Note 55: Seasonal Variation in Flower Volatiles

By Santford V. Overton & John J. Manura
1999

INTRODUCTION

The identification and quantification of volatile organic compounds (VOC's) which are responsible for the flavor and fragrance qualities in many commercial products are of significant importance to the food/fragrance industry. The most important factors for the production of characteristic flavors and aromas are the plants. Floral fragrances are the primary means by which plants attract potential pollinators. In determining a plant's floral origin and assessing its overall flavor/aroma quality, it would be extremely advantageous to have a reliable range of marker compounds characteristic of the various flowers and plants. These organic compounds can be identified and used as marker compounds in commercial products. Previous methods for the extraction and analysis of these compounds used techniques such as: solvent extraction, headspace analysis and microdistillation followed by capillary chromatography. These methods either require large sample sizes, the use of solvents, or considerable time and effort to achieve the analysis. In addition, static headspace techniques are limited to their detection and identification of many organic volatiles and especially the semi-volatile organics. Other analytical techniques are needed to profile a wider range of volatile and semi-volatile organics and to identify the flavors, fragrances, off-flavors, off-odors, and potential contaminants that may be present as flavor and fragrance additives. For this study, volatiles organic compounds were purged from several varieties of flowers followed by trapping on an adsorbent resin using a dynamic purge and trap technique. The adsorbent traps were subsequently analyzed by thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS).

Instrumentation

Figure 1 - Purge and Trap System

Samples were collected using a Scientific Instrument Services Purge and Trap System. This apparatus (Fig. 1) consists of a sparge gas inlet connected to a stainless steel purging needle that is inserted through an adaptor fitting into a 10 ml test tube. A dry purge gas inlet is located at a right angle to the sparge gas inlet at the top of the apparatus. This can be left in the closed or open position. The purpose of the dry purge is to reduce the water vapor condensation on the adsorbent trap. Opposite the dry purge inlet is the connector for the glass-lined stainless steel (GLT) desorption tube containing the adsorbent resin. The purge and trap system also contains two ball rotameters with adjustable needle valves mounted on a stationary base and permits the visual indication and independent adjustment of the carrier gas flow to each of the gas inlets.

The experiments were conducted using a Scientific Instrument Services model TD-3 Short Path Thermal Desorption System accessory connected to the injection port of an HP 5890 Series II GC with electronic pressure control interfaced to an HP 5989A Mass Spectrometer. The mass spectrometers were operated in the electron impact mode (EI) and scanned from 35 to 550 daltons during the GC run for the total ion chromatogram.

The HP 5890 Series II GC contained a short 0.5 meter by 0.53 mm diameter fused silica precolumn attached to the injection port end of a 60 meter x 0.22 mm i.d. BPX35 capillary column containing a 0.25 um film thickness. The GC injection port was set to 250 degrees C and a 10:1 split was used. The head of each column was maintained at -70 degrees C using an S.I.S. Cryotrap model 951 during the desorption and extraction process and then ballistically heated to 200 degrees C after which the oven was temperature programmed from 35 degrees C (hold for 5 minutes) to 80 degrees C at 10 degrees C/min, then to 200 degrees C at 4 degrees C/min and finally to 260 degrees C at a rate of 10 degrees C/min.

Experimental

Sample sizes of 1 g of the flower varieties were inserted into a 10mL test tube at room temperature for 45 minutes. Samples were purged with high purity helium at 20 mL/min with an additional 25 mL/min dry purge using the S.I.S. purge and trap system. Volatile analytes were gas extracted and carried to a preconditioned 4.0 mm i.d. glass-lined stainless steel desorption tube packed with 200 mg of Tenax® TA. Once the samples were collected in the desorption tubes, they were purged with helium at 50 ml/min for 5 minutes to remove any moisture that may have been collected during the sampling period. The desorption tube with sample was then attached to the Short Path Thermal Desorption System. A syringe needle was also attached. The desorption tube was injected into the GC injection port at desorption block temperatures of 220 degrees C for 5 minutes and a flow rate of 10 mL/min. The GC and mass spectrometer were operated as previously described..

