3.1 Volatile profile of RAPT with various years of storage
In the present work, an HS-SPME-GC-MS method was optimized and developed to profile the volatile compounds in RAPT. Based on the optimized method, qualitative and quantitative analysis of the volatile compounds were performed. As illustrated in Fig.S1 (Total ion chromatograms (TIC)), a large number of volatile compounds were detected in RAPT, and the profile of RAPT with different years of storage varied from each other. The data analysis was performed by using a commercial software (GSMSsolution, labSolition, Shimadzu). A total of 130 volatile compounds were finally identified by matching NIST library and comparing the retention indexes (RIs), and the results were presented in Table 1. These compounds include alcohols (22%), aldehydes (12.1%), ketones (18.2%), alkenes (14.4%), esters (8.3%), acids (8.3%), pyrroles (3.8%), and others (12.9%), as illustrated in Fig. 1B. In terms of the number of detected compounds, alcohols, ketones, and alkenes are the top 3 classes of compounds in RAPT. Of them, 64 compounds (approximately 49.23%) were shared by all RAPT samples regardless the years of storage (Fig. 1A). It is worth noting that a few of compounds, such as 1,2,3-trimethoxybenzene, hexanal and 2-hexenal, were produced with a storage time of over two years.
Table 1
Volatile compounds identified in RAPT samples from different storage durations
Volatile compounds | RI | Concentration (µg/100g) |
T0 | T1 | T2 | T3 | T4 | T5 | T6 | T7 | T8 | T9 | T10 |
Hexanal | 800 | - | 0.45 ± 0.06 | 6.04 ± 0.80 | - | - | 1.38 ± 0.22 | 0.41 ± 0.02 | 0.52 ± 0.06 | 0.53 ± 0.01 | 0.64 ± 0.12 | 0.65 ± 0.05 |
3-methyl-Butanoic acid | 811 | 4.44 ± 0.50ef | 5.86 ± 1.09g | 1.92 ± 0.03bc | 3.04 ± 0.48d | 5.12 ± 0.40f | 3.75 ± 0.37de | 0.66 ± 0.06a | 1.81 ± 0.28bc | 1.22 ± 0.06ab | 1.27 ± 0.10ab | 2.08 ± 0.03c |
2-methyl-Butanoic acid | 811 | 5.03 ± 0.82 f | 4.55 ± 0.76 f | 2.37 ± 0.03 c | 3.26 ± 0.32 e | 3.13 ± 0.05 de | 2.52 ± 0.31 cd | 0.61 ± 0.10 a | 1.09 ± 0.19 ab | 0.64 ± 0.13 a | 1.35 ± 0.23 b | 1.43 ± 0.08 b |
1-ethyl-1H-Pyrrole | 829 | 1.37 ± 0.02 | 0.95 ± 0.03 | - | - | - | - | - | - | - | - | - |
1,2,5,5-tetram ethyl-1,3-Cycl opentadiene | 840 | 2.78 ± 0.36bc | 2.32 ± 0.07bc | 1.44 ± 0.26a | 5.37 ± 0.76e | 3.68 ± 0.63d | 2.02 ± 0.25ab | 2.26 ± 0.11bc | 1.52 ± 0.23a | 2.38 ± 0.26bc | 2.2 ± 0.140bc | 4.30 ± 0.33d |
(Z)-3-Hexen-1-ol | 851 | 3.02 ± 0.21 | 2.92 ± 0.28 | - | - | - | - | - | - | - | - | - |
2-Hexenal | 853 | - | - | 3.34 ± 0.56 | 6.65 ± 0.39 | 11.02 ± 0.49 | 3.75 ± 0.44 | 5.88 ± 1.00 | 3.16 ± 0.53 | 7.68 ± 1.61 | 6.13 ± 0.51 | 6.79 ± 0.82 |
1-Hexanol | 867 | 5.58 ± 0.07f | 5.80 ± 0.79f | 3.05 ± 0.03cd | 4.06 ± 0.30e | 4.62 ± 0.73e | 3.20 ± 0.39d | 2.29 ± 0.10b | 2.14 ± 0.36ab | 3.01 ± 0.16cd | 1.53 ± 0.31a | 2.36 ± 0.35bc |
p-Xylene | 870 | 1.79 ± 0.28ab | 1.92 ± 0.21abc | 1.72 ± 0.29ab | 5.71 ± 0.40f | 4.22 ± 0.61e | 1.67 ± 0.32a | 2.62 ± 0.07d | 2.54 ± 0.26d | 2.88 ± 0.03d | 2.46 ± 0.18cd | 2.28 ± 0.46bcd |
2-Heptanone | 889 | 4.15 ± 0.54de | 3.94 ± 0.20de | 2.78 ± 0.01ab | 3.81 ± 0.57c | 5.83 ± 0.77f | 4.58 ± 0.08e | 2.99 ± 0.29ab | 2.36 ± 0.30a | 3.06 ± 0.11b | 3.98 ± 0.12de | 2.34 ± 0.17a |
Heptanal | 901 | 4.58 ± 0.76ab | 4.51 ± 0.48ab | 7.8 ± 0.18de | 6.68 ± 0.53cd | 8.71 ± 1.07e | 7.09 ± 0.42d | 7.03 ± 1.17d | 3.42 ± 0.45a | 7.60 ± 0.13de | 5.69 ± 0.35bc | 5.16 ± 1.03b |
2-Propenoic acid, butyl ester | 902 | 1.23 ± 0.13 | - | - | - | - | - | - | - | - | - | - |
2-butoxy-Ethanol | 907 | - | 0.46 ± 0.05 | - | - | 1.59 ± 0.02 | - | 0.94 ± 0.18 | 0.50 ± 0.06 | 0.38 ± 0.07 | 0.64 ± 0.07 | 0.66 ± 0.09 |
Hexanoic acid, methyl ester | 924 | - | - | - | - | 0.99 ± 0.06 | - | 0.39 ± 0.04 | 2.36 ± 0.30 | 0.50 ± 0.03 | - | - |
Camphene | 933 | - | - | - | - | - | - | 1.20 ± 0.10 | 1.32 ± 0.23 | - | - | - |
α-Pinene | 939 | 0.58 ± 0.09ab | 0.64 ± 0.07bc | 0.45 ± 0.07a | 0.50 ± 0.01a | 0.78 ± 0.03cd | 0.77 ± 0.11cd | 0.82 ± 0.12d | 1.05 ± 0.09e | 1.04 ± 0.07e | 1.04 ± 0.08e | 0.75 ± 0.04cd |
2-Octanone | 952 | 0.53 ± 0.04 | - | - | - | - | 1.36 ± 0.24 | - | - | - | - | - |
Hexanoic acid | 961 | 26.59 ± 3.60 | 25.95 ± 2.40 | 9.51 ± 1.53 | - | 27.39 ± 0.27 | 26.98 ± 5.34 | - | - | - | - | - |
1-(3-methylenecyclopentyl)-Ethanone | 963 | - | - | - | - | - | - | 0.73 ± 0.03 | 0.59 ± 0.09 | - | - | - |
Benzaldehyde | 966 | 14.77 ± 0.67a | 18.15 ± 0.58ab | 35.90 ± 4.64d | 26.23 ± 1.89c | 46.78 ± 4.13e | 24.25 ± 4.58bc | 18.65 ± 1.86ab | 20.73 ± 1.42abc | 23.83 ± 4.25bc | 21.93 ± 4.13bc | 24.15 ± 4.26bc |
3,5,5-trimethyl-2-Hexene | 968 | 37.93 ± 1.16g | 19.55 ± 2.69f | 7.66 ± 1.02d | 7.12 ± 0.70cd | 10.46 ± 1.57e | 8.69 ± 1.55de | 3.33 ± 0.50ab | 3.30 ± 0.58ab | 2.89 ± 0.56a | 4.41 ± 0.10ab | 5.28 ± 0.87bc |
2-Methylenecyclohexanol | 974 | - | - | - | - | - | - | 2.71 ± 0.49 | 3.34 ± 0.68 | 3.81 ± 0.08 | 4.78 ± 0.69 | - |
1-Octen-3-ol | 976 | 95.72 ± 2.24g | 51.67 ± 5.98f | 26.59 ± 2.40d | 23.11 ± 2.95cd | 36.69 ± 1.59e | 21.79 ± 1.34c | 11.90 ± 0.74a | 12.06 ± 2.07a | 12.57 ± 1.39ab | 17.11 ± 0.51b | 15.51 ± 2.53ab |
