Effect of superheated steam on moisture content
Figure 1a shows the effect of SS processing temperature and time on moisture content of buckwheat grains. The moisture content of SS-treated buckwheat grains decreased with the extended time and increased temperature of SS. When buckwheat grains were treated by 110 °C SS,140 °C SS,170 °C SS and 200 °C SS for 7 min, their moisture contents were decreased from 20.26–14.82%, 12.10%, 8.89% and 5.81% respectively, namely, higher temperature led to higher rate of moisture removal of buckwheat grains. In particular, after treated with SS at 170 °C for 5 min, the moisture content decreased from 20.26–12.09%, which was not significantly different from the original moisture content of untreated buckwheat (12.28%).
Effect of superheated steam on lipase inactivation
Lipase activity of buckwheat grains decreased with an increase in processing time and temperature of SS (Fig. 1b). Higher temperatures (170 °C and 200 °C) of SS treatment showed a better inactivate effect than 110 °C or 140 °C. An average level of lipase activity of buckwheat grains decreased by 26.24%, 28.98%, 55.53% and 62.70%, respectively, after the SS treatment at 110 °C, 140 °C, 170 °C and 200 °C for 7 min. Meanwhile, it was observed that when SS treatment time was shorter than 1 min, the steam temperature had no significant effect on the improvement of lipase inactivated efficiency. More than 50% of the lipase activity could be inactivated at 170 °C when the SS treatment was more than 5 min, but there was no significant difference between 170 °C-5 min and 170 °C -7 min in the reduction of lipase activity.
Free fatty acid accumulation in buckwheat during storage
The effect of SS on FFA accumulation during storage was shown in Fig. 2a-c. There was an increasing trend of FFA in untreated buckwheat at 4 and 25 °C during storage, and the highest FFA was observed after 12 weeks of storage. The FFA of untreated buckwheat stored at 50 °C increased rapidly at the beginning of storage, reached maximum levels (134.03 mg KOH/100 g d.b.) at 8 weeks, and then showed a slight decline afterward. Compared with untreated buckwheat, SS-treated buckwheat exhibited significantly fewer accumulation in FFA during storage at each storage temperature. The FFA of SS-treated buckwheat did not change significantly when stored at 4 °C and 25 °C for 12 weeks, even for 50 °C storage, which was always lower than 100 mg KOH/100 g d.b.
Changes in lipase of buckwheat during storage
Changes in lipase of untreated and SS-treated buckwheat during storage were summarized in Fig. 2e-f. In general, the lipase activity of SS-treated samples was significantly (P < 0.05) lower than that of untreated samples at each temperature. The lipase of all untreated buckwheat samples initially decreased from 7.57 KOH/g d.b and reached minimum levels of 3.16–5.84 KOH/g d.b after 8 weeks of storage. The lipase activity of untreated buckwheat decreased most by 57.85% at storage temperature of 50 °C for 8 weeks, but that of SS-treated buckwheat decreased slightly lower by 44.44%. Moreover, there was no significant difference in lipase activity of SS-treated buckwheat among different temperature conditions at the end of storage.
Fatty acid compositions of buckwheat during storage
Fatty acid compositions of untreated and SS-treated buckwheat during storage were summarized in Table 1. Before storage, the predominant fatty acids in untreated buckwheat were palmitic acid (18.04 ± 0.35%), oleic acid (34.31 ± 0.43%) and linoleic acid (35.00 ± 0.58%), which together constituted 87.35% of the total fatty acids. The percentages of stearic acid, behenic acid and docosenoic acid were significantly (P < 0.05) increased after SS treatment. There was no significant difference (P > 0.05) in percentage distribution of total SFA, total monounsaturated fatty acids (MUFA), and total polyunsaturated fatty acids (PUFA) between untreated and SS-treated buckwheat. After 12 weeks storage, the percentage of total SFA in untreated buckwheat increased significantly and that of total PUFA decreased significantly. The biggest reduction of PUFA was found in linoleic acid. However, SS treatment significantly retard the increase of SFA (mainly palmitic acid, stearic acid, arachidic acid and behenic acid) and the reduction of PUFA (mainly linoleic acid) during storage. The fatty acid compositions of stored SS-treated buckwheat were similar to those of fresh untreated and SS-treated buckwheat.