Results and Discussion

Six varieties of chrysanthemum (mums) and several different anatomical parts including the petals, sepals, stamens and stems of another variety of mum were analyzed to identify and compare the volatile organics of different flower varieties as well as the different parts of a flower (Fig. 1). Only the petals, the conspicuous colored part of the flower, were analyzed in the six mum varieties (Figs. 2-7). Over 100 volatile organics were identified in the mums studied. The mums studied produced 50 or more volatile organics which were identified in addition to many more that were either too weak or in which a good NBS library match was not achievable. The six varieties of mums possessed numerous mono- and sesquiterpenoid compounds as well as numerous straight and branched chain hydrocarbons, aldehydes, alcohols, ketones and esters (Figs. 2-7). Although they possess many common compounds, each mum variety had its own distinct fingerprint chromatograph. There exists significant variation in the quantities as well as the variety of volatile organics present in the mum varieties as well as between the different anatomical parts of the flower.

The aliphatic C6 compounds hexanal and (E)-2-hexenal and the alcohols (Z)-3-hexen-1-ol and (E)-2-hexen-1-ol (Figs. 2-6, 8), which contribute to the "green" notes of the aroma, were present in the majority of the petals of each mum. The formation of these compounds in the plant is related to cell destruction or to cellular breakdown due to maturation of the flower. The major ester found in the mums was bornyl acetate (Figs. 2, 4, 6-8). It is generally considered that esters primarily contribute to the fruity and floral notes. The aliphatic compound 1-octen-3-ol was also detected in several of the mums (Figs. 2, 6, 8) suggesting that the activity of lipoxygenase and hydroperoxide lyase producing C8 compounds from linoleic acid was occurring. High concentrations of the alcohol *-methyl-benzyl alcohol were present in each of the mums which may be characteristic of the flower during maturation. The predominant terpenes included the monoterpenes *-pinene, *-myrcene and eucalyptol with trace amounts of *-pinene, camphene and limonene (Figs. 2-8). Additional compounds included a series of sesquiterpenoid compounds, as caryophyllene and copaene. The Merck Index lists these compounds as constituents of plant derived essential oils, which are used as fragrance materials. The linear alkene 1-octene from lipid oxidation decomposition products was also identified in two of the mums (Figs. 2, 8). Additionally, numerous cyclic compounds which are characteristic of higher plants were found in each of the mums.

Although many of the same volatiles are present in the petals of these mums, there is a significant variation in the quantities as well as the variety of volatile organics in the different parts of the flower (Figs. 8-11). Numerous furan derivatives were identified in the green sepals which enclose the other flower parts in the buds (Fig. 9). Generally, volatile organic concentrations decreased in the stem portion of the flower except for the appearance of a high concentration of the cyclic sesquiterpenoid ylangene (Fig. 11). Linear alkenes, as 1-octene from lipid oxidation decomposition products, were only identified in the petals, sepals and stamens (Figs. 8-10). The higher concentration of volatile organics found in the petals, sepals and stamens of mums is indicative of the increased metabolism present in the developing flower.

Conclusion

The purge and trap or dynamic headspace technique followed by thermal desorption has been utilized for the identification and comparison of the volatile organic, aroma, and flavor components in mums. Many kinds of flavors/fragrances are used in commercial industry, and there is a demand for new and improve ones, especially natural ones. Such a source are oils which can produce a variety of flavors/fragrances. The short path thermal desorption used in conjunction with a dynamic headspace technique permits the identification and comparison of trace volatile organic compounds in flowers. The P&T technique offers several unique advantages over other techniques such as solvent extraction, static headspace sampling, and microdistillation including greater sensitivity and the detection of a wider range of volatile organics including higher molecular weight compounds and is more sensitive by a factor of at least 100 as compared to the static headspace technique. This technique can be easily incorporated into a troubleshooting technique to detect problems in a wide variety of commercial products, to compare variuos competing manufacturers products, as well as a quality control program.

Figure 2

2. 1-octene
3. 2-methyl-1-butanol
4. 1-pentanol
6. 7-methyl-1-octene
7. hexanal
9. 1-hexanol
11. (E)-2-hexen-1-ol
14. (E)-2-hexanal 15. *-pinene
18. 4-methylene-bicyclo [3.1.0] hexane
19. *-pinene
20. 1-octen-3-ol
21. 3-octanol
23. 3-octanone
26. limonene
31. eucalyptol
33. 1-methoxy-2-methyl-benzene
37. 4-methyl-1-(1-m)-3-cyclohexen-1-ol
39. octen-1-ol, acetate
41. 2-methyl-bicyclo [2.2.1] hept-2-ene
43. *-methyl-benzenemethanol
45. 6,6-dim-bicyclo [3.1.1] heptan-3-ol 49. camphor
58. bornyl acetate
63. 2-methyl-5-(1-)-2-cyclohexen-1-one
67. 6-isopropyli-bicyclo [3.1.0] hexane
73. caryophyllene
76. 3-methyl-6-(1-)-2-cyclohexen-1-one
77. 3,7-epoxy-2-methylene-6-methyloct
79. 2,6,6-tr-2,4-cycloheptadiene-1-one
86. decahydro-4a-methyl-1-naphthalene

