels, dirt plus stone. 2.2. Moisture The moisture content of the oat flour was determined by AACC44-15A [1999] method, using a hot-air oven. Ground oat flour samples (2–3 g) were dried at 130 ± 1 °C for 1 h, and percentage weight loss was calculated as the moisture content.
The moisture contents of the oat kernels were adjusted by adding distilled water to 13.0, 17.0, 20.0 and 25.0% from an initial value of 11.1%, respectively, thoroughly mixed for 10 min, then sealed with a polyethylene film in stainless steel basins and stored at room temperature for tempering. The basins were shaken occasionally for an even moisture distribution. At different tempering times (0, 4, 7, 11 and 28 h), 20.00 g oat kernels of each moisture content level were weighted, respectively, either vacuum packed in a polyethylene bag (6.5 5.5 1.3 cm3) or evenly spread on a coverless culture dish, and treated with microwave heating for 25 s followed by a 10 min cooling at room temperature. The residual lipase activities were analyzed. Each treatment and lipase activity measurement was conducted in duplicate. Oat kernels (M) which had been heated in culture dishes were weighed (M) after cooling for 10 min and the moisture loss was calculated as below:
2.3. Lipase activity
Moisture lossð%Þ ¼ Residual lipase activity was determined in 0.5 g samples using the dough method (Peterson, 1999). To each sample, 98 lL (six drops with 100 lL pipette) of glycerol trioleate (CP, 98–103%, Sinopharm Chemical Reagent Co., Ltd.) was added followed by 330 lL of buffer (0.05 M Tris–HCl, pH 7.5, containing 1% v/v Triton X-100) in to a screw-cap cuvette. The dough samples were mixed thoroughly with a glass rod and incubated at 37 °C for 1 h. The reaction was stopped by adding 100 lL of 1 M HCl and again mixing with a glass rod. For the control, HCl was added immediately. To extract oleic acid hydrolyzed from the glycerol trioleate, 5 mL of hexane was added and tightly sealed with a cap. The cuvettes were put into a boiling water bath for 5 min, and subjected to vortex shaking for 1 min to fully extract oleic acid into hexane, and then centrifuged (3000 r/min, 10 min). Four milliliter supernatant was mixed with 1 mL of copper reagent and the resulting fatty acid copper soap had a strong absorbance at 715 nm (Kwon and Rhee, 1986). The absorbance measured at 715 nm was then compared with an oleic acid standard. The lipase activity was expressed as lmol of oleic acid liberated per hour per gram of the sample. 2.4. Effect of packaging on lipase inactivation In order to study the influence of vacuum and normal packaging on the effects of lipase inactivation by microwave heating, oat kernels (20.0 g) were either sealed or vacuum packed in a polyethyl-
20:00 M 100 20:00
ð1Þ
2.6. Statistical analysis All the data are means of duplicate assays. The results were analyzed using Wilcoxon rank sum test for paired data of lipase activities with vacuum and normal packaging. Lipase activities affected by moisture and tempering time were analyzed by two-way analysis of variance. Linear regression analysis was performed between residual lipase activity and the moisture loss after 25 s microwave heating in coverless culture dishes.
3. Results and discussion 3.1. Effect of packaging on lipase inactivation by microwave heating Fig. 1 describes the variation in the residual lipase activities of the oat kernels in either sealed or vacuum packages after heat treatments varying in duration. For oat kernels in vacuum packages, with heating time prolonged, the residual lipase activity decreased slowly at first (0–25 s), and then sharply to 6.7 ± 0.5 lmol FFA h1 g1 in a period of 25–40 s. A final residual lipase activity of 2.8 ± 2.7 lmol FFA h1 g1 (about 1–2% of initial activity, 352.8 ± 3.6 lmol FFA h1 g1) remained after 45 s heating.
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Q. Keying et al. / Journal of Food Engineering 95 (2009) 280–284
450
Activity (nor.) Temp. (nor.)
The results of residual lipase activities of both packaging were analyzed with Wilcoxon rank sum test for paired data, and the results showed P < a (0.05), which also confirmed that vacuum packaging could inactivate lipase with ease during microwave heating.
160 140
350 120 300 100 250 y = 3.0463x + 10.537
80
2
200
R = 0.9644 60
150 100
40
y = 2.809x + 4.1033 2
R = 0.9963
50 0 0
Ending temperature (°C)
Lipase activities (µmol•h-1•g-1)
400
Activity (vac.) Temp. (vac.)
20 0
5
10
15
20
25
30
35
40
45
Microwave heating time (s) Fig. 1. The variation of lipase activity and final temperatures with heating time in normal or vacuum packages (nor.: normal package, vac.: vacuum package). (Note: increasing rate of temperature was compared in the same period of 10–30 s).
