Spatial-temporal variability of volatile components of Camellia azalea Wei[1]

LI Xin-lei^1,2 LI Ji-yuan¹ SUN Zhen-yuan² FAN Zheng-qi¹ YIN Heng-fu¹
1. Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Fuyang Zhejiang 311400, China
2. State Key Laboratory of Tree Genetics and Breeding, Research Institute of Forestry, Chinese Academy of Forestry, Beijing 100091, China

Introduction

Camellia, one of the top ten traditional famous flowers, originated from China (Zhang et al, 1998). Except the ornamental value, Camellia also has edible and medical functions. Cultivation and application of Camellia have a long history in China, and its ornamental, edible and medical value has been confirmed by related data (Gao et al, 2005; Liang et al, 2005). Studies have shown that the main beneficial ingredients in Camellia flowers were carbohydrates, protein, vitamins, trace elements, amino acids and flavonoids (Liang et al, 2005; Li et al, 2010). The beverages and foods on C. nitidissima have been developed (Liang et al, 2005). Studies on Rosa rugosa, Michelia alba and Jasminum sambac showed that the volatile components of flower were composed of alcohols, alkenes, esters and alkanes, etc. (Guth et al, 1993; Barkman et al, 1999; Guterman et al, 2002; Shang et al, 2002). The volatile components of three kinds of Camellia have been determined (Fan et al, 2005; 2006). C. azalea Weihas long flowering period and the main florescence is from April to December. But in appropriate cultivation conditions, C. azalea could bloom throughout the whole year (Gao et al, 2005), and this unique property make this variety a good material for extracting the volatile components of the flower. The flower volatile components of C.azaleahave not been studied until now. The volatile components in different florescence and flower parts of C. azalea were analyzed by solid phase micro-extraction and GC/MS technology, in order to make clear the variation of the volatile components and provide a scientific basis for its further development and utilization.

1 Materials and Methods

1.1 Plant material

The florescence of C. azalea was divided into four different stages, including of bud stage, first flowering stage, flowering stage and declining stage. In bud stage, most flower buds fully extend bract and the color has turn red. In the first flowering stage, petals are going to spread out and the stamen is still wrapped in petals. In the flowering stage, the petals fully open and the anthers are mature. During the declining stage, the petals are wrinkling and their texture begins to thin. The flower in flowering stage of C. azalea was divided into four parts, including of sepals, petals, stamens and pistils. The volatile components should be determined within 30 minutes after the samples were collected under the same condition.

1.2 Methods

The collected samples were placed in a sample vial, and 0.5μL decanoic acid ethyl ester of 40ng/μL was added as an internal standard substance. The samples were extracted 30 min under 40℃ by 65 µm PDMS/DVB SPME injections using solid phase micro-extraction method. The instrument was 6890N/5975B gas chromatography-mass spectrometry. Ionization mode was electron ionization with electron energy of 70eV, and injection port temperature was 250 ℃. The column temperature was programmed at 35 ℃ for 2 min, and then increased to 80℃ at 5 ℃/min, to 180 ℃and 250 ℃ at 8 ℃/min. Quadropole temperature was 150 ℃.Ion source temperature was 230 ℃, and interface temperature was 280 ℃. Mass scan range was 30～500 u. The volatile compounds were identified basing on the search results of mass spectrometry data and GC-MS standard atlas database. The relative content of each component was determined according to the ratio of the peak area of each compound and the internal standard.

2 Results

2.1 The volatile components of C. azalea in different florescence

Volatile compounds and their relative content in different florescence of C. azalea were showed in Table 1. There were 8 components identified in bud stage, 20 in first flowering stage, 21 in full flowering stage and 17 in declining stage. The relative content of octane, nonanal, caryophyllene, naphthalene and thujopsene was higher than the other components, and that increased first and then decreased with the flower blooming. The main releasing period of the five components were first flowering and full flowering stages, accounting for 65.92% and 61.02% of total volatile compounds respectively. Thujopsene, Copaene, Naphthalene, 3-Phenyl-2-propenal and 1,2,3,4,5-Pentamethyl-1,3-cyclopentadiene only existed in first flowering and full flowering stages, and alpha.-Cedrene only existed in first flowering stage. The relative content of dimethyl ether and 2-Propenal was higher in bud stage, and decreased gradually with the flower blooming. The relative content of 2-{4-[2-(4-Methoxymethylphenyl)vinyl]phenyl}propan-2-ol increased gradually with the flower blooming.

Table 1  The main volatile components and relative content of C. azalea in different florescence

Note: “-”: Not identified, the same below.
B.S: Bud stage; F.F.S: First flowering stage; F.S: Flowering stage; D.S: Declining stage, the same below.