Figure 3

Figure 4

3. 2-methyl-1-butanol
10. (Z)-3-hexen-1-ol
11. (E)-2-hexen-1-ol
13. (Z)-3-hexenal
15. *-pinene
17. camphene
18. 4-methylene--bicyclo [3.1.0] hexane
26. limonene
31. eucalyptol
39. octen-1-ol, acetate
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
49. camphor
56. 8-(1-methyle-)-bicyclo [5.1.0] octane
58. bornyl acetate
66. copaene
69. elemene
73. caryophyllene
76. 3-methyl-6-(1-)-2-cyclohexen-1-one
79. 2,6,6-tri-2,4-cycloheptadiene-1-one
85. 1,2,3,5,6,7,8,8a-octa-naphthalene
86. decahydro-4a-methyl-1-naphthalene
90. 7,11-dimethyl-1,6,10-dodecatriene

Figure 5

3. 2-methyl-1-butanol
10. (Z)-3-hexen-1-ol
11. (E)-2-hexen-1-ol
14. (E)-2-hexenal
15. *-pinene
18. 4-methylene-bicyclo [3.1.0.] hexane
29. 1-methyl-4-(1-methylethyl)-benzene
31. eucalyptol
38. 2-methyl-, 2-methyl-butanoic acid
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
49. camphor
56. 8-(1-methyle-)-bicyclo [5.1.0] octane
66. copaene
69. elemene
74. 4,11,11-bicyclo [7.2.0] undec-4-ene
76. 3-methyl-6-(1-)-2-cyclohexen-1-one
79. 2,6,6-tr-2,4-cycloheptadien-1-one
80. *-farnesene
84. 1,3-cyclohexadiene-1-carboxaldehyde
90. 7,11-dimethyl-1,6,10-dodecatriene

Figure 6

3. 2-methyl-1-butanol
11. (E)-2-hexen-1-ol
12. 2-methyl-bicyclo [3.1.0] hex-2-ene
14. (E)-2-hexenal
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene
20. 1-octen-3-ol 22. *-phellandrene
26. limonene
30. 1-methyl-2-(1-methylethyl)-benzene
31. eucalyptol
38. 2-methyl-, 2-methyl-butanoic acid
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
46. 2,5-dimethyl-3-meth-1,5-hexadiene
58. bornyl acetate
68. 3,5-dimethyl-2-cyclohexen-1-one
73. caryophyllene
79. 2,6,6-tr-2,4-cycloheptadien-1-one
81. 1-methyl-4-(5-methyl-)-cyclohexane
84. 1,3-cyclohexadiene-1-carboxaldehyde
89. 6,6-dimethyl-bicyclo [3.1.1] heptane

Figure 7

15. *-pinene
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene
31. eucalyptol
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
58. bornyl acetate
59. 6,6-di-bicyclo [3.3.1] heptan-3-one
60. 4,6-bicyclo [3.3.1] hept-3-en-2-one
61. 1,3,3-trimethyl-bicyclo [2.2.1] heptan-2-ol
73. caryophyllene
76. 3-methyl-6-(1-)-2-cyclohexen-1-one
79. 2,6,6-tr-2,4-cycloheptadien-1-one
84. 1,3-cyclohexadien-1-carboxaldehyde
90. 7,11-dimethyl-1,6,10-dodecatriene