After heat treatments varying in duration, the levels of residual activities in oat kernels heated in normal packages were all slightly higher than the corresponding levels of kernels in vacuum packages. Generally, a similar decreasing trend in lipase activity was found in both the packages, as well as a similar final level of 1–2% of initial activity after 45 s heating. In contrast, for normal package, residual lipase activity decreased slightly at first, but then increased gradually to the initial level when heated for 25 s, while that in vacuum package shows a continuous decrease. Lipoxygenase activity in groundnut seeds has been reported to be activated during microwave heating. The lipoxygenase activities in groundnut seeds (500 g) increased by about 16% when the seeds were subjected to microwave treatment (at 750 W in a laboratory microwave oven (Philips, Model M710 cooktronic) for 60 s, before a continuous decrease for even longer periods ranging from 60 to 300 s (Ramesh et al., 1995). The final temperature in the center of package surface for both packing methods is compared as shown in Fig. 1. As the heating time increased, the final temperature in vacuum and normal packages increased at a rate of 3.0 °C/s (R2 = 0.96, from 44 °C to 105 °C) and 2.8 °C/s (R2 = 0.99, from 33 °C to 88 °C) in a period of 10–30 s, and reached the point of rapid steam generation after heating for 30 and 40 s, respectively. Unlike the case of normal packages, for oats in vacuum packages, lipase activities declined continuously without any increase during 15–25 s heating. This might be due to the earlier and faster steam generation in vacuum packages at the beginning of heat treatment. In vacuum packages (e.g. absolute pressure 0.02 bar) and normal packages (absolute pressure 1 bar), moisture boils at 17.51 and 100 °C, respectively; thus much higher energy is needed to heat the same quantity of moisture from room temperature to its evaporating temperature. Therefore, compared with normal packages, the depletion of pressure in vacuum packages increases the fugacity of moisture within the packages and enables the evaporation of low temperature steam. The generations of low temperature steam increased the density of moisture inside the package and accelerated the transformation of microwave energy into thermal energy. Consequently, the increasing rate of temperature is slightly higher (0.2 °C/s in the heating period of 10–30 s) in vacuum packages and heated steam may counteract the microwave activation during the heating period of 15–25 s.
3.2. Effects of moisture content and tempering time on lipase inactivation To study the influences of moisture content and tempering time on lipase inactivation, the residual lipase activities of oat kernels with different moisture levels (13–25%) after 25 s of microwave heating at different tempering times were analyzed (Fig. 2). Fig. 2a indicates the variation in lipase activities after heating for 25 s by microwave in coverless culture dishes. The higher the moisture of oats kernels, the lower the residual lipase activity remained after heating for the same period of time. With increase in the tempering time, residual lipase activities of oat kernels with different moisture levels all increased sharply and then reposefully in a period of 0–4 h and 4–28 h after the moisture adjustment, respectively. This is mainly due to the variation in the distribution of adjusted moisture. Most moisture added stayed at the surface and outer layer of kernels just after tempering; this induced the water molecules to have more opportunity to collide with each other and vaporize. With increase in the tempering time, it was difficult to vaporize the water molecules entering the kernels, and moisture transfer rate tended to decrease; thereby the final temperature and the amount of moisture loss were also found to decrease fast at first and slowly later on.
Fig. 2. Lipase activities in oats with different moisture levels after 25 s microwave heating at different tempering times.
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Q. Keying et al. / Journal of Food Engineering 95 (2009) 280–284 Table 1 Two-way analysis of variance of the residual lipase activities after 25-s microwave heating at different moisture levels and tempering times. Moisture content/%
Residual lipase activity/lmol h1 g1
Tempering time/h
Residual lipase activity/lmol h1 g1
Effect factors
Pr > F
25 20 17 13
19.70 d 49.76 c 75.73 b 196.40 a
0 4 7 11 28
43.52 b 98.71 a 83.61 a 99.83 a 101.31 a
Moisture content (a) Tempering time (b) Interaction (a b)
<0.0001 0.0003 0.0180
Means of the same column followed by different letters differ significantly (P < 0.05), according to Duncan’s New Multiple Range Test.
Fig. 2b shows the changes in residual lipase activity of samples after 25 s of microwave heating treatments using vacuum packing. Lipase activities in oat kernels with 17–25% moisture content were inactivated by 98–99% when heat treatment was conducted immediately after moisture adjustment. Similarly, higher levels of residual lipase activity remained with the tempering time increased. Nevertheless, lower residual lipase activity remained in oat kernels using vacuum packages in comparison with that in coverless culture dishes, mainly due to the retention of the steam that was generated in vacuum packages during heating. Table 1 shows the results of a two-way analysis of variance of the lipase activities in oats with different moisture levels after 25 s of microwave heating at different tempering times. The results indicated that the interaction of moisture content and tempering time had a significant effect on the lipase inactivation using microwave heating. This implies that both the factors affected the efficiency of lipase inactivation, mainly due to their correlation with the steam generation: the increase in the moisture content accelerated the steam generation, and the extension in tempering time made the release of moisture difficult as explained above. To confirm this conclusion, the effect of steam quantity on lipase inactivation was further studied by analyzing the correlation between the moisture loss and the residual lipase activities after 25 s heating under different moisture levels at different tempering times. Steam quantity during heating was denoted by the ratio of moisture loss, which was expressed as percentage (%) of moisture loss in every 20.00 g of oat samples (wet based). As shown in Fig. 3, with increase in the tempering time, the ratio of moisture loss decreased dramatically in a period of 0–4 h after moisture adjustment and decreased slowly later on, just the opposite of the changes in residual lipase activities as shown in Fig. 2a. The decrease in the moisture loss of oat kernels with the same moisture level was mainly caused by the prolongation of tempering time when more moisture was transferred from outer to inner layer, which consequently prevented the steam generation and release during heating by microwave.