2,3-Octanedione | 986 | 7.45 ± 0.87c | 4.24 ± 0.16ab | 5.39 ± 0.86b | 9.51 ± 1.89d | 7.45 ± 1.40c | 4.07 ± 0.83ab | 3.12 ± 0.16a | 2.74 ± 0.52a | 3.71 ± 0.43a | 2.65 ± 0.02a | 3.08 ± 0.32a |
6-methyl-5-Hepten-2-one | 987 | 19.46 ± 1.55ef | 14.85 ± 0.99cd | 12.83 ± 1.47abc | 16.39 ± 3.17cde | 21.93 ± 3.56f | 17.33 ± 3.32de | 10.80 ± 1.05ab | 9.99 ± 1.93ab | 9.30 ± 1.33a | 13.45 ± 0.59bc | 12.93 ± 0.69abc |
β-Myrcene | 992 | - | 17.95 ± 0.84 | - | - | - | 21.02 ± 3.18 | - | - | - | 16.13 ± 0.84 | 12.85 ± 1.02 |
(2R,5R)-2-Methyl-5-(prop-1-en-2-yl)-2-vinyltetrahydrofuran | 993.8 | 4.74 ± 0.32a | 5.62 ± 0.07ab | 4.89 ± 0.76ab | 7.50 ± 1.42cd | 11.66 ± 1.55e | 6.53 ± 1.16abc | 5.03 ± 0.01ab | 5.78 ± 1.14abc | 6.62 ± 0.14bc | 8.92 ± 1.46d | 6.30 ± 0.58abc |
Octanal | 1001 | 7.28 ± 0.09ab | 6.62 ± 0.67ab | 6.01 ± 0.27a | 6.26 ± 1.19a | 18.80 ± 0.89d | 11.07 ± 1.91c | 6.98 ± 0.59ab | 6.21 ± 0.97a | 8.56 ± 1.28b | 8.36 ± 1.42b | 7.43 ± 1.07ab |
(E,E)-2,4-Heptadienal | 1011 | 11.52 ± 0.35abc | 10.40 ± 0.30a | 10.68 ± 2.06ab | 15.28 ± 0.40e | 18.85 ± 1.86f | 11.11 ± 1.87ab | 13.18 ± 1.21bcde | 12.34 ± 2.32abcd | 14.41 ± 0.18de | 14.13 ± 0.28de | 13.92 ± 1.45cde |
α-Terpinene | 1018 | 4.93 ± 0.23ab | 5.96 ± 0.12cd | 4.64 ± 0.50ab | 6.40 ± 0.19d | 8.56 ± 0.11e | 5.97 ± 0.39cd | 4.82 ± 0.46ab | 5.12 ± 0.45bc | 5.40 ± 0.35bc | 6.67 ± 1.18d | 4.03 ± 0.64a |
Benzyl chloride | 1019 | 1.50 ± 0.05d | 1.01 ± 0.13abc | 1.06 ± 0.18bc | 1.62 ± 0.08de | 1.72 ± 0.07e | 1.05 ± 0.18abc | 1.15 ± 0.15bc | 0.97 ± 0.16ab | 1.22 ± 0.09c | 0.82 ± 0.09a | 0.94 ± 0.01ab |
o-Cymene | 1026 | 4.22 ± 0.16 | - | - | 15.27 ± 2.08 | - | 19.3 ± 3.55 | - | - | - | - | - |
2-ethyl-1-Hexanol | 1026.3 | 3.74 ± 0.64a | 56.79 ± 4.80f | 33.71 ± 6.19c | 32.19 ± 3.17c | 60.63 ± 5.05f | 19.3 ± 3.55b | 28.46 ± 0.87c | 49.97 ± 1.65e | 42.02 ± 0.52d | 56.39 ± 0.84f | 49.57 ± 3.43e |
D-Limonene | 1030 | 44.37 ± 7.79b | 68.31 ± 1.50d | 74.91 ± 12.27cd | 27.88 ± 3.64d | 74.48 ± 4.30a | 57.35 ± 11.33bc | 45.45 ± 3.18b | 55.43 ± 8.92b | 53.03 ± 4.01b | 69.44 ± 5.84cd | 43.89 ± 7.40b |
Benzyl alcohol | 1034.4 | 14.21 ± 0.19 | - | - | 12.43 ± 0.26 | - | - | - | - | - | - | - |
2,6-dimethyl-5-Heptenal | 1044 | 25.51 ± 0.37e | 21.35 ± 1.30cd | 18.09 ± 3.51bc | 25.65 ± 2.28e | 24.54 ± 3.10de | 20.66 ± 2.77c | 13.24 ± 0.91a | 14.65 ± 2.43ab | 15.89 ± 0.06ab | 13.44 ± 1.49a | 12.94 ± 0.81a |
Benzeneacetaldehyde | 1049 | 11.93 ± 0.26ab | 9.98 ± 0.62a | 11.99 ± 1.36ab | 22.67 ± 0.96d | 22.44 ± 3.00d | 11.63 ± 1.93ab | 12.24 ± 1.19ab | 12.24 ± 2.10ab | 19.25 ± 0.65c | 14.11 ± 0.30b | 17.16 ± 0.73c |
(Z)-3,7-dimethyl-1,3,6-Octatriene | 1051 | 6.53 ± 0.26ab | 8.29 ± 0.68cde | 7.43 ± 0.68abcd | 9.44 ± 0.35e | 10.86 ± 0.37f | 7.81 ± 0.93bcd | 6.94 ± 0.38ab | 6.44 ± 0.86a | 7.14 ± 0.44abc | 8.52 ± 0.54de | 7.54 ± 1.29abcd |
1H-Pyrrole-2-carboxaldehyde, 1-ethyl- | 1054 | 25.94 ± 0.51e | 23.39 ± 2.05de | 23.00 ± 4.15de | 24.66 ± 3.13de | 32.01 ± 2.66f | 20.12 ± 3.50cd | 10.05 ± 0.31a | 12.15 ± 2.43a | 9.05 ± 1.36a | 13.20 ± 0.29ab | 17.39 ± 3.04bc |
2-Octenal, (E)- | 1056.7 | - | - | - | - | - | - | - | 2.11 ± 0.31 | - | 2.87 ± 0.05 | - |
Ethanone, 1-(1H-pyrrol-2-yl)- | 1063.2 | 13.99 ± 1.34bc | 13.35 ± 0.74b | 13.25 ± 0.65b | 25.82 ± 0.14d | 23.12 ± 2.73d | 16.67 ± 2.63c | 11.84 ± 0.82ab | 12.01 ± 1.22ab | 12.96 ± 2.29ab | 12.39 ± 2.09ab | 9.93 ± 1.76a |
γ-Terpinene | 1064 | 12.38 ± 2.06bc | 13.80 ± 1.93cd | 15.40 ± 2.70d | 12.75 ± 0.50bcd | 10.23 ± 0.88b | 6.90 ± 0.21a | 6.32 ± 0.08a | 10.18 ± 1.49b | 7.18 ± 0.67a | 13.09 ± 0.20cd | 12.61 ± 2.44bcd |
(E)-2-Octen-1-ol | 1067 | 59.44 ± 2.59g | 34.28 ± 3.24f | 28.8 ± 2.29e | 27.26 ± 0.19e | 27.93 ± 0.66e | 18.67 ± 1.05d | 13.39 ± 0.38c | 9.72 ± 1.11b | 6.22 ± 0.82a | 9.99 ± 0.55b | 9.81 ± 0.55b |
1-Octanol | 1068 | 20.41 ± 0.43c | 25.39 ± 1.53d | 27.68 ± 5.06d | 49.66 ± 0.06f | 39.84 ± 2.93e | 26.91 ± 4.28d | 8.53 ± 1.21ab | 7.62 ± 0.37a | 12.67 ± 0.51b | 10.15 ± 0.95ab | 10.55 ± 1.29ab |
trans-Linalool oxide (furanoid) | 1076 | 46.04 ± 4.44ab | 44.25 ± 4.77ab | 61.16 ± 6.85cd | 76.5 ± 14.48e | 67.97 ± 2.44de | 52.65 ± 7.51bc | 41.15 ± 3.95ab | 38.55 ± 5.53a | 43.51 ± 7.40ab | 43.03 ± 2.98ab | 49.44 ± 1.17ab |
endo-2-Methylbicyclo[3.3.1]nonane | 1083.34372 | 0.85 ± 0.12 | - | - | - | - | - | - | 1.53 ± 0.16 | - | - | - |
cis-5-ethenyltet rahydro-α, α, 5-trimethyl-2-Fu ranmethanol | 1088 | 90.90 ± 3.28cd | 88.30 ± 8.28cd | 103.91 ± 8.67de | 93.83 ± 18.50cde | 108.56 ± 3.73e | 79.86 ± 12.01bc | 65.34 ± 8.57ab | 52.30 ± 9.05a | 61.88 ± 6.83a | 66.3 ± 3.94ab | 66.69 ± 8.19ab |
3,5-Octadien-2-one | 1093 | - | - | - | - | 19.47 ± 0.73 | - | 9.12 ± 0.41 | - | - | - | - |
3-Cyclohexen-1-one, 3,5,5-trimethyl- | 1097 | 1.56 ± 0.25 | - | - | - | - | - | - | - | - | - | - |
2-Cyclohexen-1-one, 3,4,4-trimethyl- | 1097 | - | - | 0.63 ± 0.05 | - | 0.94 ± 0.10 | 0.73 ± 0.14 | - | - | 0.67 ± 0.11 | - | - |
