**1. Introduction**

There are currently 93 recognized species of *Curcuma* L. (Zingiberaceae) [1]. These perennial rhizomatous herbs originated in subtropical and tropical areas of Asia, Australia, and South America [2], and a number of *Curcuma* species are cultivated in large scales in India, Nepal, Pakistan, Bangladesh, Indonesia, Malaysia, and Thailand [3]. *Curcuma* species are herbaceous perennial herbs with tuberous rhizomes (underground stems). Among them, some important species such as *Curcuma amada* Roxb. (mango ginger), *Curcuma angustifolia* Roxb. (wild arrowroot), *C. aromatica* Salisb. (wild turmeric), *Curcuma caesia* Roxb. (black turmeric), *Curcuma decipiens* Dalzell, *Curcuma kwangsiensis* S. G. Lee & C. F. Liang, *Curcuma longa* L., *Curcuma montana* Roxb., *Curcuma ochrorhiza* Valeton, *Curcuma pierreana* Gagnep., *Curcuma roscoeana* Wall., and *Curcuma zedoaria* (Christm.) Roscoe (zedoary) have economical value as they are used in medicine, cosmetics, and both the floricultural and culinary industries [4]. Turmeric is mainly used for culinary, medicinal, and aromatic purposes. Its

**Citation:** Duong, L.; Mentreddy, S.R.; Satyal, R.; Satyal, P.; Setzer, W.N. Essential Oil Chemotypes of Four Vietnamese *Curcuma* Species Cultivated in North Alabama. *Horticulturae* **2022**, *8*, 360. https:// doi.org/10.3390/horticulturae8050360

Academic Editors: Dasha Mihaylova and Aneta Popova

Received: 25 March 2022 Accepted: 19 April 2022 Published: 21 April 2022

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**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

rhizomes are the ancient colorful spice source and have a bitter and pungen<sup>t</sup> taste and a pepper-like aroma. Turmeric is also known as the "Golden Spice of India" [5] or "Kitchen Queen" [6]. For example, it has been used in curries in India; in Japan and Korea it is popularly served as a herbal tea; and it is used as a preservative and a coloring agen<sup>t</sup> in mustard sauce, cheese, butter, and chips in the western world [7]. Curcuminoids and the essential oil of turmeric are associated with a myriad of medicinal, culinary and industrial properties of curcuma species [8], which are derived from the underground plant part, rhizomes (actually the stem), which are tuberous, with a rough and segmented skin. The primary rhizome is known as "mother rhizome" or bulb, and is pear-shaped in the center (Figure 1). The branches of mother rhizomes are the secondary rhizomes, called lateral or "finger rhizomes" [9].

**Figure 1.** Rhizomes of Vietnamese *Curcuma* species cultivated in North Alabama: *Curcuma aromatica* (green rhizome, CA22; white rhizome, CA46), *Curcuma caesia* (black rhizome, CC38), *Curcuma zanthorrhiza* (lime rhizome, CZ44), *Curcuma longa* (yellow rhizome, CL56; red rhizome, CL63). Photographs taken by Lam Duong.

Though turmeric has been known for its multiple uses for over 4000 years in India [10], its use as a medicinal and health supplement in the United States is of recent origin. The interest in turmeric in the U.S. has been increasing over the past two decades mainly due to a large number of scientific publications on its medicinal benefits [8]. To meet the growing demand for turmeric, the U.S. imports 90% of its market demand mainly from India. The U.S. import market was estimated at USD 87.28 million in 2018 [11]. The large market for turmeric in the United States suggests that there is opportunity for cultivation of turmeric in this country provided varieties with high curcumin yield and desirable essential oil composition are available.

*Curcuma aromatica* Salisb. (wild turmeric) is found naturally in South Asia, including southern China, Bhutan, Myanmar, India, Nepal, Sri Lanka [12], and Vietnam [13], and is widely cultivated in China, India, and Japan [14]. The plant is used in traditional medicines throughout its range for its wound-healing, anti-inflammatory, anti-tumor, immunomodulatory, antimicrobial effects and as an antidote for snake venom [15–17]. The rhizome essential oils are generally dominated by camphor, curzerenone, germacrone, curdione, and 1,8-cineole [14].

*Curcuma caesia* Roxb. (black turmeric) grows wild in northeastern and central India, Malaysia, Thailand, and Indonesia [14,18]. The rhizome of *C. caesia* has been used as a traditional medicine to treat leprosy, bronchitis, asthma, cancer, epilepsy, fever, wounds, impotence, fertility, vomiting, and pain [19]. *Curcuma caesia* is considered to be endangered in its native range in India [18], however, it has been underexplored in terms of cultivation and commercialization [20]. The major components in the rhizome essential oil of *C. caesia* from northeastern India were camphene, 1,8-cineole, camphor, borneol, (*E*)-β-caryophyllene, and *ar*-turmerone, which defined two chemotypes, a camphor/*ar*-turmerone chemotype and a 1,8-cineole/(*E*)-β-caryophyllene chemotype [20].

*Curcuma longa* L. (turmeric) is cultivated worldwide, especially in tropical countries in Asia, Australia, and the Neotropics [9]. It is a well-known medicinal agen<sup>t</sup> and culinary ingredient. In addition to curcumin and other non-volatile curcuminoids, the essential oil of turmeric has been employed in the treatment of various maladies in humans and animals [21]. Turmeric essential oils are made up of hundreds of components and the major components, however, are α-turmerone, β-turmerone, *ar*-turmerone, β-sesquiphellandrene, α-zingiberene, germacrone, terpinolene, *ar*-curcumene, and α-phellandrene [3,22].