A significant negative correlation (R2 = 0.9252) was found using the linear regression analysis between residual lipase activity of oat kernels (in coverless culture dishes) and the moisture loss after 25 s of microwave heating when the amount of moisture loss was fairly large (over 3%) (as shown in Fig. 4). No significant linear correlation (R2 = 0.6612) between the moisture loss and the residual lipase activity was found when the quantity of steam was not enough to inactivate the lipase (moisture loss <3%). Moreover, lower moisture loss from oat kernels with a higher moisture content than that with a lower moisture content was found when the tempering time of the former was much longer than that of the latter, and vice versa (as shown in Fig. 3). This suggested that the steam quantity expressed as moisture loss, rather than the moisture content, was the critical factor in the inactivation of lipase. However, the combination of increasing moisture content to 17–25%, conducting heating treatments immediately after a moisture adjustment procedure, and using vacuum packaging could promote the steam release and inactivate the lipase more efficiently. In addition to the steam quantity, temperature is another important factor affecting lipase inactivation. The interaction of temperature and moisture played an important role in the traditional thermal inactivation treatments as reported. For example, 1 h of kilning at 120 and 64 °C was required to inactivate 97–98% of lipase in oat kernels with 20% and 6% moisture (Hutchinson and Martin, 1951). As shown in Table 2, the higher the end temperature was, the lower the residual lipase activity remained at the same tempering time (in the same row) and same moisture content (in the same column). However, no significant correlation between temperature and residual lipase activity was found in this experiment. The possibility of inactivating lipase completely by microwave heating was examined using the maximal heating duration (49 s, up to the pressure tolerance of the package) and the heating treatment was conducted immediately after moisture adjustment to 20%. Residual lipase activity was 0.3 ± 0.3 lmolh1 g1. This suggested a complete lipase inactivation. In addition, the increase of free fatty acids (FFAs) in these microwave-treated oat kernels was even less than 0.5-fold of the initial value (<30 mg KOH/
Fig. 3. Moisture loss in oat kernels with different moisture levels after 25 s microwave heating in coverless culture dishes at different tempering times.
Fig. 4. The correlation between residual activity and moisture loss of oats after 25 s microwave heating (in coverless culture dishes).
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Table 2 The final temperature (°C) and residual lipase activity (lmol h1 g1) after 25-s microwave heating. Tempering time/h
0 4 7 11 28
Moisture content (%) 25
20
17
13
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
105.9 ± 0.9 103.5 ± 1.2 104.0 ± 0.3 105.0 ± 0.0 101.3 ± 0.3
3.3 25 17.1 24 29.2
102.3 ± 1.3 101.2 ± 0.5 100.7 ± 3.0 99.0 ± 2.7 100.0 ± 0.0
4.2 75.5 44.1 94.4 30.6
102.3 ± 1.3 93.5 ± 2.2 98.8 ± 0.2 94.0 ± 1.7 98.2 ± 1.2
14.9 115.7 73.6 98.7 75.8
94.8 ± 4.2 91.7 ± 3.7 93.3 ± 1.3 92.8 ± 0.8 84.7 ± 8.3
151.7 178.8 199.7 182.2 269.7
100 g oat flours, dry-based), whereas in raw oat kernels up to 18fold increase was found, after 1-month storage at 40 °C and 80% relative humidity. 4. Conclusions Steam quantity was a critical factor in lipase inactivation by microwave heating. Microwave heating was able to stabilize the oat flour using a sufficient heating process. Appropriately increasing moisture content of oat kernels to 17–25%, conducting microwave heating treatment immediately after adjusting moisture, and using vacuum packaging could promote the steam release and inactivate the lipase completely. Modification of these conditions was required before they can be applied on a continuous and commercial scale processing. Reusable high-pressure containers may substitute for the disposable polyethylene pouches. The similarity of the role played by steam quality in lipase inactivation both by microwave and steaming provides an interesting topic for a future research. The effects of these two types of treatments on the variation of lipase activity, FFA values during storage and sensory property of oat foods would be investigated in our future work. References Bryngelsson, S., Dimberg, L.H., Kamal-Eldin, A., 2002. Effects of commercial processing on levels of antioxidants in oats (Avena sativa L.). Journal of Agricultural and Food Chemistry 50 (7), 1890–1896. Ekstrand, B., Gangby, I., Akesson, G., 1992. Lipase activity in oats-distribution, pH dependence, and heat inactivation. Cereal Chemistry 69 (4), 379–381. Faiyaz Ahmed, K.P.S.V.S.P.K.S., 2007. Improved shelf-life of rice bran by domestic heat processing and assessment of its dietary consumption in experimental rats. Journal of the Science of Food and Agriculture 87 (1), 60–67. Farag, R.S., El-Baroty, G., Abd-El-Aziz, N., Basuny, A.M., 1997. Stabilization of olive oil by microwave heating. International Journal of Food Sciences and Nutrition 48 (6), 365–371.