Linalool | 1106 | 271.51 ± 2.55de | 253.54 ± 15.61de | 275.46 ± 46.89e | 482.41 ± 5.36g | 332.09 ± 29.48f | 231.47 ± 27.77cd | 182.61 ± 19.81ab | 161.46 ± 29.32a | 209.00 ± 6.52bc | 201.65 ± 8.40abc | 184.53 ± 20.76ab |
3,7-dimethyl-1,5,7-Octatrien-3-ol | 1106 | 70.30 ± 1.19def | 63.29 ± 5.05cd | 82.77 ± 13.45f | 78.88 ± 14.00ef | 112.34 ± 3.17g | 67.20 ± 11.20de | 51.62 ± 2.36bc | 43.34 ± 8.27ab | 29.66 ± 4.87a | 41.49 ± 1.24ab | 37.53 ± 1.89a |
(E,E)-2,4-Octadienal | 1113 | - | - | - | - | - | - | - | 3.16 ± 0.45 | - | 3.53 ± 0.07 | - |
2,6-dimethyl-Cyclohexanol | 1114 | 28.45 ± 0.56bc | 27.36 ± 3.32bc | 43.77 ± 7.89d | 31.94 ± 3.72c | 32.85 ± 5.38c | 23.78 ± 4.75b | 13.27 ± 2.32a | 11.37 ± 1.90a | 13.43 ± 1.89a | 11.54 ± 0.61a | 16.37 ± 0.01a |
Phenylethyl Alcohol | 1116 | 22.40 ± 0.32 | 19.74 ± 1.41 | - | 34.19 ± 5.41 | 32.15 ± 0.47 | - | - | 12.63 ± 2.50 | - | 13.73 ± 1.58 | - |
2-ethyl-Hexanoic acid | 1116.7 | - | 4.03 ± 0.50 | - | 6.62 ± 1.15 | - | - | - | - | 4.68 ± 0.53 | - | - |
2,6,6-trimethyl-2-Cyclohexene-1-carboxaldehyde | 1122.9 | - | - | - | - | 4.65 ± 0.74 | - | 6.71 ± 0.41 | 8.39 ± 0.84 | 5.51 ± 0.94 | 8.16 ± 0.76 | 9.35 ± 1.53 |
Isophorone | 1124 | 2.69 ± 0.28a | 2.79 ± 0.48a | 4.72 ± 0.86cd | 5.18 ± 0.67d | 5.61 ± 0.60d | 3.94 ± 0.47bc | 2.52 ± 0.09a | 2.67 ± 0.45a | 2.84 ± 0.52a | 2.67 ± 0.21a | 3.72 ± 0.45b |
(E,Z)-2,6-dimethyl-2,4,6-Octatriene | 1131 | - | - | 1.99 ± 0.22 | - | - | - | - | - | - | - | 1.59 ± 0.12 |
E,E-2,6-Dimethyl-1,3,5,7-octatetraene | 1132 | 2.73 ± 0.18 | 1.91 ± 0.15 | 2.7 ± 0.07 | - | 2.64 ± 0.37 | 1.17 ± 0.18 | - | - | - | 0.92 ± 0.06 | - |
3-Nonen-2-one | 1136 | 1.37 ± 0.22ab | 1.49 ± 0.14abc | 1.73 ± 0.23abc | 2.79 ± 0.34d | 3.50 ± 0.62e | 2.51 ± 0.27d | 1.24 ± 0.17a | 1.32 ± 0.16a | 1.48 ± 0.24abc | 1.96 ± 0.18c | 1.89 ± 0.14bc |
Benzyl nitrile | 1143 | - | - | - | - | 3.57 ± 0.37 | 2.35 ± 0.31 | 2.43 ± 0.36 | 2.60 ± 0.09 | 1.22 ± 0.04 | - | 1.67 ± 0.28 |
(R,S)-5-Ethyl-6-methyl-3E-hepten-2-one | 1143.9 | 6.47 ± 1.25 | - | - | - | - | - | - | - | - | - | - |
1,2-dimethoxy-Benzene | 1146 | 6.51 ± 1.21a | 11.85 ± 1.11de | 16.85 ± 0.59f | 17.19 ± 1.41f | 17.24 ± 0.52f | 16.19 ± 0.95f | 9.68 ± 1.61bc | 8.28 ± 1.34ab | 7.99 ± 0.63ab | 10.65 ± 0.19cd | 13.52 ± 0.09e |
(E,E)-2,6-Nonadienal | 1146 | - | - | - | - | - | 1.89 ± 0.26 | - | 1.38 ± 0.11 | - | - | 1.49 ± 0.27 |
2,3,3-trimethyl-Bicyclo[2.2.1]heptan-2-ol | 1148 | 5.41 ± 0.61ef | 4.97 ± 0.92cde | 5.39 ± 0.21ef | 7.85 ± 0.97g | 6.21 ± 0.16f | 5.14 ± 0.52de | 4.32 ± 0.49bcd | 3.15 ± 0.04a | 4.01 ± 0.83abc | 5.94 ± 0.12ef | 3.78 ± 0.01ab |
1-Nonanol | 1169.2 | 10.81 ± 0.17cd | 8.38 ± 0.42ab | 11.53 ± 1.74d | 17.39 ± 1.81e | 19.69 ± 2.21f | 10.48 ± 0.52cd | 8.98 ± 0.36bc | 8.03 ± 1.46ab | 6.94 ± 0.48ab | 7.74 ± 0.07ab | 6.53 ± 0.09a |
3,4-Dimetho xytoluene | 1172 | - | - | - | - | - | - | 2.03 ± 0.29 | - | - | 3.09 ± 0.13 | 4.14 ± 0.84 |
DL-Menthol | 1173 | 48.93 ± 0.51ab | 58.07 ± 1.32bc | 74.09 ± 10.77d | 77.28 ± 11.84d | 67.90 ± 7.93cd | 45.88 ± 9.17ab | 48.32 ± 2.93ab | 46.46 ± 5.97ab | 45.21 ± 0.99ab | 39.29 ± 2.84a | 176.36 ± 15.6e |
Octanoic acid | 1173 | 14.04 ± 0.41c | 15.77 ± 0.98c | 26.17 ± 4.85de | 29.59 ± 4.36ef | 32.27 ± 2.50f | 23.99 ± 1.71d | 4.60 ± 0.34a | 7.59 ± 1.22ab | 7.69 ± 1.30ab | 9.55 ± 1.81b | 15.06 ± 2.67c |
(R)- 4-methyl-1-(1-meth ylethyl)-3-Cycl ohexen-1-ol | 1175 | 13.83 ± 0.27a | 19.14 ± 1.47b | 22.41 ± 3.93bc | 32.01 ± 5.03d | 31.49 ± 3.08d | 25.00 ± 2.56c | 19.52 ± 1.30b | 18.24 ± 3.07ab | 21.57 ± 1.50bc | 22.86 ± 0.80bc | 21.79 ± 1.69bc |
(3R,6S)-2,2,6-Trimethyl-6-vinyltetrahydro-2H-pyran-3-ol | 1183 | 8.88 ± 0.55c | 8.23 ± 1.24abc | 12.01 ± 2.03d | 14.64 ± 2.63e | 11.63 ± 1.26d | 8.25 ± 0.38abc | 7.13 ± 1.16abc | 6.01 ± 0.71a | 8.44 ± 0.70bc | 6.29 ± 0.56ab | 7.52 ± 0.01abc |
Methyl salicylate | 1187 | 39.34 ± 0.18 | 29.65 ± 2.16 | 35.28 ± 4.83 | 37.53 ± 3.98 | - | - | - | - | - | - | - |
4-trimethyl-α, α-Benzeneme thanol | 1188 | - | - | - | - | - | - | - | - | - | - | 7.23 ± 0.58 |
α-Terpineol | 1190 | 102.34 ± 0.95a | 136.06 ± 8.53bc | 165.38 ± 28.08d | 193.80 ± 8.27e | 200.51 ± 12.53e | 152.52 ± 11.32bcd | 125.56 ± 20.12ab | 103.74 ± 18.79a | 152.97 ± 13.65bcd | 133.27 ± 8.64bc | 159.20 ± 20.92cd |
Naphthalene | 1190 | 2.48 ± 0.16 | 3.40 ± 0.20 | 4.29 ± 0.63 | - | 7.94 ± 0.61 | - | - | - | 4.14 ± 0.18 | - | - |
1-ethyl-2,5-Pyrrolidinedione | 1191 | 2.90 ± 0.36 | - | - | 14.26 ± 2.61 | - | - | 6.88 ± 1.17 | 4.39 ± 0.86 | 6.94 ± 1.22 | 6.30 ± 0.38 | 5.52 ± 0.25 |
2,6,6-trimethyl-1,3-Cyclohexadiene-1-carboxaldehyde | 1205 | 24.66 ± 0.15d | 22.10 ± 1.60c | 26.42 ± 0.11de | 32.52 ± 3.37f | 35.64 ± 0.08g | 27.49 ± 2.06e | 20.81 ± 0.18c | 14.67 ± 0.09a | 17.96 ± 0.67b | 16.29 ± 1.32ab | 16.06 ± 0.77ab |
2,3-dihydro-Benzofuran | 1219 | 6.05 ± 0.18 | 5.11 ± 0.56 | 5.99 ± 0.93 | 7.64 ± 1.31 | 8.01 ± 1.3 | 4.69 ± 0.09 | 3.74 ± 0.58 | 2.98 ± 0.37 | 4.18 ± 0.48 | - | - |
2,6,6-trimethyl-1-Cyclohexene-1-carboxaldehyde | 1220 | 24.58 ± 0.27d | 17.79 ± 1.31c | 19.28 ± 1.60c | 26.38 ± 0.49e | 27.02 ± 1.39e | 13.92 ± 1.17b | 10.2 ± 0.25a | 8.32 ± 1.44a | 9.82 ± 0.26a | 9.07 ± 0.27a | 9.93 ± 1.29a |