Table 1
Fatty acid compositions (% of total fatty acids) of buckwheat grains during storage
Fatty acids | 0 week | | 12 week |
Untreated | SS-treated | Untreated | SS-treated |
C14:0 Myristic | 0.16 ± 0.14a | 0.17 ± 0.07a | | 0.16 ± 0.06a | 0.18 ± 0.02a |
C15:0 Pentadecanoic | 0.16 ± 0.01a | 0.13 ± 0.02b | 0.16 ± 0.02a | 0.14 ± 0.00ab |
C16:0 Palmitic | 18.04 ± 0.35b | 17.51 ± 0.14b | 19.04 ± 0.65a | 17.85 ± 0.79b |
C16:1 Palmitoleic | 0.21 ± 0.06b | 0.17 ± 0.00b | 0.17 ± 0.00b | 0.28 ± 0.01a |
C18:0 Stearic | 2.06 ± 0.04c | 2.14 ± 0.01b | 2.49 ± 0.05a | 2.00 ± 0.02c |
C18:1 Oleic | 34.31 ± 0.43a | 33.96 ± 0.28a | 34.02 ± 0.35a | 34.62 ± 0.37a |
C18:2 Linoleic (ω-6) | 35.00 ± 0.58a | 34.91 ± 0.30a | 33.04 ± 0.12b | 34.83 ± 0.29a |
C18:3 Linolenic (ω-3) | 2.85 ± 0.04a | 2.72 ± 0.04a | 2.43 ± 0.03b | 2.47 ± 0.17b |
C20:0 Arachidic | 1.32 ± 0.15b | 1.50 ± 0.13b | 1.86 ± 0.05a | 1.41 ± 0.06b |
C20:1 Gadoleic | 2.84 ± 0.12ab | 3.07 ± 0.06a | 2.64 ± 0.09b | 2.73 ± 0.20b |
C22:0 Behenic | 1.69 ± 0.11c | 1.97 ± 0.01b | 2.41 ± 0.09a | 1.94 ± 0.11b |
C22:1 Docosenoic | 0.07 ± 0.12b | 0.25 ± 0.01a | 0.14 ± 0.12ab | 0.23 ± 0.01ab |
C24:0 Lignoceric | 1.29 ± 0.19a | 1.49 ± 0.08a | 1.44 ± 0.05a | 1.32 ± 0.10a |
∑SFA | 24.72 ± 0.15b | 24.92 ± 0.18b | 27.56 ± 0.49a | 24.84 ± 0.59b |
∑MUFA | 37.43 ± 0.45a | 37.45 ± 0.35a | 36.97 ± 0.56a | 37.86 ± 0.55a |
∑PUFA | 37.85 ± 0.58a | 37.63 ± 0.34a | 35.47 ± 0.09b | 37.31 ± 0.29a |
Values are mean ± standard deviation (n = 3). Mean in a row with different lowercase letters indicate statistical difference (P < 0.05). SFA, saturated fatty acids; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids. |
Lipidomics profile differences of buckwheat samples
Figure S1 (Supplementary Material 1) shows representative UPLC-Q-Exactive Orbitrap MS spectra of lipid extract from different buckwheat samples. The signals in positive ionization mode were mainly attributed to sphingoid (So), phosphatidylcholine (PC), phosphatidylethanolamine (PE), triacylglycerol (TG), lysophosphatidylcholine (LPC), diacylglycerol (DG), and in negative ionization mode were mainly attributed to ceramide (Cer), cardiolipin (CL), phosphatidic acid (PA), phosphatidylglycerol (PG), phosphatidylserine (PS), phosphatidylinositol (PI), monogalactosyl diacylcerol (MGDG), digalactosyl diacylglycerol (DGDG). The different molecular species of lipids in buckwheat were shown in Tables S1-S4 (Supplementary Material 2). A total of 457 lipid molecular species belonging to 23 lipid classes were detected. The lipids in buckwheat were dominantly composed of GPs and GLs, which accounted for 67.57–87.59% of the total lipids. The number of lipid molecular species of each lipid subclass in buckwheat was shown in Table 2. The sum of lipid molecular species of the buckwheat were: stored SS-treated buckwheat > fresh SS-treated buckwheat > stored untreated buckwheat > fresh untreated buckwheat. Before storage, the total species of GPs decreased and that of GLs increased after SS processing. Storage significantly increased the GLs and GPs species in untreated buckwheat, and SS treatment slow down this change in GPs species during storage. The lipids categories were analyzed by summing the relative abundance of individual lipids in the same class (Fig. 3). The relative abundance of PC and PE decreased significantly after SS treatment before storage, and that of PS, PG, So, phosphatidylinositol monophosphate (PIP), lysophosphatidic acid (LPA), TG, DG and monoglyceride (MG) increased significantly (P < 0.05). Meanwhile, the relative abundance of PA, PC, LPA, digalactosyl monoacylglycerol (DGMG) and TG significantly decreased after storage, and that of CL, DGDG, sulfoquinovosyl diacylglycerol (SQDG), phosphatidylinositol diphosphate (PIP2), PE, PG and PS increased. However, SS treatment could retard the changes of SQDG, PE, PS and LPA during storage. It was noteworthy that the relative abundance of TG, DG and MG in stored SS-treated buckwheat decreased significantly, while that of monogalactosyl monoacylglycerol (MGMG), MGDG, DGMG and DGDG significantly increased, compared with the fresh untreated buckwheat.