Ganbmann, W., Vorwerck, K., 1995. Oat milling, processing and storage. In: The Oat Crop. Chapman and Hall, London, pp. 369–408. Gong, H., Li, C., Wang, Y., 1999. Analysis of oat germplasm quality characters. Journal of Shanxi Agricultural Sciences 27 (2), 16–19. Grant, P.J.A., 1999. Different heating techniques for oat lipase stabilization. Lincoln,Nebraska, University of Nebraska. Doctor: 159. Hutchinson, J.B., Martin, H.F., 1951. Location and destruction of lipase in oats. Nature 167 (4254), 758–759. Jiaxun, T., Rao, R., Liuzzo, J., 1993. Microwave heating for rice bran stabilization. Journal of Microwave Power and Electromagnetic Energy 28 (3), 156–164. Kwon, D.Y., Rhee, J.S., 1986. A simple and rapid colorimetric method for determination. Journal of American oil chemists society 63 (1), 89–92. Liukkonen, K.H., Montfoort, A., Laakso, S.V., 1992. Water-induced lipid changes in oat processing. Journal of Agricultural and Food Chemistry 40 (1), 126– 130. O’Connor, J., Perry, H.J., Harwood, J.L., 1992. A comparison of lipase activity in various cereal grains. Journal of Cereal Science 16 (2), 153–163. Peterson, D.M., 1999. Lipase activity and lipid metabolism during oat malting. Cereal Chemistry 76 (1), 159–163. Ponne, C.T., Moeller, A.C., Tijskens, L.M.M., Bartels, P.V., Meijer, M.M.T., 1996. Influence of microwave and steam heating on lipase activity and microstructure of rapeseed (Brassica napus). Journal of Agricultural and Food Chemistry 44 (9), 2818–2824. Ramesh, M., Haridas Rao, P., Ramadoss, C.S., 1995. Microwave treatment of groundnut (Arachis hypogaea): extractability and quality of oil and its relation to lipase and lipoxygenase activity. Lebensmittel Wissenschaft und Technologie 28 (1), 96–99. Ramezanzadeh, F.M., Rao, R.M., Windhauser, M., Prinyawiwatkul, W., Tulley, R., Marshall, W.E., 1999. Prevention of hydrolytic rancidity in rice bran during storage. Journal of Agricultural and Food Chemistry 47 (8), 3050– 3052. Stevenson, D.G., Inglett, G.E., Chen, D.J., Biswas, A., Eller, F.J., Evangelista, R.L., 2008. Phenolic content and antioxidant capacity of supercritical carbon dioxide-treated and air-classified oat bran concentrate microwave-irradiated in water or ethanol at varying temperatures. Food Chemistry 108 (1), 23–30. Vetrimani, R., Jyothirmayi, N., Rao, P.H., Ramadoss, C.S., 1992. Inactivation of lipase and lipoxygenase in cereal bran, germ and soybean by microwave treatment. Lebensmittel-Wissenschaft und Technologie 25 (6), 532–535. Youngs, V.L., 1978. Oat lipids. Cereal Chemistry 55 (5), 591–597. Zadernowski, R., Nowak-Polakowska, H., Rashed, A.A., 1999. The influence of heat treatment on the activity of lipo- and hydrophilic components of oat grain. Journal of Food Processing and Preservation 23 (3), 177–191.
Contents lists available at ScienceDirect
Journal of Food Engineering journal homepage: www.elsevier.com/locate/jfoodeng
An investigation on pretreatments for inactivation of lipase in naked oat kernels using microwave heating Qian Keying a,1, Ren Changzhong a,b,1, Li Zaigui a,* a b
Lab. of Cereal Science, China Agricultural University, Beijing 100083, PR China Engineering and Technology Research Center of Oats, Baicheng Academy of Agricultural Sciences, Baicheng City, Jilin 137000, PR China
a r t i c l e
i n f o
Article history: Received 26 July 2008 Received in revised form 11 March 2009 Accepted 3 May 2009 Available online 8 May 2009 Keywords: Naked oat kernels Microwave heating Lipase activity Inactivation Steam quantity
a b s t r a c t To investigate the primary factors affecting the efficiency of microwave heating pretreatments in inactivating lipase in naked oat kernels, the effects of temperature, duration of heating, moisture content of oat kernels, tempering time and packaging conditions on the inactivation of lipase activities were examined. Microwave heat treatment for 45 s was found to inactivate 98–99% lipase in oat kernels (20 g in polyethylene package) with 11.1% moisture content, thereby stabilizing oat kernels against triglyceride lipolysis. The steam quantity played a crucial role in lipase inactivation. A trend was found: the higher the quantity of steam evaporated, the lower the residual lipase activities remained in whole kernels. Pretreatments, such as vacuum packaging, increasing the moisture content of oat kernels to a range of 17.0–25.0% from 11.1%, and conducting heat treatments as soon as the kernel moisture contents were adjusted, were found to be beneficial in accelerating steam evaporation; the efficiency of microwave heating on lipase inactivation was thereby increased. Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction Oats originated in China, of which 95% are naked oats (Avena nuda L.), the other 5% are hulled oats. They have a similar nutritional component as hulled oats and also contain higher fat content than other cereal grains. The fat content in 90% of 4000 entries in the world oat collection is 5.0–9.0% (Youngs, 1978), while that of 710 entries of Chinese oat kernels is 3.7–9.4%, with most of them being above 5.0% (Gong et al., 1999). Oats also have a higher lipase activity than any other cereals (O’Connor et al., 1992). Since triglycerides in whole kernels comprise about 78% of total lipids, the increase of free fatty acids at the expense of triglycerides catalyzed by lipase is the primary difficulty in the application of oat in new food. Prevention of lipid hydrolysis is the main goal in the manufacturing of oat products (Liukkonen et al., 1992). In effect, lipid content in intact oat kernels stored for 1 year at room temperature is quite stable (Hutchinson and Martin, 1951), for the protection of endogenous antioxidants, such as tocopherols, Lascorbic acid, thiol, phenolic amino acid and phenol compounds. Though some of these chemicals are heat sensitive under commercial heat treatment (Bryngelsson et al., 2002), some research also showed that heat treatment at 100 °C for 30 min did not significantly reduce the phenolic content in oats (Zadernowski et al., 1999). Even microwave heating at 150 °C for 15 min significantly * Corresponding author. Tel.: +86 10 6273 7293; fax: +86 10 6273 7392. E-mail address: [email protected] (L. Zaigui). 1 Authors with equal contributions. 0260-8774/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.jfoodeng.2009.05.002