*Curcuma zanthorrhiza* Roxb. (Javanese turmeric) is often referred to in the literature as *Curcuma xanthorrhiza* Roxb., however, that name is not recognized by World Flora Online [23]. The plant is native to Indonesia, although is also cultivated in Malaysia, the Philippines, Thailand, Vietnam, and to a lesser extent in China, India, Japan, and South Korea [24]. Traditional medicinal uses of the plant include treatment for stomach illness, liver ailments, constipation, bloody diarrhea, dysentery, arthritis, rheumatism, fevers, hemorrhoids, vaginal discharge, and skin eruptions [24].

As part of our research program investigating potential cultivation of *Curcuma* in Alabama, *Curcuma aromatica* (both green- and white-colored rhizomes), *C. caesia* (blackcolored rhizome), *C. zanthorrhiza* (lime-green rhizome), and *C. longa* (both yellow-, and red-colored rhizomes), obtained from Vietnam, were cultivated in North Alabama (Figure 1). The rhizome essential oils were obtained by hydrodistillation and analyzed by gas chromatographic methods. Both the "mother" or main rhizomes as well as the "daughter" or finger rhizomes were obtained and analyzed. The six *Curcuma* varieties used in this study were selected out of 64 genotypes according to three criteria: high yield but low curcuminoid content (variety, CL56), high yield but no curcumin content (CA22, CA46, CC38, and CZ44), and high yield and high curcumin content (CL63) based on unpublished data by the authors, Lam Duong and S.R. Mentreddy at Alabama A&M University.

### **2. Materials and Methods**

## *2.1. Plant Material*

The six *Curcuma* varieties used in this study were collected by Lam Duong from various locations in Vietnam: CA22 (Quang Nam province), CA46 (Gia Lai province), CC38 (Nghe An province), CL56 (Bac Giang province), CL63 (Quang Tri province), and CZ44 (Gia Lai province). The *Curcuma* rhizomes varieties were initially planted in a glass greenhouse at Alabama A&M University (Normal, AL, USA) and subsequently cultivated at the Alabama A&M Winfred Thomas Agricultural Research Station (Hazel Green, AL, USA, 3489 N, 8656 W) as previously described for *C. longa* cultivation [9]. Each of the fresh *Curcuma* rhizomes were collected on 18 March 2021, and stored at −20 ◦C until processed. The mother and daughter rhizomes were, separately, chopped and hydrodistilled using a Likens–Nickerson apparatus for 4 h (see Table 1).


**Table 1.** *Curcuma* rhizome hydrodistillation yields.

### *2.2. Gas Chromatographic Analyses*

The *Curcuma* rhizome essential oils were analyzed by gas chromatography—mass spectrometry (GC-MS) as previously reported [3]: Shimadzu GCMS-QP2010 Ultra (Shimadzu Scientific Instruments, Columbia, MD, USA), electron impact (EI) mode (70 eV), scan range 40–400 m/z, scan rate 3.0 scans/s; ZB-5ms GC column (60 m length × 0.25 mm diameter × 0.25 μm film thickness) (Phenomenex, Torrance, CA, USA), He carrier gas, 208.3 kPa head pressure, flow rate 2.0 mL/min, injector temperature 260 ◦C, ion source temperature 260 ◦C, oven temperature program 50 ◦C to 260 ◦C at 2 ◦C/min then held at 260 ◦C for 5 min; 0.3 μL of 5% *w*/*v* solutions in CH2Cl2 were injected, split ratio 1:24. Essential oil components were identified by comparison of MS fragmentation and retention index (RI) with those provided in the databases [25–28].

Gas chromatography with flame ionization detection (GC-FID) was carried out as previously reported [29]: Shimadzu GC 2010 (Shimadzu Scientific Instruments, Columbia, MD, USA) equipped with flame ionization detector, ZB-5 capillary column (60 m × 0.25 mm i.d.; film thickness 0.25 μm) (Phenomenex, Torrance, CA, USA); oven temperature programmed as above for GC-MS; injector and detector temperatures 260 ◦C; He carrier gas, flow rate 1.0 mL/min; 0.3 μL of 5% *w*/*v* solution in CH2Cl2 were injected, ratio 1:31. The percent compositions of the essential oils were calculated from peak areas with quantification using the external standard method; calibration curves of representative compounds from each class were used for quantification.

Chiral GC-MS was carried out as previously reported [29]: Shimadzu GCMS-QP2010S instrument (Shimadzu Scientific Instruments, Columbia, MD, USA), Restek B-Dex 325 column (Restek Corporation, Bellefonte, PA, USA); oven temperature program 50 ◦C to 120 ◦C at 1.5 ◦C/min, then to 200 ◦C at 2.0 ◦C/min; 0.1 μL of 5% *w*/*v* solutions in CH2Cl2 were injected, with a split ratio of 1:25. The enantiomers were determined by comparison of retention times with authentic samples obtained from Sigma-Aldrich (Milwaukee, WI, USA) and the relative enantiomer percentages were calculated from peak integration.
