Essential Oil from Korean Native Thyme Bak-Ri-Hyang

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This entry is adapted from the peer-reviewed paper 10.3390/molecules27134251

Thymus quinquecostatus Celak. (Korean name: bak-ri-hyang) is an important medicinal and aromatic herb in Korea, which is named for the spread of its fragrance over a distance of approximately 40 km. In traditional Korean systems of medicine, T. quinquecostatus has been used to treat cancer, constipation, hepatic disease, arteriosclerosis, poor circulation in women, constipation, and menstrual irregularities.

bak-ri-hyang chemical composition essential oil biological activity

1. Introduction

Essential oils are hydrophobic concentrates containing volatile aromatic compounds extracted from plants. Approximately 200 kinds of industrially used essential oils are obtained from the flowers, leaves, roots, stems, bark, fruits, buds, and seeds of various aromatic plants. Several methods are employed to isolate volatile components from medicinal and aromatic plants including hydro-distillation, steam distillation, extraction, supercritical fluid extraction (SFE), organic solvent extraction (OSE), and microwave extraction (AE). Among them, the hydro-distillation and steam distillation extraction methods are the standardized methods to isolate essential oils. Essential oils contain a variety of aromatic components, mainly belonging to monoterpenes and sesquiterpenes due to the differences in their odor characteristics. Essential oils are used in numerous industries to develop perfumery, aromatherapy, cosmetics, food, medicinal, and household cleaning products ^[1].

Essential oils from aromatic plants mainly contain monoterpenes and sesquiterpenes and their oxygenated derivatives with diverse chemical structures. These aroma components have been reported to possess various biological activities including anti-inflammatory, antispasmodic, analgesic, antioxidant, immunomodulatory, psychotropic, antiviral, antidiabetic, anticancer, and significant antibacterial properties ^[2]^[3]. Further, essential oils are applied in the fields of aromatherapy to maintain the physical and mental balance of humans. In agriculture, essential oils are used to inhibit the growth of plant fungal pathogens ^[4]^[5] and to control weeds ^[6]^[7]. Hence, essential oils can be used in eco-friendly agriculture in the future. In general, essential oils are widely used in the perfumery (fragrance) and the food industries (flavor) more than other industries. Although essential oils are produced from various aromatic plants in Korea, they are hardly used at the industrial level.

The genus Thymus belongs to the family Lamiaceae with approximately 300 species distributed worldwide ^[8]^[9]. Thymus species have traditionally been used to treat digestive and respiratory diseases such as colds, indigestion, nausea, and dysentery ^[10]. Thyme species possess various pharmacological properties including anti-inflammatory, analgesic, antispasmodic, antitussive, carminative, antihypertensive, antidiabetic, and anthelmintic ^[11]^[12]. This plant has also been used to enhance the flavor of tea or spice in daily life. In particular, the main components in the essential oils of Thymus, thymol and carvacrol, are classified as Generally Recognized As Safe (GRAS) by the United States Food and Drug Administration (FDA). However, in the United States, the daily allowance of essential oils used in food is stipulated ^[13]. In European Pharmacopoeia, only the thymol chemotype is selectively used among thyme essential oils. Thymol (36.0–55.0%), carvacrol (1.0–4.0%), p-cymene (15.0–28.0%), γ-terpinene (5.0–10.0%), linalool (4.0–6.5%), β-myrcene (1.0–3.0%), and terpinen-4-ol (0.2–2.5%) are the most abundant components in the essential oils of Thymus species. Among them, thymol is considered in pharmacopeias and used as a food preservative in the US, UK, Japan, India, and China ^[10].