increased the phenolic content and antioxidant activity of oat bran extracts (Stevenson et al., 2008). However, rather than the antioxidants, any damage brought about by processing to the oat intact kernels will instantly start the hydrolysis, due to a highly increased exposure of lipid to the lipase. In this way, any residual lipase can affect the storage quality of the ground oat meal and can cause offflavors such as soapiness and bitterness in oatcakes and biscuits made from oatmeal dough (Ekstrand et al., 1992). It is therefore generally considered necessary to inactivate the lipases completely. Up to date, however, steaming and/or kilning are still the typical industrial pretreatments, which require temperature up to 95–100 °C for 1.5–2.0 h per batch and pose difficult working conditions (Ganbmann and Vorwerck, 1995). One of the research focuses is to seek for an alternative, efficient and energy-saving technique of lipase inactivations. Microwave heat treatment varying in severity has been able to inactivate lipase and lipoxygenase in cereal bran, germ, soybean (Vetrimani et al., 1992; Jiaxun et al., 1993), groundnuts (Ramesh et al., 1995), rapeseeds (Ponne et al., 1996) and olive oil (Farag et al., 1997). Heating with domestic microwave oven has also been reported to be effective in extending the shelf-life of rice bran by reducing about 90% of increasing rate of free fatty acids (FFAs) in 3-month (Ramezanzadeh et al., 1999) and 4-month (Faiyaz Ahmed, 2007) storage. Further, it has also been reported that the inactivation of rice bran lipase depends upon temperature, duration of heat treatment, and moisture content, the latter being critical (Jiaxun et al., 1993). Earlier research (Hutchinson and Martin, 1951) has also shown that to inactivate 97–98% of lipase, oat kernels with a
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moisture content of 6, 8, 10, 12, 14, 16, 18 or 20% should be roasted for 1 h under 120, 104, 92, 83, 76, 71, 67 or 64 °C, respectively. Although microwaving has been applied to many cereals and other food materials, and does serve rapid and efficient heating for it requires no heat transmission or loss of heat medium (e.g., steam in steaming), it has seldom been applied to oat processing. In Grant’s experimental setup, 130 g of whole oats was heated in a 22.5 cm tall, 5 cm wide glass cylinder with four 0.3 cm holes randomly placed around the cylinder and covered with a glass lid. Samples were treated for 180 s to a final temperature of 108 °C using a microwave oven (1500 W, 2450 MHz) and the lipase activity was not significantly inactivated (Grant, 1999). This was attributed to the continuous moisture loss during microwave treatment. However, the lipase inactivation by microwave heat treatment, as well as the role of moisture in the process, has not been investigated in previous researches. This work examines the feasibility of microwave heating for inactivating lipase in oat kernels. The effects of moisture content of oat kernels, heating time, temperature, tempering time and packaging on the extent of inactivation are investigated.
281
ene bag (6.5 5.5 1.3 cm3). Sample bags were placed in the center of the revolving glass tray and heated by a domestic microwave oven (Glanz, 2450 MHz, 800 W, Foshan Shunde Galanz Microwave Oven Electrical Appliance Ltd.) for a period of 10– 45 s, respectively, with a starting temperature of 19–22 °C. The center temperature of the packages (with oat kernels sealed inside) by the end of heating was immediately measured in triplicates using a non-contact infrared thermometer (DT-8811H, CEM Shenzhen Ever-best Machinery Industry Co., Ltd.). Treated oat kernels were cooled for 1 min at room temperature (19–22 °C), then poured out from each bag onto a paper tray and air-dried at 40 °C for 1 h. Dried kernels were then ground using a high-speed pulverizer (HY-04 B, Beijing Huanya Tianyuan Machinery technology Co., Ltd.) until over 90% of the oat flour had a particle size of less than 0.425 mm (>90% could pass through a 40mesh sieve). Oat flour obtained was taken out (3 g), packed with a filter paper and defatted by Soxhlet extraction for 6 h using diethyl ether. Defatted flour was dried at 40 °C for 1 h and then stored at 18 °C for the determination of lipase activity. 2.5. Effect of moisture content and tempering time on lipase inactivation
2. Materials and methods 2.1. Materials Oats (Huawan No. 6) cultivated in Zhangjiakou, Hebei province, China, were harvested in September 2007. The whole oat kernels were stored at 10 °C in a sealed polyethylene bag after threshing and removing husked kernels (as contaminants in naked oat, about 5%), fragmentary kernels, dirt plus stone. 2.2. Moisture The moisture content of the oat flour was determined by AACC44-15A [1999] method, using a hot-air oven. Ground oat flour samples (2–3 g) were dried at 130 ± 1 °C for 1 h, and percentage weight loss was calculated as the moisture content.
The moisture contents of the oat kernels were adjusted by adding distilled water to 13.0, 17.0, 20.0 and 25.0% from an initial value of 11.1%, respectively, thoroughly mixed for 10 min, then sealed with a polyethylene film in stainless steel basins and stored at room temperature for tempering. The basins were shaken occasionally for an even moisture distribution. At different tempering times (0, 4, 7, 11 and 28 h), 20.00 g oat kernels of each moisture content level were weighted, respectively, either vacuum packed in a polyethylene bag (6.5 5.5 1.3 cm3) or evenly spread on a coverless culture dish, and treated with microwave heating for 25 s followed by a 10 min cooling at room temperature. The residual lipase activities were analyzed. Each treatment and lipase activity measurement was conducted in duplicate. Oat kernels (M) which had been heated in culture dishes were weighed (M) after cooling for 10 min and the moisture loss was calculated as below:
2.3. Lipase activity
Moisture lossð%Þ ¼ Residual lipase activity was determined in 0.5 g samples using the dough method (Peterson, 1999). To each sample, 98 lL (six drops with 100 lL pipette) of glycerol trioleate (CP, 98–103%, Sinopharm Chemical Reagent Co., Ltd.) was added followed by 330 lL of buffer (0.05 M Tris–HCl, pH 7.5, containing 1% v/v Triton X-100) in to a screw-cap cuvette. The dough samples were mixed thoroughly with a glass rod and incubated at 37 °C for 1 h. The reaction was stopped by adding 100 lL of 1 M HCl and again mixing with a glass rod. For the control, HCl was added immediately. To extract oleic acid hydrolyzed from the glycerol trioleate, 5 mL of hexane was added and tightly sealed with a cap. The cuvettes were put into a boiling water bath for 5 min, and subjected to vortex shaking for 1 min to fully extract oleic acid into hexane, and then centrifuged (3000 r/min, 10 min). Four milliliter supernatant was mixed with 1 mL of copper reagent and the resulting fatty acid copper soap had a strong absorbance at 715 nm (Kwon and Rhee, 1986). The absorbance measured at 715 nm was then compared with an oleic acid standard. The lipase activity was expressed as lmol of oleic acid liberated per hour per gram of the sample. 2.4. Effect of packaging on lipase inactivation In order to study the influence of vacuum and normal packaging on the effects of lipase inactivation by microwave heating, oat kernels (20.0 g) were either sealed or vacuum packed in a polyethyl-
20:00 M 100 20:00
ð1Þ
2.6. Statistical analysis All the data are means of duplicate assays. The results were analyzed using Wilcoxon rank sum test for paired data of lipase activities with vacuum and normal packaging. Lipase activities affected by moisture and tempering time were analyzed by two-way analysis of variance. Linear regression analysis was performed between residual lipase activity and the moisture loss after 25 s microwave heating in coverless culture dishes.