Table 2
The number of lipid molecular species of each lipid subclass in buckwheat grains during storage
Lipid classes | RT(time) | 0 week | | 12 week |
Untreated | SS-treated | Untreated | SS-treated |
LPA | 2.16–3.08 | 3 | 3 | | ND | 3 |
LPC | 2.01–4.37 | 3 | 3 | 2 | 8 |
LPE | 4.42–6.83 | 3 | 3 | ND | 1 |
PA | 7.50-12.47 | 5 | 5 | 6 | 6 |
PC | 6.74–12.58 | 26 | 8 | 25 | 19 |
PE | 2.54–12.85 | 25 | 5 | 23 | 13 |
PG | 4.57–13.16 | 10 | 14 | 20 | 11 |
PS | 6.19–12.66 | 11 | 16 | 17 | 13 |
PI | 6.43–12.99 | 7 | 11 | 9 | 12 |
CL | 7.22–13.71 | 10 | 12 | 18 | 17 |
PIP | 6.62–12.08 | 6 | 9 | 5 | 9 |
PIP2 | 9.65–9.85 | ND | ND | 2 | ND |
PIP3 | 8.67 | 1 | 1 | 1 | 1 |
Total GPs | 2.01–13.71 | 109 | 89 | 127 | 112 |
DGDG | 6.27–11.75 | 5 | 6 | 7 | 10 |
DGMG | 1.62–3.19 | 3 | 3 | 1 | 3 |
MGDG | 4.09–11.65 | 6 | 7 | 6 | 13 |
MGMG | 2.23–3.80 | 1 | 1 | 2 | 3 |
SQDG | 2.35–11.63 | 7 | 9 | 14 | 7 |
MG | 11.75–12.09 | ND | 2 | ND | 2 |
DG | 7.21–15.71 | 19 | 25 | 19 | 24 |
TG | 6.57–16.08 | 90 | 135 | 95 | 123 |
Total GLs | 1.62–16.08 | 131 | 188 | 144 | 185 |
Cer | 1.79–13.35 | 19 | 16 | 15 | 17 |
So | 0.71–4.29 | 5 | 6 | 7 | 8 |
Total SPs | 0.71–13.35 | 24 | 22 | 22 | 25 |
SUM | | 264 | 299 | | 293 | 322 |
ND means not detected. |
Association of hydrolytic rancidity with lipase activity and lipidomics profile
The scatter plots and linear regression of FFA value vs. lipase activity in untreated and SS-treated buckwheat are presented in Fig. 4a and b. The association of FFA value with lipase activity in untreated and SS-treated buckwheat had a R2 values of 0.8221 and 0.6777, respectively. The slopes of SS-treated buckwheat (-8.28) was lower than that of untreated buckwheat (-10.27). The correlations of hydrolytic rancidity and lipid profiles were analyzed using Pearson correlation analysis and the correlation coefficients were given in Fig. 4c. The results showed that FFA value was significantly positively (P < 0.01) correlated with the relative abundance of SQDG, PE and PIP2, while had negative correlation with LPE (P < 0.01), TG, DG and MG. In addition, lipase activity was negatively correlated with the relative abundance of So, CL, MGMG, MGDG DGDG, SQDG, PA, PG, PI, PIP and PIP3.