2. Botanical Characteristics

Thyme is a perennial deciduous shrub known to be native to the Mediterranean coast of southern Europe ^[14]. It is also native to Korea ^[15]^[16], China ^[17], Japan, Mongolia, Central Asia ^[18], and other subtropical and temperate regions. In the Lamiaceae family, the genus Thymus has a large number of species and is considered important among botanists because of its high commercial and medicinal value. Thymus is a Nepetoideae Kostel (subfamily), Mentheae Dumort (tribe) Menthinae Endl. (subtribe) ^[19], with 250 taxa in Europe, Northwest Africa, Ethiopia, Asia, and Greenland ^[20], and it is generally classified into 215 species ^[21]. Different species of Thymus have significant morphological variations and do not have clear diagnostic characteristics for plant characteristics. Hence, the total number of Thymus species varies greatly among researchers, making it the most controversial plant among those of European origin ^[22].

T. quinquecostatus has many branches from the stem and the leaves are opposite and egg-shaped, oval, or lanceolate. The length of the leaf is 5~12 mm and the width is 3~8 mm; the edge has almost no saw teeth, there are small hairs, and it has concave dots on both sides of the leaf. Thyme leaves have capillary secretory hairs, and Shin and Yu ^[23] observed this microstructure with an electron microscope, inferring that the oleaginous substance would be secreted by granular secretion. Jing et al. ^[24] also observed trichomes according to the development of thyme leaves and found that there are two types of peltate and capitate trichomes in thyme, and the components accumulated at each developmental stage of the leaf are different. These findings indicate that the chemical composition of thyme’s essential oils significantly vary according to the growing season.

The flowers of thyme are white or pink, 7–9 mm long, and bloom in July–August. The fruits are small, dark brown, and bear fruit in September ^[25]. Nakada and Sugawara ^[26] investigated the floral variation and reproduction mode of thyme. As for the stamen, there are long L-stamens and short S-stamens. It has been reported that anthers of type S-stamen produce pollen of reproductive fertility, whereas anthers of type S-stamen produce pollen of complete infertility.

In particular, two varieties of T. quinquecostatus are found in Korea: bak-ri-hyang (T. quinquecostatus Celakovsky), found throughout the Korean Peninsula; island thyme (T. quinquecostatus var. japonica), found only in island areas including Ulleungdo. Thymol was found to be the most abundant component in the essential oils of bak-ri-hyang (39.8%) and island thyme (54.7%). ^[27]^[28]. T. vulgaris L. is reported to be a completely different species from Korean native thyme. T. quinquecostatus is also distributed in China, but significant variations were observed in their essential oil compositions. Carvacrol (28.54%) is the major component of thyme from the Ningxia Guyuan region, whereas p-cymene (17.39%) is from the Shandong Yimeng region. These data suggest that the main component in the essential oil of thyme remarkably varied according to the region. The difference in the essential oil composition might be due to the differences in environmental conditions, harvesting time, cross-pollination, extraction, and analysis methods of the essential oils ^[29]^[30]. There is a great morphological difference between thyme.

Currently, it has been investigated that several types of thyme introduced from foreign countries are being sold as domestic thyme in the private sector in Korea. Further, the correct identification of domestic thyme is too difficult due to the presence of many crossbreeds between native and imported thymes ^[31]. Therefore, a clear species characterization study of Korean native thyme is required. These studies will support the industrial needs following the entry into force of the Nagoya Protocol.