3. Results and discussion 3.1. Effect of packaging on lipase inactivation by microwave heating Fig. 1 describes the variation in the residual lipase activities of the oat kernels in either sealed or vacuum packages after heat treatments varying in duration. For oat kernels in vacuum packages, with heating time prolonged, the residual lipase activity decreased slowly at first (0–25 s), and then sharply to 6.7 ± 0.5 lmol FFA h1 g1 in a period of 25–40 s. A final residual lipase activity of 2.8 ± 2.7 lmol FFA h1 g1 (about 1–2% of initial activity, 352.8 ± 3.6 lmol FFA h1 g1) remained after 45 s heating.
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450
Activity (nor.) Temp. (nor.)
The results of residual lipase activities of both packaging were analyzed with Wilcoxon rank sum test for paired data, and the results showed P < a (0.05), which also confirmed that vacuum packaging could inactivate lipase with ease during microwave heating.
160 140
350 120 300 100 250 y = 3.0463x + 10.537
80
2
200
R = 0.9644 60
150 100
40
y = 2.809x + 4.1033 2
R = 0.9963
50 0 0
Ending temperature (°C)
Lipase activities (µmol•h-1•g-1)
400
Activity (vac.) Temp. (vac.)
20 0
5
10
15
20
25
30
35
40
45
Microwave heating time (s) Fig. 1. The variation of lipase activity and final temperatures with heating time in normal or vacuum packages (nor.: normal package, vac.: vacuum package). (Note: increasing rate of temperature was compared in the same period of 10–30 s).
After heat treatments varying in duration, the levels of residual activities in oat kernels heated in normal packages were all slightly higher than the corresponding levels of kernels in vacuum packages. Generally, a similar decreasing trend in lipase activity was found in both the packages, as well as a similar final level of 1–2% of initial activity after 45 s heating. In contrast, for normal package, residual lipase activity decreased slightly at first, but then increased gradually to the initial level when heated for 25 s, while that in vacuum package shows a continuous decrease. Lipoxygenase activity in groundnut seeds has been reported to be activated during microwave heating. The lipoxygenase activities in groundnut seeds (500 g) increased by about 16% when the seeds were subjected to microwave treatment (at 750 W in a laboratory microwave oven (Philips, Model M710 cooktronic) for 60 s, before a continuous decrease for even longer periods ranging from 60 to 300 s (Ramesh et al., 1995). The final temperature in the center of package surface for both packing methods is compared as shown in Fig. 1. As the heating time increased, the final temperature in vacuum and normal packages increased at a rate of 3.0 °C/s (R2 = 0.96, from 44 °C to 105 °C) and 2.8 °C/s (R2 = 0.99, from 33 °C to 88 °C) in a period of 10–30 s, and reached the point of rapid steam generation after heating for 30 and 40 s, respectively. Unlike the case of normal packages, for oats in vacuum packages, lipase activities declined continuously without any increase during 15–25 s heating. This might be due to the earlier and faster steam generation in vacuum packages at the beginning of heat treatment. In vacuum packages (e.g. absolute pressure 0.02 bar) and normal packages (absolute pressure 1 bar), moisture boils at 17.51 and 100 °C, respectively; thus much higher energy is needed to heat the same quantity of moisture from room temperature to its evaporating temperature. Therefore, compared with normal packages, the depletion of pressure in vacuum packages increases the fugacity of moisture within the packages and enables the evaporation of low temperature steam. The generations of low temperature steam increased the density of moisture inside the package and accelerated the transformation of microwave energy into thermal energy. Consequently, the increasing rate of temperature is slightly higher (0.2 °C/s in the heating period of 10–30 s) in vacuum packages and heated steam may counteract the microwave activation during the heating period of 15–25 s.
3.2. Effects of moisture content and tempering time on lipase inactivation To study the influences of moisture content and tempering time on lipase inactivation, the residual lipase activities of oat kernels with different moisture levels (13–25%) after 25 s of microwave heating at different tempering times were analyzed (Fig. 2). Fig. 2a indicates the variation in lipase activities after heating for 25 s by microwave in coverless culture dishes. The higher the moisture of oats kernels, the lower the residual lipase activity remained after heating for the same period of time. With increase in the tempering time, residual lipase activities of oat kernels with different moisture levels all increased sharply and then reposefully in a period of 0–4 h and 4–28 h after the moisture adjustment, respectively. This is mainly due to the variation in the distribution of adjusted moisture. Most moisture added stayed at the surface and outer layer of kernels just after tempering; this induced the water molecules to have more opportunity to collide with each other and vaporize. With increase in the tempering time, it was difficult to vaporize the water molecules entering the kernels, and moisture transfer rate tended to decrease; thereby the final temperature and the amount of moisture loss were also found to decrease fast at first and slowly later on.