(Z)- 3,7-dimethyl-2,6-Octadien-1-ol | 1229 | 13.34 ± 0.56a | 15.85 ± 1.18ab | 20.89 ± 3.5c | 30.34 ± 5.58d | 28.89 ± 4.37d | 19.98 ± 0.65bc | 12.41 ± 1.81a | 12.58 ± 2.31a | 16.28 ± 0.63abc | 14.45 ± 1.12a | 13.31 ± 1.17a |
Geraniol | 1258 | 43.95 ± 1.12bcd | 48.22 ± 3.67cd | 64.18 ± 11.11e | 78.40 ± 1.24f | 82.65 ± 3.82f | 51.55 ± 1.73d | 32.38 ± 6.4a | 31.83 ± 6.24a | 41.33 ± 2.65abc | 35.81 ± 2.48ab | 38.93 ± 7.02abc |
Nonanoic acid | 1272.1 | 23.46 ± 1.49c | 22.45 ± 1.56c | 41.20 ± 3.75ef | 36.43 ± 5.82e | 43.80 ± 4.25f | 28.84 ± 3.00d | 9.37 ± 1.39a | 10.63 ± 1.78ab | 14.64 ± 2.14ab | 15.60 ± 1.83b | 11.26 ± 0.02ab |
6-Undecanone | 1274 | 16.73 ± 0.62de | 13.70 ± 1.14bc | 16.81 ± 2.70de | 14.53 ± 0.10cd | 19.20 ± 3.09e | 11.11 ± 1.64b | 6.71 ± 0.69a | 8.20 ± 1.35a | 7.09 ± 0.79a | 7.56 ± 1.28a | 8.49 ± 0.18a |
Indole | 1294 | - | - | - | - | - | - | - | - | - | 1.26 ± 0.09 | - |
1-Tridecene | 1294 | - | - | 1.81 ± 0.25 | 2.63 ± 0.04 | 2.38 ± 0.27 | 0.94 ± 0.13 | 1.56 ± 0.15 | 1.49 ± 0.25 | 1.49 ± 0.26 | 1.37 ± 0.28 | 1.28 ± 0.02 |
2,6,10,10-tetramethyl-1-Oxaspiro[4.5]dec-6-ene | 1305.2 | 6.59 ± 0.89 | 6.91 ± 1.18 | 5.19 ± 0.52 | - | - | 4.41 ± 0.59 | - | - | 2.56 ± 0.30 | - | 3.61 ± 0.42 |
5,6,7,7a-tetrahydro-3,6-dimethyl-2(4H)-Benzofuranone | 1314 | 6.23 ± 0.89 | - | - | - | - | - | - | - | - | - | - |
1,2,3-Trimethoxybenzene | 1317 | - | - | 2.33 ± 0.32 | - | 3.47 ± 0.22 | 2.52 ± 0.16 | 2.88 ± 0.44 | 2.14 ± 0.30 | 0.92 ± 0.11 | 2.68 ± 0.24 | 1.90 ± 0.20 |
2,6-dimethoxy-Phenol | 1357 | 1.32 ± 0.06 | 1.84 ± 0.36 | - | - | - | - | - | - | - | - | - |
dihydro-5-pentyl-2(3H)-Furanone | 1362 | 7.83 ± 0.15f | 2.84 ± 0.17a | 5.15 ± 0.63d | 3.83 ± 0.63bc | 9.06 ± 0.17g | 6.00 ± 0.60e | 4.38 ± 0.50c | 3.08 ± 0.54ab | 3.07 ± 0.51ab | 5.79 ± 0.36de | 3.60 ± 0.26abc |
10-Undecenoic acid, methyl ester | 1371 | 13.9 ± 0.39 | 14.65 ± 1.05 | 16.25 ± 1.70 | - | 16.55 ± 1.88 | - | - | - | - | - | - |
Propanoic acid, 2-methyl-, | 1380 | 0.99 ± 0.07a | 1.09 ± 0.03a | 3.45 ± 0.50bc | 4.33 ± 0.05cd | 5.16 ± 0.73d | 2.87 ± 0.43b | 1.54 ± 0.30a | 9.35 ± 1.48e | 3.96 ± 0.54c | 4.34 ± 0.17cd | 3.98 ± 0.43c |
(E)-1-(2,6,6-trimethyl-1,3-cyclohexadien-1-yl)-2-Buten-1-one | 1388 | 3.38 ± 0.01ab | 4.05 ± 0.39bc | 5.4 ± 0.48d | 5.69 ± 0.69d | 6.73 ± 0.80e | 4.62 ± 0.07c | 2.99 ± 0.01a | 2.79 ± 0.25a | 3.10 ± 0.26a | 2.82 ± 0.01a | 2.93 ± 0.36a |
1-Tetradecene | 1392 | - | - | 0.62 ± 0.11 | - | - | - | 2.99 ± 0.07 | - | 1.20 ± 0.24 | - | 1.98 ± 0.32 |
6,10-dimethyl-2-Undecanone | 1407 | 3.55 ± 0.02 | 3.94 ± 0.33 | 4.05 ± 0.34 | - | 6.08 ± 0.39 | 5.47 ± 0.68 | - | 4.02 ± 0.26 | 4.20 ± 0.05 | - | - |
[3R-(3α,3aβ,7β,8aα)]− 2,3,4,7,8,8a-hexahydro-3,6,8 ,8-tetramethyl-1H-3a,7-Meth anoazulene | 1408 | - | - | 17.11 ± 2.37 | 20.25 ± 3.83 | 16.42 ± 1.83 | 20.17 ± 3.77 | 6.75 ± 0.57 | 6.85 ± 1.16 | 6.44 ± 0.97 | 10.93 ± 0.46 | 18.69 ± 3.47 |
Dodecanal | 1412 | - | 1.42 ± 0.20 | - | 4.66 ± 0.65 | 1.25 ± 0.20 | 1.18 ± 0.09 | 0.84 ± 0.14 | 0.98 ± 0.09 | - | 2.45 ± 0.14 | 1.27 ± 0.02 |
Longifolene | 1412.6 | - | - | - | - | 4.11 ± 0.66 | 2.94 ± 0.54 | 4.58 ± 0.39 | 3.23 ± 0.43 | - | 3.40 ± 0.71 | 1.43 ± 0.16 |
α-Ionone | 1421 | 15.43 ± 2.62b | 15.14 ± 0.62b | 15.13 ± 1.86b | 22.50 ± 4.34c | 20.08 ± 1.97c | 12.99 ± 0.56ab | 10.08 ± 0.28a | 11.30 ± 1.48a | 10.96 ± 0.53a | 13.04 ± 2.33ab | 15.44 ± 1.88b |
6-Methyl-6-(5-methylfuran-2-yl)heptan-2-one | 1426 | - | - | 3.79 ± 0.44 | 33.09 ± 5.04 | - | 2.83 ± 0.31 | - | - | - | - | - |
4-(2,2,6-trimethyl-7-oxabicyclo[4.1.0]hept-1-yl)-3-Buten-2-one | 1428 | - | - | 8.27 ± 1.49 | - | - | - | - | - | - | 2.53 ± 0.35 | - |
Ionone | 1429 | - | - | 23.66 ± 0.28 | - | - | 19.49 ± 1.88 | 10.24 ± 0.92 | 4.26 ± 0.54 | - | 4.04 ± 0.52 | - |
4-(2,6,6-trimethyl-1-cyclohexen-1-yl)-2-Butanone | 1433 | 4.48 ± 0.36 | - | 2.48 ± 0.32 | 2.75 ± 0.24 | - | - | - | - | - | - | 3.73 ± 0.25 |
6,10-dimethyl-5,9-Undecadien-2-one | 1434 | 34.61 ± 6.73d | 29.03 ± 0.59c | 31.08 ± 3.02cd | 16.66 ± 2.97ab | 32.01 ± 3.25cd | 20.95 ± 0.92b | 11.81 ± 0.46a | 13.46 ± 1.74a | 12.94 ± 0.14a | 14.56 ± 0.89a | 14.19 ± 2.36a |
4-(2,6,6-Trimethylcyclohexa-1,3-dienyl)but-3-en-2-one | 1440 | 8.04 ± 0.23a | 13.17 ± 0.81b | 17.77 ± 2.81c | 23.54 ± 1.55d | 26.62 ± 2.64e | 19.55 ± 0.75c | 11.73 ± 0.86b | 11.58 ± 1.50b | 13.08 ± 0.23b | 11.62 ± 0.47b | 13.19 ± 2.58b |
trans-β-Ionone | 1457 | 61.66 ± 2.27c | 59.18 ± 2.87c | 63.9 ± 9.36c | 56.33 ± 5.05c | 75.95 ± 5.35d | 47.80 ± 1.19b | 35.30 ± 2.34a | 28.51 ± 3.80a | 32.33 ± 0.55a | 28.8 ± 1.16a | 31.45 ± 5.12a |
1-Dodecanol | 1457 | 3.56 ± 0.06a | 5.59 ± 0.20b | 6.27 ± 0.75bc | 5.68 ± 0.74b | 9.00 ± 0.62d | 7.30 ± 0.22c | 5.71 ± 0.46b | 5.73 ± 1.13b | 7.46 ± 0.68c | 6.96 ± 0.93c | 7.22 ± 0.24c |
(+)-Valencene | 1491 | 4.85 ± 0.33 | 4.41 ± 0.29 | 5.4 ± 0.91 | - | 4.94 ± 0.35 | 2.21 ± 0.23 | 2.57 ± 0.09 | 2.00 ± 0.32 | - | - | - |
cis-hexahydro-8a-methyl-1,8(2H,5H)-Naphthalenedione | 1517 | 28.42 ± 0.95 | - | - | - | - | 22.57 ± 0.2 | - | - | 16.95 ± 0.67 | - | - |