3. Agronomic Characteristics

Since thyme is an aromatic plant, the standard of cultivation in industrial terms is high yield and high quality of natural essential oil. The soil, light, planting distance, and cultivation methods for cultivating thyme containing high-content and high-quality essential oil have been studied. T. quinquecostatus can be propagated by seeds, but generally, cuttings are used. This takes advantage of the botanical characteristics of thyme stems that take root in the soil when they grow to a certain extent, but it is not possible to cut them at any time of the year. Choi et al. ^[32] reported that more than 80% of rooting was achieved when thyme was cultivated between April and June. After 30 days, the seedling qualities, such as rooting, plant length, leaf length, leaf width, and the number of branches, were investigated. The authors found that the appropriate cutting time for thyme propagation was between April and June. Farmers use plant growth regulators when making cuttings (personal communication with Kim Hee-yeon), but there has been no study on the effect obtained by using them. Choi et al. ^[32] reported that the rooting rate was the highest (93%) under a perlite:vermiculite ratio of 1:1 (v/v). This ratio is suitable for the propagation of thyme cuttings for improving the plant length and width of seedling quality. Sunlight, along with moisture, has a great influence on plant growth. When thyme was grown without shading, the fresh weight was high, and the flowering period was early. Conversely, due to the low root depth of thyme, if periodic irrigation is not performed, it will suffer from drying out. The planting distance affects the overall yield. In general, it is known that an increase in plant spacing is found to increase the yield of the plant. A similar pattern was observed in the case of baechohyang, which is grown as a medicinal plant on domestic farms. As the planting distance of thyme was farther, the fresh weight and dry weight of the plant increased. When the planting distance was 15 × 15 cm, the plant height of thyme was 27.8 cm, and the fresh weight was 48.7 g/plant. However, the planting distance was doubled, and when it was 30 × 30 cm, the plant height was 25.3 cm, and the fresh weight was 156.7 g/plant. From these results, it can be seen that the greater the distance of thyme planting, the higher the yield. The essential oil content was 3.4% in non-forestry cultivation, whereas the oil content was as low as 2.6% in open-field cultivation, which indicates that stable moisture supply and protection from damage caused by pests were effective ^[32].

4. Chemical Composition of Essential Oils

The essential oils from Thymus sp. Have long been used for medicinal purposes, perfumery, and in food industries due to the presence of various bioactive components and their unique aroma. Hence, thyme has been of interest to many academic and industrial researchers.

A total of 30 volatile compounds were detected in the steam distilled oil of thyme using the solid-phase microextraction (SPME) technique. The main components were trans-geraniol (36.85%), citral (15.64%), 3-octanone (3.70%), geranyl acetate (3.43%), borneol (2.48%), and nerol (2.25%) ^[33]. A study indicated that 29 different compounds were detected in the essential oil of thyme extracted by steam distillation. Trans-geraniol (39.75%) was the main components followed by citral (20.04%), geranyl acetate (6.00%), borneol (3.22%), nerol (3.21%), and 3-octanone (2.49%). In China, the main component of thyme from Cheongwon is trans-geraniol, but the main component of thyme from Shandong Yimeng is p-cymene. The study suggested that the chemotype is different from that of Cheongwon and Shandong Yimeng of China ^[29]. It is reported that the different chemical types of thyme grown in different regions are due to the differences in genotype and environmental conditions ^[8]^[31]. Choi et al. ^[34] reported that the yield of thyme essential oil was 2.62%, and this level of content is not low compared to essential oils of other herbal plants; therefore, it is useful in industrial extraction. It was reported that the essential oil yield of other herbal plants was gamguk: 2.0%; pear cloves: 0.6%; lavender: 3.49%; rosemary: 2.89%; chamomile: 1.16%.

Oh et al. ^[35] reported that cymene was the most abundant (93.5%) in the essential oil of Jeju thyme. Cymene was also found in the form of p-cymen-3-ol (50.41%), p-cymen-2-ol (24.06%), and cymene (19.04%). The results demonstrated that the chemical type of Jeju thyme is closer to thyme from Shandong Yimeng in China than thyme from Cheongwon Herbalang in Chungbuk ^[33]. From these results, it can be seen that thyme inhabiting Korea has various chemical types.

Lee et al. ^[36] extracted essential oil from the leaves and stems of thyme to test the functionality of native Korean herbal plants using three methods: supercritical extraction (SFE), steam distillation (SDE), and microwave extraction (MAE). When the leaves were extracted by the SFE, SDE, and MAE methods, the essential oil yields were 5.77, 5.72, and 5.70%, respectively, and there was no significant difference. There was no difference between the SFE method and the SDE method in the comparative study between the extraction methods performed on baechohyang in the same study by the authors. The MAE extraction method, used by Lee et al. ^[36] for the extraction of essential oils, uses microwaves and has the advantage of selectively heating and extracting only the desired components in natural substances ^[37]. It is also widely used in the extraction of essential oils ^[38]^[39].