Fig. 2. Lipase activities in oats with different moisture levels after 25 s microwave heating at different tempering times.
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Q. Keying et al. / Journal of Food Engineering 95 (2009) 280–284 Table 1 Two-way analysis of variance of the residual lipase activities after 25-s microwave heating at different moisture levels and tempering times. Moisture content/%
Residual lipase activity/lmol h1 g1
Tempering time/h
Residual lipase activity/lmol h1 g1
Effect factors
Pr > F
25 20 17 13
19.70 d 49.76 c 75.73 b 196.40 a
0 4 7 11 28
43.52 b 98.71 a 83.61 a 99.83 a 101.31 a
Moisture content (a) Tempering time (b) Interaction (a b)
<0.0001 0.0003 0.0180
Means of the same column followed by different letters differ significantly (P < 0.05), according to Duncan’s New Multiple Range Test.
Fig. 2b shows the changes in residual lipase activity of samples after 25 s of microwave heating treatments using vacuum packing. Lipase activities in oat kernels with 17–25% moisture content were inactivated by 98–99% when heat treatment was conducted immediately after moisture adjustment. Similarly, higher levels of residual lipase activity remained with the tempering time increased. Nevertheless, lower residual lipase activity remained in oat kernels using vacuum packages in comparison with that in coverless culture dishes, mainly due to the retention of the steam that was generated in vacuum packages during heating. Table 1 shows the results of a two-way analysis of variance of the lipase activities in oats with different moisture levels after 25 s of microwave heating at different tempering times. The results indicated that the interaction of moisture content and tempering time had a significant effect on the lipase inactivation using microwave heating. This implies that both the factors affected the efficiency of lipase inactivation, mainly due to their correlation with the steam generation: the increase in the moisture content accelerated the steam generation, and the extension in tempering time made the release of moisture difficult as explained above. To confirm this conclusion, the effect of steam quantity on lipase inactivation was further studied by analyzing the correlation between the moisture loss and the residual lipase activities after 25 s heating under different moisture levels at different tempering times. Steam quantity during heating was denoted by the ratio of moisture loss, which was expressed as percentage (%) of moisture loss in every 20.00 g of oat samples (wet based). As shown in Fig. 3, with increase in the tempering time, the ratio of moisture loss decreased dramatically in a period of 0–4 h after moisture adjustment and decreased slowly later on, just the opposite of the changes in residual lipase activities as shown in Fig. 2a. The decrease in the moisture loss of oat kernels with the same moisture level was mainly caused by the prolongation of tempering time when more moisture was transferred from outer to inner layer, which consequently prevented the steam generation and release during heating by microwave.
A significant negative correlation (R2 = 0.9252) was found using the linear regression analysis between residual lipase activity of oat kernels (in coverless culture dishes) and the moisture loss after 25 s of microwave heating when the amount of moisture loss was fairly large (over 3%) (as shown in Fig. 4). No significant linear correlation (R2 = 0.6612) between the moisture loss and the residual lipase activity was found when the quantity of steam was not enough to inactivate the lipase (moisture loss <3%). Moreover, lower moisture loss from oat kernels with a higher moisture content than that with a lower moisture content was found when the tempering time of the former was much longer than that of the latter, and vice versa (as shown in Fig. 3). This suggested that the steam quantity expressed as moisture loss, rather than the moisture content, was the critical factor in the inactivation of lipase. However, the combination of increasing moisture content to 17–25%, conducting heating treatments immediately after a moisture adjustment procedure, and using vacuum packaging could promote the steam release and inactivate the lipase more efficiently. In addition to the steam quantity, temperature is another important factor affecting lipase inactivation. The interaction of temperature and moisture played an important role in the traditional thermal inactivation treatments as reported. For example, 1 h of kilning at 120 and 64 °C was required to inactivate 97–98% of lipase in oat kernels with 20% and 6% moisture (Hutchinson and Martin, 1951). As shown in Table 2, the higher the end temperature was, the lower the residual lipase activity remained at the same tempering time (in the same row) and same moisture content (in the same column). However, no significant correlation between temperature and residual lipase activity was found in this experiment. The possibility of inactivating lipase completely by microwave heating was examined using the maximal heating duration (49 s, up to the pressure tolerance of the package) and the heating treatment was conducted immediately after moisture adjustment to 20%. Residual lipase activity was 0.3 ± 0.3 lmolh1 g1. This suggested a complete lipase inactivation. In addition, the increase of free fatty acids (FFAs) in these microwave-treated oat kernels was even less than 0.5-fold of the initial value (<30 mg KOH/
Fig. 3. Moisture loss in oat kernels with different moisture levels after 25 s microwave heating in coverless culture dishes at different tempering times.
Fig. 4. The correlation between residual activity and moisture loss of oats after 25 s microwave heating (in coverless culture dishes).