Butylated Hydroxytoluene | 1517.5 | 1.39 ± 0.13 | 1.50 ± 0.08 | 1.66 ± 0.30 | 1.03 ± 0.02 | 0.99 ± 0.09 | 0.66 ± 0.13 | - | - | - | - | - |
(R)-5,6,7,7a-tetrahydro-4,4,7a-trimethyl-2(4H)-Benzofuranone, | 1525 | 31.72 ± 1.51ab | 33.55 ± 3.10abc | 43.40 ± 6.44bc | 72.09 ± 14.23d | 72.4 ± 10.57d | 45.21 ± 6.16c | 33.06 ± 1.47abc | 30.34 ± 5.95a | 44.20 ± 5.88bc | 34.16 ± 1.14abc | 33.48 ± 1.59abc |
(E)-3,7,11-trimethyl-1,6,10-Dodecatrien-3-ol | 1551 | 4.31 ± 0.14 | 7.80 ± 0.22 | - | - | - | 7.56 ± 0.24 | - | - | 4.03 ± 0.20 | - | 4.24 ± 0.15 |
2,4-Di-tert-butylphenol | 1555 | 16.86 ± 1.65a | 19.54 ± 1.14ab | 21.46 ± 3.21abc | 19.50 ± 3.05ab | 35.94 ± 2.87d | 26.11 ± 0.39c | 22.37 ± 1.31bc | 19.52 ± 2.73ab | 24.38 ± 3.57bc | 24.94 ± 4.67c | 24.68 ± 3.04bc |
n-Nonylcyclohexane | 1556 | - | - | - | - | - | - | - | - | 4.66 ± 0.29 | 6.94 ± 0.95 | 6.10 ± 0.88 |
Methanone, dicyclohexyl- | 1576 | 18.32 ± 0.71bcd | 18.43 ± 1.31bcd | 21.86 ± 3.22d | 21.84 ± 4.21d | 28.70 ± 3.46e | 19.76 ± 0.95bcd | 16.31 ± 2.43abc | 12.81 ± 2.04a | 18.14 ± 2.48bcd | 15.45 ± 0.71ab | 20.52 ± 3.81cd |
2,2,4-Trimethyl-1,3-pentanediol diisobutyrate | 1587.5 | 7.82 ± 0.15b | 15.79 ± 0.66g | 16.20 ± 0.46g | 10.78 ± 0.20de | 9.14 ± 0.68bc | 20.42 ± 0.46h | 5.73 ± 0.87a | 10.15 ± 1.67cd | 11.94 ± 0.60e | 13.91 ± 0.21f | 13.90 ± 1.77f |
Cedrol | 1607.9 | - | 11.23 ± 0.02 | 18.63 ± 1.21 | 7.36 ± 1.05 | 17.59 ± 1.29 | 12.45 ± 0.08 | 15.94 ± 0.56 | 30.98 ± 4.02 | 30.57 ± 2.25 | 37.09 ± 0.57 | 31.47 ± 4.95 |
Tetradecanal | 1614 | - | - | - | 1.63 ± 0.05 | - | - | 5.98 ± 0.23 | - | - | - | 7.23 ± 1.46 |
Benzophenone | 1664 | - | - | 14.89 ± 2.63 | 3.83 ± 0.12 | 9.30 ± 0.49 | 11.12 ± 1.08 | - | 8.18 ± 0.94 | 10.74 ± 0.29 | 7.49 ± 1.23 | 7.25 ± 0.74 |
2,2',5,5'-tetramethyl-1,1'-Biphenyl | 1668.5 | - | - | - | - | - | - | 1.56 ± 0.09 | 1.22 ± 0.18 | - | 1.22 ± 0.02 | 1.39 ± 0.16 |
2-hexyl-1-Decanol | 1790 | 5.40 ± 0.18 | - | - | - | - | - | - | - | 6.05 ± 0.77 | 9.29 ± 0.05 | 6.56 ± 1.18 |
3,4-diethyl-1,1'-Biphenyl | 1792 | - | 11.24 ± 0.26 | - | - | - | 28.4 ± 1.82 | - | 12.51 ± 1.53 | - | - | 10.7 ± 0.49 |
Neophytadiene | 1840 | 21.12 ± 0.02e | 22.90 ± 2.62ef | 25.74 ± 5.10f | 16.80 ± 1.99cd | 25.51 ± 0.77f | 19.01 ± 2.47de | 13.37 ± 2.20bc | 13.39 ± 2.31bc | 16.89 ± 0.72cd | 11.20 ± 0.05ab | 8.99 ± 1.41a |
Dodecylcyclohexane | 1864 | 2.73 ± 0.36a | 3.48 ± 0.13ab | 8.83 ± 1.48e | 6.67 ± 0.73d | 4.66 ± 0.41bc | 6.84 ± 0.55d | 5.97 ± 0.20cd | 4.67 ± 0.68bc | 4.80 ± 0.75bc | 5.84 ± 0.95cd | 4.01 ± 0.55b |
Phytol | 2045 | 16.50 ± 1.15de | 24.40 ± 2.06f | 23.47 ± 1.53f | 14.73 ± 2.26cd | 16.69 ± 1.21de | 18.51 ± 3.07e | 14.36 ± 2.81cd | 11.28 ± 2.02bc | 13.83 ± 0.87cd | 8.48 ± 1.24ab | 7.63 ± 0.99a |
Decanoic acid, methyl ester | 1282 | - | - | - | 16.38 ± 1.25 | - | 14.87 ± 1.95 | 10.06 ± 0.38 | 9.18 ± 0.42 | - | - | - |
n-Hexadecanoic acid | 1968 | - | - | - | - | 40.59 ± 4.43 | - | - | - | - | - | - |
1-Decanol | 1258 | - | - | - | - | 9.67 ± 0.92 | 5.48 ± 0.3 | - | 4.07 ± 0.29 | - | - | - |
Sulfurous acid, butyl tetradecyl ester | 2434 | 3.34 ± 0.56 | 2.15 ± 0.19 | - | - | 2.22 ± 0.20 | 1.87 ± 0.08 | - | - | - | 1.25 ± 0.15 | 1.33 ± 0.12 |
a RI = Retention index of aroma compounds on HP-5MS UI Column; RI were obtained from NIST Chemistry WebBook. (https://webbook.nist.gov/chemistry/name-ser/) |
b Concentrations of aroma compounds in T0, T1, T2, T3, T4, T5, T6, T7, T8, T9 and T10 were represented RAPT samples from eleven different aging years, respectively; values with different letters (a-h) in a row are significantly using Duncan’s multiple comparison tests ( p < 0.05 ) . |
c All the above aroma compounds were identified by MS and RI values, except. Phytol, Decanoic acid, methyl ester, n-Hexadecanoic acid, 1-Decanol and Sulfurous acid, butyl tetradecyl ester is identified by MS. |
As illustrated in Fig. 1 and Table 1, the composition and contents of volatile compounds in RAPT underwent apparent changes during storage. At the same years of storage, the difference in contents between compounds was also significant (p < 0.05). As shown in Fig. 1C, alcohol compounds are the most abundant component, which is independent with the years of storage. Most of the alcohols in tea, such as benzyl alcohol, phenylethyl alcohol and (Z)-3,7-dimethyl-2,6-octadien-1-ol, contribute to the rose-like and citrus-like aroma characteristics. These alcohols are produced from the degradation of Strecker aldehydes or the hydrolysis of glycoside precursors (Zhu et al., 2021). In addition to alcohols, aldehydes, ketones, alkenes and esters also showed relative high contents in RAPT. The total concentration of volatiles generally fluctuated during storage (Fig. 1C). Moreover, after 5 years of storage, the total volatile contents decreased and although the overall content remains relatively stable, it is still lower compared to the samples with shorter storage durations. This distinction may be attributed to the differences between RAPT and other tea processing techniques.