In a comparative study on the content of island thyme essential oil, Lee et al. ^[37] reported that the efficiency of extraction with the SFE method was 0.20 to 0.27%, and the efficiency of the water steam distillation method was 0.03%. The authors found that this low extraction efficiency was different depending on the sampling location and harvest time, and this difference was also reported in the essential extraction efficiency of baechohyang ^[1].

Choi et al. ^[34] conducted a study on the harvest time, cultivation type, and drying method for the production of high-quality essential oil. In thyme, the essential oil content started to increase before the flowering period and was the highest during the flowering period (2.62%, steam distillation method), and after that, the content decreased significantly. Similar results were reported for baechohyang ^[40], indicating that the harvest time is very important for a high yield of essential oil. In a study on the volatile fragrance component of thyme according to the domestic production area, Chiang et al. ^[41] reported that there was a difference in the main component of essential oil. When thyme collected from the high mountains of Jeju Island was analyzed with a thermal desorption gas chromatograph-mass spectrometer, a total of 50 volatile components were detected in the essential oil, and the most abundant component was γ-terpinene (14.95%). A total of 62 volatile components were detected in thyme essential oil collected from the mid-mountainous region of Jeju, and the main component was p-thymol (27.51%). In the thyme collected from Ulleungdo, 41 types of volatile components were detected in the essential oil, and the main component was phenol (13.48%). A total of 54 components were detected in the essential oil of thyme collected in Gapyeong, Gyeonggi-do, and the main component was carvacrol (18.25%).

The results of these studies can be summarized as follows. First, the content of thyme essential oil differed by region. Second, it is possible to classify the chemical type as the main component of the essential oil of thyme. Chiang et al. ^[41] reported that there are at least four chemical types (i.e., γ-terpinene, p-thymol, phenol, and carvacrol) for thyme. The fact that the main component, as well as the content of the essential oil, may differ by region, even in the same species of plant, was reported in a study of thyme (T. vulgaris). The essential oil content of Thymus spathulafolia was 3.74% ^[42], Thymus capitatus was 6.0% ^[43], and Thymus linearis was 11.2% ^[44]. The main component of thyme from Casola and Modena, Italy, was thymol ^[45], but the main component of thyme from Palmaria Island was carvacrol ^[46].

In addition, Tohidi et al. ^[47] extracted essential oils of the genus Thymus collected from various regions in Iran. Through cluster analysis, they could be divided into three groups: thymol, geraniol/linalool, and carvacrol. These results indicate that even the same plant (thyme) living in one country may have different main components depending on the habitation area, and this difference is inferred due to the severe morphological variation of plants of the genus Thymus. In the western and central parts of the Balkan Peninsula, 250 taxa were identified ^[48] and only 110 taxa from the Mediterranean ^[49]. Hence, thyme has caused many taxonomic controversies among plant species of European origin ^[22]. Paying attention to the fact that bak-ri-hyang and island thyme are different in appearance, Shin and Kim ^[50] investigated the chemical composition of the essential oils. Thyme contains volatile organic compounds: thymol (41.7%) > γ-terpinene (16.0%) > p-cymene (13.0%) > β-caryophyllene (4.7%) > carvacrol (4.0%) > β-bisabolene (2.7%) > α-terpinene (2.6%). On the other hand, island thyme contains thymol (39.8%) > γ-terpinene (9.9%) > p-cymene (5.5%) > camphor (5.0%) > β-caryophyllene (4.7%) > α-pinene (3.5%). However, the most abundant component of both plants was thymol; thus, both plants can be classified as the same chemical type.

In a study by Shin and Choi ^[51], Korean thyme, which inhabits the entire Korean Peninsula, was classified into Jeju-do, Ulleung-do, and Korean-style. Baik et al. ^[52] reported that the content of the extracted components was different when domestic thyme was extracted with different solvents. The content of thymol was 64.23% in methanol extraction, 72.65% in ethanol extraction, and 57.98% in hexane extraction, indicating that the extraction solvent affects the extraction of specific components.

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