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Table 2 The final temperature (°C) and residual lipase activity (lmol h1 g1) after 25-s microwave heating. Tempering time/h
0 4 7 11 28
Moisture content (%) 25
20
17
13
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
Final temperature
Residual lipase activity
105.9 ± 0.9 103.5 ± 1.2 104.0 ± 0.3 105.0 ± 0.0 101.3 ± 0.3
3.3 25 17.1 24 29.2
102.3 ± 1.3 101.2 ± 0.5 100.7 ± 3.0 99.0 ± 2.7 100.0 ± 0.0
4.2 75.5 44.1 94.4 30.6
102.3 ± 1.3 93.5 ± 2.2 98.8 ± 0.2 94.0 ± 1.7 98.2 ± 1.2
14.9 115.7 73.6 98.7 75.8
94.8 ± 4.2 91.7 ± 3.7 93.3 ± 1.3 92.8 ± 0.8 84.7 ± 8.3
151.7 178.8 199.7 182.2 269.7
100 g oat flours, dry-based), whereas in raw oat kernels up to 18fold increase was found, after 1-month storage at 40 °C and 80% relative humidity. 4. Conclusions Steam quantity was a critical factor in lipase inactivation by microwave heating. Microwave heating was able to stabilize the oat flour using a sufficient heating process. Appropriately increasing moisture content of oat kernels to 17–25%, conducting microwave heating treatment immediately after adjusting moisture, and using vacuum packaging could promote the steam release and inactivate the lipase completely. Modification of these conditions was required before they can be applied on a continuous and commercial scale processing. Reusable high-pressure containers may substitute for the disposable polyethylene pouches. The similarity of the role played by steam quality in lipase inactivation both by microwave and steaming provides an interesting topic for a future research. The effects of these two types of treatments on the variation of lipase activity, FFA values during storage and sensory property of oat foods would be investigated in our future work. References Bryngelsson, S., Dimberg, L.H., Kamal-Eldin, A., 2002. Effects of commercial processing on levels of antioxidants in oats (Avena sativa L.). Journal of Agricultural and Food Chemistry 50 (7), 1890–1896. Ekstrand, B., Gangby, I., Akesson, G., 1992. Lipase activity in oats-distribution, pH dependence, and heat inactivation. Cereal Chemistry 69 (4), 379–381. Faiyaz Ahmed, K.P.S.V.S.P.K.S., 2007. Improved shelf-life of rice bran by domestic heat processing and assessment of its dietary consumption in experimental rats. Journal of the Science of Food and Agriculture 87 (1), 60–67. Farag, R.S., El-Baroty, G., Abd-El-Aziz, N., Basuny, A.M., 1997. Stabilization of olive oil by microwave heating. International Journal of Food Sciences and Nutrition 48 (6), 365–371.
Ganbmann, W., Vorwerck, K., 1995. Oat milling, processing and storage. In: The Oat Crop. Chapman and Hall, London, pp. 369–408. Gong, H., Li, C., Wang, Y., 1999. Analysis of oat germplasm quality characters. Journal of Shanxi Agricultural Sciences 27 (2), 16–19. Grant, P.J.A., 1999. Different heating techniques for oat lipase stabilization. Lincoln,Nebraska, University of Nebraska. Doctor: 159. Hutchinson, J.B., Martin, H.F., 1951. Location and destruction of lipase in oats. Nature 167 (4254), 758–759. Jiaxun, T., Rao, R., Liuzzo, J., 1993. Microwave heating for rice bran stabilization. Journal of Microwave Power and Electromagnetic Energy 28 (3), 156–164. Kwon, D.Y., Rhee, J.S., 1986. A simple and rapid colorimetric method for determination. Journal of American oil chemists society 63 (1), 89–92. Liukkonen, K.H., Montfoort, A., Laakso, S.V., 1992. Water-induced lipid changes in oat processing. Journal of Agricultural and Food Chemistry 40 (1), 126– 130. O’Connor, J., Perry, H.J., Harwood, J.L., 1992. A comparison of lipase activity in various cereal grains. Journal of Cereal Science 16 (2), 153–163. Peterson, D.M., 1999. Lipase activity and lipid metabolism during oat malting. Cereal Chemistry 76 (1), 159–163. Ponne, C.T., Moeller, A.C., Tijskens, L.M.M., Bartels, P.V., Meijer, M.M.T., 1996. Influence of microwave and steam heating on lipase activity and microstructure of rapeseed (Brassica napus). Journal of Agricultural and Food Chemistry 44 (9), 2818–2824. Ramesh, M., Haridas Rao, P., Ramadoss, C.S., 1995. Microwave treatment of groundnut (Arachis hypogaea): extractability and quality of oil and its relation to lipase and lipoxygenase activity. Lebensmittel Wissenschaft und Technologie 28 (1), 96–99. Ramezanzadeh, F.M., Rao, R.M., Windhauser, M., Prinyawiwatkul, W., Tulley, R., Marshall, W.E., 1999. Prevention of hydrolytic rancidity in rice bran during storage. Journal of Agricultural and Food Chemistry 47 (8), 3050– 3052. Stevenson, D.G., Inglett, G.E., Chen, D.J., Biswas, A., Eller, F.J., Evangelista, R.L., 2008. Phenolic content and antioxidant capacity of supercritical carbon dioxide-treated and air-classified oat bran concentrate microwave-irradiated in water or ethanol at varying temperatures. Food Chemistry 108 (1), 23–30. Vetrimani, R., Jyothirmayi, N., Rao, P.H., Ramadoss, C.S., 1992. Inactivation of lipase and lipoxygenase in cereal bran, germ and soybean by microwave treatment. Lebensmittel-Wissenschaft und Technologie 25 (6), 532–535. Youngs, V.L., 1978. Oat lipids. Cereal Chemistry 55 (5), 591–597. Zadernowski, R., Nowak-Polakowska, H., Rashed, A.A., 1999. The influence of heat treatment on the activity of lipo- and hydrophilic components of oat grain. Journal of Food Processing and Preservation 23 (3), 177–191.