To investigate the variation in the volatile profile of RAPT samples stored for different durations, this work conducted principal component analysis (PCA) and hierarchical cluster analysis (HCA) using the semi-quantitative results of 64 volatile compounds. The PCA scoring plot (Fig. 1D) demonstrates the clear differentiation of the samples based on the first two principal components, which collectively account for 65.3% of the total variance. The RAPT samples can be broadly categorized into four distinct groups, indicating significant changes in the volatile profile during storage. The results of HCA also revealed a clear classification of the samples into four categories: T0, T1, T2 and T5 formed one category, T3 and T4 formed another category, T6, T7, T8 and T9 constituted a third category, and T10 represented a separate category (Fig. 1E). Notably, the samples with a storage duration of 10 years (T10) exhibited a distinct separation from other samples, suggesting a significant change in the volatile composition following a decade of storage. In addition, the samples with a storage duration of 1–5 years were predominantly located in the left region of the scoring plot. Overall, the volatile profile of RAPT may experience a significant shift after 3–5 years of storage, representing the first turning point, followed by another notable transformation after 10 years of storage, signifying the second turning point. The loading plot (Fig.S2) indicated that volatile compounds underwent significant changes during the storage period, particularly in linalool, α-terpineol, 3,7-dimethyl-1,5,7-octatrien-3-ol, geraniol, cis-5-ethenyltetrahydro-α,α,5-trimethyl-2-furanmethanol, and (R)-5,6,7,7a-tetrahydro-4,4,7a-trimethyl-2(4H)-benzofuranone. These results further demonstrated the non-negligible influence of storage on the volatile composition of RAPT.
3.2 Dynamic changes of different compounds during storage
As indicated by the quantitative results, the contents of many volatile compounds varied greatly during the storage of RATP. To visually illustrate the change patterns of volatile compounds in RAPT for different storage years, this study employed heatmap analysis, as depicted in Fig. 2.
As illustrated in Fig. 2, three distinct patterns of changes can be observed in the content of these compounds. The first pattern consists of a gradual decrease in content over time, the second pattern involves an increase in content with increasing storage time, and the third pattern exhibits an initial increase followed by a gradual decrease after 5 years. The first group of volatile compounds includes 1-octen-3-ol, 3,5,5-trimethy-2-hexene, (E)-2-octen-1-ol, 1-hexanol, 2-methl-butanoic acid, 3-methyl-butanoic acid, and 6,10-dimethyl-5,9-undecadien-2-one. These seven volatile compounds showed a tendency to decrease in content during storage, and their dominant aroma characteristics were described as mushroom-like, green, roasted, fatty and cheese-like. It is worth noting that most of these compounds belong to the alcohol chemical class, which is an interesting finding. Alcohols have been found to be the main volatile compounds in RAPT and contribute significantly to its characteristic aroma (Fan et al., 2021). Thus, it is hypothesized that tea with shorter storage times may have stronger grassy, roasted, and fatty aromas. In addition, the second group of compounds includes α-pinene, D-limonene, 2-ethyl-1-hexanol, and DL-menthol. These volatile compounds are mainly associated with a woody and mint-like aroma. This finding explains the disappearance of green and roasted aroma, and the emergence of woody and mint-like aroma in RAPT. Furthermore, most of these compounds showed a pattern towards the third group of compounds (approximately 75%). For instance, benzaldehyde increased from 14.77 µg /100g (stored for 0 years) to 46.78 µg /100g (stored for 5 years) and then slowly decreased. Similarly, 2,4-Di-tert-butylphenol (16.86–35.94) increased from 16.86 to 35.94 (µg /100g) initially and then slowly decreased. Here, benzaldehyde is produced through the deamination of amino acids by aminotransferase (Y. Liu et al., 2023). Interestingly, a majority of the third group of volatile compounds decreased in 5 years of storage (T5), which is consistent with a previous study indicating that five years of storage is a significant turning point (Ren et al., 2022). Moreover, most of the third group of volatile compounds contribute to the floral and fruity aroma. This result suggested that the green and roasted aromas diminish over time, while the floral and fruity aromas gradually become more prominent during the 0–5 years of storage.
FigureS3 shows a decrease in the relative contents of 12 volatile compounds during storage. These compounds include 1-hexanol, 2,6-dimethy-5-heptenal, 1-octen-3-ol, 2,3-octanedione, 2,6,6-trimethyl-1,3-cyclohexadiene-1-carboxaldehyde, 3,5,5-trimethyl-2-hexene, (E)-2-octen-1-ol, 2-methyl-butanoic acid, 1-ethyl-1H-pyrrole-2-carboxaldehyde, 6,10-dimethyl-5,9-undecadien-2-one, trans-β-ionone, 6-undecanone. Specifically, they predominantly green, fatty, floral, fruity and roasted aroma. Moreover, the relative contents of some volatile compounds have been increased during storage, notably α-ionone, isophorone, 1,2-dimethoxy-benzene, 1,2,5,5-trimethyl-1,3-cyclopentadiene. These compounds contribute to violet-like, woody, fruity, vanilla and phenolic-like aromas. It is speculated that these changes in volatile compounds may explain the development of a woody aroma in RAPT with extended storage periods.
3.3 Aroma characteristics of RAPT during storage
Odor Activity Values (OAV) is a quantitative measure used to assess the odor potency of a compound or mixture, and is typically determined by measuring the concentration of a volatile compound in a sample and comparing it to a corresponding odor threshold. Based on the OAV, the contribution of each volatile compound can be evaluated, and the correlation between the quantified volatile compounds and the aroma can be determined. In general, aroma compounds with OAV greater than or equal to one are considered as active aromatic compounds and are believed to significantly contribute to the overall aroma profile of tea (Russo et al., 2020; Yin et al., 2022). It is also important to consider the relative contribution of the components in a mixture, in addition to assessing the contribution of individual compounds. Therefore, the contribution of the mixed compounds to the aroma was further analyzed by calculating the aroma contribution index (ACI). Based on an OAV of ≥ 1, fourty-three volatile compounds were identified as potential aroma compounds. (Table 2). Figure 3. shows that the change of these 43 volatile compounds during storage. Interestingly, 22 of them had the highest OAV at T3 or T4 (after 3 or 4 years of storage). In addition, according to their different aroma characteristics, the 43 volatiles were classified into nine groups: green, fruity, floral, roasted, fatty, chemical, cheese-like, stale, and woody. These aroma attributes have been previously used to describe the aroma characteristics of RAPT in other studies.