Figure 9

1. acetic acid
2. 1-octene
3. 2-methyl-1-butanol
6. 7-methyl-1-octene
7. hexanal
9. 1-hexanol
11. (E)-2-hexen-1-ol
14. (E)-2-hexenal
15. *-pinene
16. 2-furancarboxaldehyde
17. camphene
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene 19. *-pinene
20. 1-octen-3-ol
21. 3-octanol 22. *-phellandrene
24. 1-methyl-4-(1-) 1,3-cyclohexadiene
27. 3,7-dimethyl-, 1,3,6-octatriene
31. eucalyptol
32. 5-methyl-2-furancarboxaldehyde
34. 1-methyl-4-(1-) 1,4-cyclohexadiene
35. 3-methyl-2,5-furandione
36. 1-methyl-4-(1-methyle-)-cyclohexane
38. 2-methyl-, 2-methyl-butanoic acid
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
46. 2,5-dimethyl-3-meth-1,5-hexadiene
49. camphor
51. 6,6-di-bicyclo [3.3.1] heptan-3-one
57. 4,6-bicyclo [3.3.1] hept-3-en-2-one
58. bornyl acetate
64. 3-methyl-6-(1-) 2-cyclohexen-1-one
66. copaene
75. 7,11-dimethyl-1,6,10-dodecatriene
76. 3-methyl-6-(1-) 2-cyclohexen-1-one
78. tricyclo [4.3.1.13,8] undecane
80. *-farnesene
82. 1H-cyclopenta [1,3] cyclopropa [1,2] b
92. linalyl-3-methylbutanoate
93. caryophyllene oxide

Figure 8

2. 1-octene
3. 2-methyl-1-butanol
5. toluene
6. 7-methyl-1-octene
7. hexanal
8. 4-methyl-1-pentanol
10. (Z)-3-hexen-1-ol
11. (E)-2-hexen-1-ol
14. (E)-2-hexenal
15. *-pinene
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene
20. 1-octen-3-ol
21. 3-octanol 23. 3-octanone
25. 6-methyl-5-hepten-2-one
28. 1,7,7-tricyclo [2.2.1.02,6] heptane
29. 1-methyl-4-(1-methylethen)-benzene
31. eucalyptol
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
49. camphor
53. 2-methyl-5-(1-)-2-cyclohexen-1-one
58. bornyl acetate
66. copaene
68. 3,5-dimethyl-2-cyclohexen-1-one
72. 4,6-bicyclo [3.1.1] hept-3-en-2-one
73. caryophyllene
76. 3-methyl-6-(1-)-2-cyclohexen-1-one
79. 2,6,6-tr-2,4-cycloheptadien-1-one
80. *-farnesene
82. 1H-cyclopenta [1,3] cyclopropa [1,2] b
88. 2,5-dimethyl-3-met-1,5-heptadiene
90. 7,11-dimethyl-1,6,10-dodecatriene
91. 3,7-dimethyl-2,6-octadien-1-ol

Figure 10

2. 1-octene
3. 2-methyl-1-butanol
5. toluene
6. 7-methyl-1-octene
7. hexenal
8. 4-methyl-1-pentanol
10. (Z)-3-hexen-1-ol
11. (E)-2-hexen-1-ol
12. 2-methyl-bicyclo [3.1.0] hex-2-ene
14. (E)-2-hexenal 15. *-pinene
17. camphene
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene
20. 1-octen-3-ol
21. 3-octanol
23. 3-octanone
25. 6-methyl-5-hepten-2-one
31. eucalyptol
38. 2-methyl-, 2-methyl-butanoic acid
39. octen-1-ol, acetate
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
49. camphor
53. 2-methyl-5-(1-)-2-cyclohexen-1-one
55. *-methyl-benzenepropanol
58. bornyl acetate
62. 4-methyl-cyclohexene
64. 3-methyl-6-(1-)-2-cyclohexen-1-one
70. 2,5-dimethyl-3-meth-1,5-hexadiene
73. caryophyllene
79. 2,6,6-tr-2,4-cycloheptadien-1-one
83. 8-(1-methyle)-bicyclo [5.1.0] octane
84. 1,3-cyclohexadiene-1-carboxaldehyde
93. caryophyllene oxide

Figure 11

3. 2-methyl-1-butanol
10. (Z)-3-hexen-1-ol
11. (E)-2-hexen-1-ol
14. (E)-2-hexenal
15. *-pinene
18. 4-methylene-bicyclo [3.1.0] hexane
94. *-myrcene
19. *-pinene
20. 1-octen-3-ol
21. 3-octanol 2
3. 3-octanone
24. 1-methyl-4-(1-)-1,3-cyclohexadiene
27. 3,7-dimethyl-,1,3,6-octatriene
31. eucalyptol
36. 1-methyl-4-(1-methyl-)-cyclohexene
42. bicyclo [2.2.2] oct-5-en-2-one
43. *-methyl-benzenemethanol
49. camphor
66. copaene
80. *-farnesene
82. 1H-cyclopenta [1,3] cyclopropa [1,2] b
65. ylangene
87. 1-ethenyl-1-methyl-2-cyclohexane

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