The volatile compounds of C. azalea in different florescence were divided into six types, including of alkenes, alcohol, aldehydes and ketones, esters, alkanes and others such as ether, acids and amines. The change of relative content of various compounds in different florescence was showed in Figure 1. The relative content of alkenes was the highest among the six types of compounds, followed by alkanes and aldehydes and ketones, accounting for 44.17%, 16.42% and 16.38% of the total components respectively. Alcohols and esters accounted for 9.08% and 4.50% respectively. The relative content of alkenes, esters and alkanes first increased and then decreased with the flower blooming, that in first flowering and full flowering stage accounting for 89.16%, 81.09% and 62.62% of the total content of four flowering stages. The relative content of alcohols increased with the flower blooming, accounting for 56.89% in first flowering and full flowering stages together. The relative content of aldehydes and ketones decreased with the flower blooming, accounting for 40.96% in first flowering and full flowering stages together.

2.2 The volatile components of C. azalea in different flower parts

The relative content of each volatile compound in different flower parts of C. azalea was showed in Table 2. There were 20 components identified in sepals, 22 in petals, 21 in stamens and 13 in pistils. Compared with petals and stamens, there were fewer volatile components in sepals and pistils, and their relative content was also lower, that showed that the petals and stamens were the main parts of volatile components releasing of C. azalea. There were seven same compounds in the four flower parts, and the relative content of each compound was far higher in petals and stamens than that in sepals and pistils. For example, the relative content of Hexane in petals and stamens was 9.3435 and 6.6015 respectively, and that in sepals and pistils was 2.1732 and 0 respectively. The relative content of hexane was the highest in petals, followed by diethyl phthalate, 2-ethyl-1-hexanol and 2-methyl-butane, that of the four compounds accounting for 65.30% and becoming the main components of the petals. The total content of hexane, 2,2,4-trimethyl-pentane, nonanal and 2-(4-Ethoxycarbonyl) benzylidene-benzofuran-6-ol-3-one accounted for 58.42% and become the main components of the stamens.

Table 2  The main volatile components and relative content of C. azalea in different flower parts

 The classification of the main volatile components in different flower parts was showed in Figure 2. The relative content of alkanes was the highest among the six type compounds, followed by aldehydes and ketones, alcohol and alkenes, that accounting for 48.21%, 17.93%, 13.12% and 12.22% of the total content respectively. The relative content of alkanes was the highest in petals, which accounting for 49.86%, and followed by alcohol, alkenes and esters. The relative content of alkanes was also the highest in stamens and sepals, that accounting for 51.59% and 44.66% respectively, and followed by aldehydes and ketones, alkenes and alcohol. The relative content of aldehydes and ketones was the highest in pistils, that accounting for 83.87%, and the other compounds content were all lower.

Fig. 1  Classification of main volatile components of flower of C. azalea in different florescence	Fig. 2  Classification of main volatile components of C. azalea in different flower parts

3 Discussion

The main compounds were alcohol, aldehyde, ester, alkyl, alkene and Linaloloxide in C.grijsii, C.japonica ‘Kramer’s supreme’ and C.japonica ‘Scentsation’. Cis-Linaloloxide II and Phenylethyl Alcohol were dominant in C. grijsii, and linalool was higher in C.japonica ‘Kramer’s supreme’(Fan et al, 2005;2006). The main volatile components and their relative content of C. azalea were quite different, which may be due to the difference of species or its specific components. The study on Chimonanthus praecox showed that the change of alkanes, alcohols and esters was positively correlated with the flower opening process (Xie et al, 2008). The relative content of alcohols of Jasminum sambac in immature, maturity and wilting stage was positively correlated with its storage time after theflower was harvested, and ester was the highest in maturity (Guo et al, 1994). The volatile components of C. azalea released gradually with the flower blooming, and the relative content of alcohols, esters and alkanes raised from the bud stage to the full flowering stage, that was basically same with the study on Jasminum sambac and Chimonanthus praecox. But the relative content of aldehydes and ketones of C. azalea decreased with the flower blooming, which should be further studied.

Petals and stamens of C. azalea were the main parts of volatile components releasing. There were some differences on volatile components in different flower parts, and the relative content of the same components was also different obviously in different parts, that suggested that there were differences on volatile components and their relative content in different flower parts(Yuan et al, 2008).The volatile components of C. azalea were studied by solid phase micro-extraction and GC/MS technology in the experiment, and the spatial-temporal variability of volatile components was make clear, which could provide a scientific basis for the further development and utilization. Because of the complexity of the formation and releasing mechanism of volatile components, the specific sources pathway of the main volatile compounds and the relationship between the changes of enzyme activities and the volatile compounds releasing of C. azalea need to be further studied.

Acknowledgement:

This work was supported by the National Key Twelfth-Five Science and Technology Program (2012BAD01B0703), National Natural Science Foundation (No.30800886), International Cooperation Project of China (2011DFA30490), and Zhejiang Key Flower Breeding Program (2012C12909-6).

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 [1] Author for correspondence: lixinlei2020@163.com, Associate Professor