Table 2
Key volatile compounds identified in RAPT samples from different years of storage
Volatile compounds | Class | Odor | Threshold(µg/kg) | OAV | | | | | | | | | | | ACI(%) |
| | | | T0 | T1 | T2 | T3 | T4 | T5 | T6 | T7 | T8 | T9 | T10 | |
DL-Menthol | Woody | mint-like,woody | 1000 | 0.5 | 0.6 | 0.7 | 0.8 | 0.7 | 0.5 | 0.5 | 0.5 | 0.5 | 0.4 | 1.8 | < 0 |
Linalool | Floral&fruity | Citrus-like, flowery | 0.6 | 4525.1 | 4225.6 | 4591.1 | 8040.2 | 5534.9 | 3857.9 | 3043.5 | 2691.1 | 3483.4 | 3360.8 | 3075.5 | 0.72 ~ 8.28 |
α-Terpineol | Floral | flowery | 4.08 | 250.8 | 333.5 | 405.3 | 475 | 491.5 | 373.8 | 307.7 | 254.3 | 374.9 | 326.7 | 390.2 | 0.06 ~ 0.63 |
Geraniol | Floral&fruity | Rose-like, citrus-like | 3.2 | 137.3 | 150.7 | 200.6 | 245 | 258.3 | 161.1 | 101.2 | 99.5 | 129.1 | 111.9 | 121.7 | 0.03 ~ 0.25 |
trans-Linalool oxide (furanoid) | Floral | floral | 3000 | 0.2 | 0.1 | 0.2 | 0.3 | 0.2 | 0.2 | 0.1 | 0.1 | 0.1 | 0.1 | 0.2 | < 0 |
trans-β-Ionone | Floral | Flowery, violet-like | 0.007 | 88081.6 | 84544.4 | 91283.8 | 80478.2 | 108493 | 68278.6 | 50430.3 | 40730.3 | 46186 | 41143.2 | 44931.8 | 14.41 ~ 85.82 |
D-Limonene | Fruity | Citrus-like, carrot-like | 13 | 34.1 | 52.5 | 57.6 | 21.4 | 57.3 | 44.1 | 35 | 42.6 | 40.8 | 53.4 | 33.8 | 0.01 ~ 0.09 |
Benzaldehyde | Fruity | Cherry-like | 320 | 0.5 | 0.6 | 1.1 | 0.8 | 1.5 | 0.8 | 0.6 | 0.6 | 0.7 | 0.7 | 0.8 | < 0 |
1H-Pyrrole-2-carboxaldehyde, 1-ethyl- | Roasted | Bread-like, roasty | 65000 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0 |
Octanoic acid | Cheese flavor | Carrot-like, musty Cheese, fat | 190 | 0.7 | 0.8 | 1.4 | 1.6 | 1.7 | 1.3 | 0.2 | 0.4 | 0.4 | 0.5 | 0.8 | < 0 |
α-Ionone | Floral | Flowery, violet-like | 0.4 | 385.8 | 378.5 | 378.1 | 562.5 | 501.9 | 324.9 | 251.9 | 282.4 | 274.1 | 326 | 386 | 0.06 ~ 0.62 |
Benzeneacetaldehyde | Floral | Lilac and hyacinth-like | 4 | 29.8 | 24.9 | 30 | 56.7 | 56.1 | 29.1 | 30.6 | 30.6 | 48.1 | 35.3 | 42.9 | 0 ~ 0.06 |
2,6,6-trimethyl-1,3-Cyclohexadiene-1-carboxaldehyde | Woody | Woody,herbal | 3 | 82.2 | 73.7 | 88.1 | 108.4 | 118.8 | 91.6 | 69.4 | 48.9 | 59.9 | 54.3 | 53.5 | 0.01 ~ 0.12 |
(E,E)-2,4-Heptadienal | Floral | Fatty, flowery | 0.032 | 3598.6 | 3248.6 | 3338.5 | 4774.2 | 5892 | 3473.1 | 4117.8 | 3857.1 | 4501.8 | 4416.6 | 4351.2 | 0.55 ~ 7.76 |
1,2-dimethoxy-Benzene | Stale | Sweet and Musty | 3.17 | 20.5 | 37.4 | 53.2 | 54.2 | 54.4 | 51.1 | 30.5 | 26.1 | 25.2 | 33.6 | 42.7 | 0.01 ~ 0.06 |
(Z)-3,7-dimethyl-2,6-Octadien-1-ol | Floral&fruity | Citrus-like,Floral | 49 | 2.7 | 3.2 | 4.3 | 6.2 | 5.9 | 4.1 | 2.5 | 2.6 | 3.3 | 2.9 | 2.7 | 0 ~ 0.01 |
6-methyl-5-Hepten-2-one | Fruity | Citrus-like, apple-like | 50 | 3.9 | 3 | 2.6 | 3.3 | 4.4 | 3.5 | 2.2 | 2 | 1.9 | 2.7 | 2.6 | 0 ~ 0.01 |
1-Octanol | Floral&fruity | Rose-like or lemon-like | 0.8 | 255.1 | 317.3 | 346 | 620.8 | 497.9 | 336.4 | 106.6 | 95.2 | 158.4 | 126.8 | 131.8 | 0.06 ~ 0.64 |
Nonanoic acid | Stale | Moldy, pungent | 26 | 9 | 8.6 | 15.8 | 14 | 16.8 | 11.1 | 3.6 | 4.1 | 5.6 | 6 | 4.3 | 0 ~ 0.01 |
2,6,6-trimethyl-1-Cyclohexene-1-carboxaldehyde | - | - | 5 | 49.2 | 35.6 | 38.6 | 52.8 | 54 | 27.8 | 20.4 | 16.6 | 19.6 | 18.1 | 19.9 | 0.01 ~ 0.05 |
1-Octen-3-ol | Green | Mushroom-like, green | 45 | 21.3 | 11.5 | 5.9 | 5.1 | 8.2 | 4.8 | 2.6 | 2.7 | 2.8 | 3.8 | 3.4 | 0 ~ 0.01 |
1-(1H-pyrrol-2-yl)-Ethanone | Nutty | Nutty, musty | 3.4 | 41.2 | 39.3 | 39 | 75.9 | 68 | 49 | 34.8 | 35.3 | 38.1 | 36.4 | 29.2 | 0.01 ~ 0.08 |
Octanal | Fruity | Citrus-like, soapy | 0.0455 | 1600.9 | 1455.3 | 1320.7 | 1376.8 | 4131.4 | 2432.5 | 1534.7 | 1365 | 1880.6 | 1836.4 | 1632.4 | 0.25 ~ 3.52 |
1-Dodecanol | Fatty | Fatty, soapy | 6.1 | 5.8 | 9.2 | 10.3 | 9.3 | 14.8 | 12 | 9.4 | 9.4 | 12.2 | 11.4 | 11.8 | 0 ~ 0.02 |
1-Nonanol | Floral | rose-like | 11 | 9.8 | 7.6 | 10.5 | 15.8 | 17.9 | 9.5 | 8.2 | 7.3 | 6.3 | 7 | 5.9 | 0 ~ 0.02 |
Heptanal | Fatty | Citrus-like, fatty | 9.7 | 4.7 | 4.7 | 8 | 6.9 | 9 | 7.3 | 7.2 | 3.5 | 7.8 | 5.9 | 5.3 | 0 ~ 0.01 |
Isophorone | Woody | Woody | 3000 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0 |
Dihydro-5-pentyl-2(3H)-Furanone | Cheese flavor | Coconut, creamy, waxy with fatty milky notes | 4.5 | 17.4 | 6.3 | 11.5 | 8.5 | 20.1 | 13.3 | 9.7 | 6.8 | 6.8 | 12.9 | 8 | 0 ~ 0.02 |
(E)-1-(2,6,6-trimethyl-1,3-cyclohexadien-1-yl)-,2-Buten-1-one | Floral&fruity | apple-like, rose-like | 1000 | < 0.1 | < 0.1 | 0.1 | 0.1 | 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0 |
2-Heptanone | Fruity | cabbage-like | 70 | 0.6 | 0.6 | 0.4 | 0.5 | 0.8 | 0.7 | 0.4 | 0.3 | 0.4 | 0.6 | 0.3 | < 0 |
3-methyl-Butanoic acid | Cheese flavor | Cheese-like | 10000 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0 |
1-Hexanol | Green | Green, grassy | 40 | 1.4 | 1.4 | 0.8 | 1 | 1.2 | 0.8 | 0.6 | 0.5 | 0.8 | 0.4 | 0.6 | < 0 |
p-Xylene | Chemical | Plastic, green, | 60 | 0.3 | 0.3 | 0.3 | 1 | 0.7 | 0.3 | 0.4 | 0.4 | 0.5 | 0.4 | 0.4 | < 0 |
2-methyl-Butanoic acid | Cheese flavor | Butter, cheese-like | 15 | 3.4 | 3 | 1.6 | 2.2 | 2.1 | 1.7 | 0.4 | 0.7 | 0.4 | 0.9 | 1 | < 0 |
α-Pinene | Woody | Fresh, woody | 48 | 0.1 | 0.1 | 0.1 | 0.1 | 0.2 | 0.2 | 0.2 | 0.2 | 0.2 | 0.2 | 0.2 | < 0 |
(Z)-3-Hexen-1-ol | Green | Grass | 0.0002 | 151243.8 | 145866.1 | ND | ND | ND | ND | ND | ND | ND | ND | ND | 0 ~ 40.33 |
1-ethyl-1H-Pyrrole | Roasted | Burnt, roasted | 3 | 4.6 | 3.2 | ND | ND | ND | ND | ND | ND | ND | ND | ND | < 0 |
Methyl salicylate | Green | wintergreen | 10 | 39.3 | 29.7 | 35.3 | 37.5 | ND | ND | ND | ND | ND | ND | ND | 0 ~ 0.04 |
Phenylethyl Alcohol | Floral&fruity | Rose-like | 9 | 24.9 | 21.9 | 29.4 | 38 | 35.7 | 19.1 | ND | 14 | 7 | 15.3 | ND | 0 ~ 0.04 |
β-Myrcene | Woody | Pepper-like,carrot-like | 0.75 | ND | 239.3 | ND | ND | ND | 280.2 | ND | ND | ND | 215.1 | 171.3 | 0 ~ 0.41 |
Tetradecanal | Chemical | Alkane | 60 | ND | ND | ND | 0.3 | ND | ND | 1 | ND | ND | ND | 1.2 | < 0 |
1-Decanol | Floral&fruity | orange-like, floral | 23 | ND | ND | ND | ND | 4.2 | 2.4 | ND | 1.8 | ND | ND | ND | < 0 |
Naphthalene | Chemical | pungent dry tarry | 50 | 0.5 | 0.7 | 0.9 | ND | 1.6 | ND | ND | ND | 0.8 | ND | ND | < 0 |
2,3-dihydro-Benzofuran | Chemical | Tar-like, pungent | 48 | 1.3 | 1.1 | 1.2 | 1.6 | 1.7 | 1 | 0.9 | 0.6 | 0.9 | ND | ND | < 0 |
1-Oxaspiro[4.5]dec-6-ene, 2,6,10,10-tetramethyl- | Woody | Herbal | 0.0002 | 329594.6 | 345510 | 259316.6 | ND | ND | 220607.8 | ND | ND | 128021.3 | ND | 180540.2 | 0 ~ 76.48 |
Benzyl alcohol | Floral&fruity | Rose-like, cherry-like | 5500 | < 0.1 | ND | ND | < 0.1 | ND | ND | ND | ND | ND | ND | ND | < 0 |
Ionone | Floral | Flowery, violet-like | 1500 | ND | ND | 0.2 | ND | ND | 0.1 | 0.1 | < 0.1 | ND | < 0.1 | ND | < 0 |
(E)-2-Octenal | Green | Green | 3 | ND | ND | ND | ND | ND | ND | ND | 7 | ND | 9.6 | ND | 0 ~ 0.02 |
2,6-dimethoxy-Phenol | Chemical | phenolic, smoky, woody | 400 | v | < 0.1 | ND | ND | ND | ND | ND | ND | ND | ND | ND | < 0 |
(E,E)-2,6-Nonadienal | Green | cucumber-like, grren | 1 | ND | ND | ND | ND | ND | 18.9 | ND | 13.8 | ND | ND | 14.9 | 0 ~ 0.03 |
Dodecanal | Green | Green,soapy,citrus-like | 1.07 | ND | 13.3 | ND | 43.5 | 11.7 | 11.1 | 7.8 | 9.2 | ND | 22.9 | 11.9 | 0 ~ 0.04 |
