**1. Introduction**

An important task in natural products chemistry is finding new compounds with novel properties and structures. Moreover, in fragrance chemistry, the odors of constituents are evaluated and key compounds are identified that contribute to the scent profiles of fragrance materials. Figure 1 shows the general investigative process for achieving these goals of fragrance chemistry.

Fig. 1. General investigative process for identifying new odor compounds with novel properties and structures in fragrance organic chemistry.

Extraction of odor plants gives oils containing many compounds. A popular and useful method for analyzing the composition is gas chromatography/mass spectrometry (GC-MS),

Separation of Odor Constituents by Microscale Fractional Bulb-To-Bulb Distillation 201

Commercially available, a bulb-to-bulb distillation apparatus (see Fig. 2) is usually used to purify a small amount of organic material (10-500 mg) by distillation under vacuum. Glass bulbs of about 3-3.5 cm in diameter are connected in series. The joint between bulbs, for example, between bulb A and bulb B, is made of ground glass. The general procedure for

3. Bulbs A, B, and C are placed into an oven and heated slowly. While the other bulbs are

5. Next, bulb C is removed from the oven and becomes a collection bulb. After cooling to room temperature, bulb C, along with bulb D, is placed in a cooling bath. Then, bulbs A

Cooling all the bulbs outside the oven is a critical part of this procedure. Failure to do so will

Using this method, we can perform separations on small amounts of material (down to 10

We used fractional bulb-to-bulb distillation to evaluate the odors of oils obtained from incense materials (sandalwood, frankincense, etc.). We extracted the essential oils of the incense first with hexane and then with methanol. We compared the odors of the hexaneextracted essential oils to the original base materials. We found that the odors of the extracted oils were similar to the odors of the base materials. We performed the following fractional distillations to obtain the minimum odorant groups constituting the fragrances of

**2.2 Separation of odor constituents of representative incense by bulb-to-bulb** 

being heated, bulb D is placed in a cooling bath (e.g., ice or water).

6. Finally, bulb A is heated and bulbs B, C, and D are cooled.

**2. Microscale fractional bulb-to-bulb distillation** 

performing a distillation on this apparatus is given below.

Fig. 2. Sketch of bulb-to-bulb distillation apparatus.

4. The first fraction is collected from bulb D.

result in low recovery and poor separation.

mg) to obtain fractions with different boiling points.

1. Oily sample is placed in bulb A. 2. The apparatus is evacuated.

and B are heated.

**distillation technique** 

**2.1 General procedure of microscale fractional bulb-to-bulb distillation** 

which provides important information about the scent profiles of fragrant plants. The analytical advances in the latest GC-MS technology are remarkable. In testing for the presence of a particular compound, detection of trace amounts of the substance has become possible. Recently, the olfactometry (O) functionality was incorporated into the GC-MS method. That analytical method, GC-MS-O, enables the odors of each constituent in a mixture to be precisely evaluated. These advances in analytical technology are associated with the discovery of natural products with important properties, for instance, a unique flavor. However, because the level of analysis is extremely detailed, the number of components in the characteristic scent of a plant can become huge. For this reason, indentifying the important scent components out of the many components is nontrivial. As a solution, a method for measuring the relative strength of each constituent's scent has recently been developed, and key findings have been reported. However, this method has the following problems.


Such difficulties arise in elucidating the key fragrance constituents of essential oils obtained from fragrance materials, such as sandalwood and vetiver, which are currently used as base notes.

We found that the fragrances of these scent materials can be expressed by combining groups of scent components. These groups consist of several organic compounds that have similar structures and thus similar odor. The interaction between these constituents is vital to the odor. From an oil, it is important to separate the *minimum odorant groups* necessary to retain the characteristic fragrance of the scent material.

In this chapter, I will explain how to divide the odor oils from fragrant plants into groups with similar properties (structure, odor, etc.). I will also present the results of experiments conducted in my laboratory, which used popular and important scent materials.

Sandalwood (*Santalum album,* L.), vetiver (*Vetiveria zizanioides*), patchouli (*Pogostemon patchouli*), and frankincense (*Boswellia papyrifera*) are typical materials in traditional Japanese incense, and the essential oils obtained from these materials possess unique and valuable features such as providing the base notes for perfume. Although many studies on the constituents of these materials have been reported, the key compounds of their scent profiles remain unknown. Determining these scent profiles is a prerequisite to elucidating the key components of the characteristic odors of these materials.

In attempting such a determination, however, the following main issues must be overcome. Firstly, the fragrances of the scent materials are not formed by a mere superposition of individual scents. Secondly, the scent components of these materials have weak odors. Almost all researchers have recognized these issues, but previously developed methods have not been suitable for clarifying the odor characteristics. In our work, we reconsidered and revised previous methods and were able to successfully evaluate the scent profiles of these materials