Indole | Floral | Floral, animal-like | 500 | 0.1 | ND | < 0.1 | ND | ND | ND | ND | ND | < 0.1 | < 0.1 | ND | < 0 |
Hexanoic acid, methyl ester | Fruity | Pine apple-like | 10 | ND | ND | ND | ND | 1 | ND | 0.4 | 2.4 | 0.5 | ND | ND | < 0 |
2-Octanone | Woody | Woody, Herbal | 50 | 0.1 | ND | ND | ND | ND | 0.3 | ND | ND | ND | ND | ND | < 0 |
2-butoxy-Ethanol | - | - | 2600 | ND | < 0.1 | ND | ND | < 0.1 | ND | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0.1 | < 0 |
Hexanal | Green | Green | 9 | ND | 0.5 | 6.7 | ND | ND | 1.5 | 0.5 | 0.6 | 0.6 | 0.7 | 0.7 | < 0 |
1,2,3-Trimethoxybenzene | Stale | Stale and Musty | 0.75 | ND | ND | 31.1 | ND | 46.3 | 33.6 | 38.4 | 28.5 | 12.3 | 35.7 | 25.3 | 0 ~ 0.07 |
a OT = Odor thresholds, |
b ND, the compounds was not detected. |
c All the odor thresholds were obtained from: (Guo et al., 2021; Hao et al., 2023; Tatsu et al., 2020; Zhai et al., 2022; Zheng et al., 2023; Zhu et al., 2021) |
Table 2 shows the aroma compounds in RAPT decreased with increasing storage time, including 1-octen-3-ol, 2-methyl-butanoic acid, 1-ethyl-1H-pyrrole. These compounds are related to green and roasted aroma attributes. On the one hand, linalool (OAV = 8040.2) and 1-octanol (OAV = 620.8) exhibited the highest OAV at T3 (3 years of storage). (E, E)-2,4-heptadienal (OAV = 5892) and octanal (OAV = 4131.4) had the highest OAV at T4 (4 years of storage). On the other hand, DL-menthol (OAV = 1.8) reached its maximum value at T10 (10 years of storage) and was characterized by a peppermint and woody aroma. This could explain the formation of woody and mint-like aroma in RAPT after 10 years of storage. Besides, (Z)-3,7-dimethyl-2,6-octadien-1-ol was found to contribute to the aroma profile of RAPT during storage. Its OAVs gradually increase from 2.7 at T0 to a peak value (6.2) at T3, before slowly decreasing with increasing storage time to 2.7 at T10. Furthermore, it is interesting to note that α-ionone (OAV = 562.5) and linalool (OAV = 8040.2) has violet- like, citrus-like, and flowery aroma characteristics, reaching their peak levels at T3. Meantime, geraniol (OAV = 258.3), octanal (OAV = 4131.4), and α-terpineol (OAV = 491.5) reached the highest at T4, displaying rose-like, citrus-like, and flowery aroma characteristics. This suggests that the storage of RAPT for about 3 to 4 years results in an abundance of floral and fruity aroma. Moreover, the contribution of aroma compounds in the mixture was compared using the aroma contribution index (ACI). Five volatile compounds were screened based on thier high OAV (≥ 100) and ACI (≥ 1). These compounds include linalool (OAV:2691.1–8040.2, ACI:0.72–8.28), (E,E)-2,4-heptadienal (OAV: 3248.6–5892, ACI:0.55–7.76), (Z)-3-hexen-1-ol (OAV: 0–151243.8, ACI: 0–40.33), 2,6,10,10-tetramethyl-1-oxaspiro[4.5]dec-6-ene (OAV: 0–345510, ACI: 0–76.48), and octanal (OAV: 1320.7–4131.4, ACI: 0.25–3.52).
3.4 Identification of the key aroma characteristics of RAPT during storage
Floral, fruity and green aromas are considered as favorable aroma characteristics in tea. Therefore, we analyzed the floral and fruity aromas of volatile compounds in RAPT. In Fig. 4A, it is shown that most of the volatile compounds in RAPT have pleasant floral and fruity aromas during storage. Previous results have also indicated that the floral and fruity aromas were increased over time during the first four years of storage. RAPT is unique compared to other teas because it undergoes autoclaving and compressing treatments. These treatments result in an increase in the content of certain aroma compounds such as phytol, linalool and geraniol, which have a significant contribution to the floral aroma of tea (Su et al., 2022). Consequently, RAPT has a distinctive floral and fruity aroma which may be dramatically changed during storage. As shown in Fig. 4B, the roasted aroma, particularly that of 1-ethyl-1H-pyrrole, gradually diminished over the storage period. After ten years of storage, the green aroma decreased, while the woody aroma increased. However, it is important to note that t the floral and fruity aromas remained dominant, indicating their overall persistence in the RAPT.
In addition, the intensity of the floral and fruity were gradually enhanced during storage. Hence, eight aroma compounds were screened with OAV ≥ 100, namely linalool (OAV:2691.01–8040.2), α-terpineol (OAV:250.8–491.5), geraniol (OAV:99.5–258.3), trans-β-ionone (OAV:40730.3–108493), α-ionone (OAV:274.1–562.5), (E,E)-2,4-heptadienal (OAV:3248.6–5892), 1-octanol (OAV:95.2–620.8), octanal (OAV:1320.7–4131.4). Interestingly, all of them contribute to the aroma of floral and fruity. Notably, (E,E)-2,4-heptadienal is one of the dominant volatile compounds in high-grade Jingshan green tea, and it is produced through the degradation of linolenic acid, providing flowery aroma (Flaig et al., 2020). In oolong tea (Camellia sinensis), geraniol and linalool are the main contributors to the floral, rose-like and sweet aroma characteristics. In addition, Yang et al. (2021) found that trans-β-ionone contributed to the aroma characteristics of Rougui tea (P. Yang et al., 2021). Therefore, these eight volatile compounds are considered to be the key contributors to the floral and fruity characteristics during RAPT storage.
3.5 Metabolic pathways of the key aroma compounds
The changes in aroma characteristics may be influenced by several factors during storage. Geraniol and linalool, which are monoterpenes, serve as the primary constituents responsible for the pleasant floral and fruity scent found in plants. These compounds are synthesized in tea through the metabolic conversion of two primary precursors, namely isopentenyl diphosphate (IPP) and dimethylallyl pyrophosphate (DMAPP). The mevalonate pathway (MVA) plays a critical role in the synthesis of secondary metabolites, including sterols, sesquiterpenes, and triterpenes within the cytoplasm and endoplasmic reticulum. The 2-C-Methyl-D-erythritol-4-phosphate pathway (MEP) and carotenoid cleavage dioxygenase (CCD) are two key metabolic pathways involved in the biosynthesis of monoterpenes and oxidation of carotenoids, respectively (Schwab et al., 2008). It is hypothesized that changes in the metabolism of microorganisms during storage may result in changes in aroma characteristics. Trans-β-ionone and α-ionone are secondary oxidation products derived from carotenoids, including β-carotene, α-carotene, phytoene, lutein, and lycopene, which undergo oxidation by CCD to form primary oxidation products and then further converted to secondary oxidation products (Y. Liu et al., 2023). Furthermore, the conversion of linolenic acid into (E,E)-2,4-heptadienal occurs through the involvement of leucoanthocyanidin reductase (LOX) and glutathione peroxidase (HPL). A previous work has shown that LOX continues to facilitate the synthesis of aldehydes from fatty acids throughout the fermentation stage (Q. Chen et al., 2022). The storage of RAPT, in addition to being a natural aging and fermentation process, is also believed to contribute to the changes in aroma. These changes may be attributed to microbial metabolism, enzymatic digestion and natural oxidation.