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Chemical Composition of Essential Oils and Supercritical Carbon Dioxide Extracts from Cambodian Spices

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26 October 2023

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27 October 2023

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Abstract
Cambodian spices Amomum kravanh, Citrus hystrix and Piper nigrum 'Kampot' have long history of seasoning of food products. In this study, essential oils (EOs) and supercritical CO2 extracts from all three species have been analyzed using GC-MS with two columns of different polarity, whereas EO and CO2 extract from P. nigrum fruits and CO2 extract from the peel of C. hystrix have been isolated and analyzed for the first time. The results showed that A. kravanh EO contained mostly eucalyptol (78.8/ 72.6 %), while CO2 extract was rich in oleic acid (29.26 %) and tricosane (14.74 %). C. hystrix EO was rich in β-pinene (29.95/29.45 %), followed by D-limonene (24.54/23.24 %) and sabinene (9.94/10.23 %). β-pinene (30.2/28.9 %), D-limonene (23.99/23.74 %) and sabinene (13.36/19.55 %) were also predominant in the CO2 extracts. β-Caryophyllene was the main constituent of both EO and CO2 extract from P. nigrum 'Kampot' with contents ranging from 37.84 to 55.84 %, followed by 3-carene (from 7.18 to 18.72 %). Such findings suggest that supercritical CO2 can potentially be used for extraction of all three Cambodian spices. Nevertheless, further research determining the most efficient extraction parameters will be needed before its application in the spice processing practice.
Keywords: 
Subject: Biology and Life Sciences  -   Plant Sciences

1. Introduction

The term spice refers to dried plants or their parts that are used to enhance food flavor, taste, and color [1,2]. Nowadays, more than 400 spices and condiments are used worldwide from which among 275 species have their origin in tropical Southeast Asia. Cardamom, cinnamon, clove, ginger, nutmeg, pepper, and turmeric are examples of commodities of global economic importance. In 2021, the global spice market accounted for 21.3 billion US dollars, and it is forecasted to reach 27.4 billion USD by the end of 2026 [3,4]. Economically, the most important spice in international trade is black pepper, known as “king,” of the spices. Pepper or peppercorn refers to dried fruits of Piper nigrum, a perennial vine native to Western Ghats in India, which belongs to Piperaceae family. Throughout history, black pepper was one of the most traded spices worldwide and was utilized even as a currency all around the commercial routes established between Europe and India [5]. At the present time, the black pepper market is estimated to have reached 4400 million USD in 2022 and is likely to increase to almost 8 million USD by 2032 [6]. Apart from their seasoning properties, spices are used as natural colorants in food industry due to the presence of pigments. Moreover, due to numerous proven beneficial effects attributed to active biochemicals present in spices, they are also utilized in aromatherapy, cosmetics, nutraceuticals, perfumes and pharmaceuticals. Spices can be added to foods in various forms, such as whole, ground or in form of highly concentrated extracts [7].
In most spices, essential oils (EOs) are the main constituents responsible for their taste and olfactory sensations. EOs are usually comprised of large number (up to 400) of individual constituents with one or two dominant compounds, mostly classified as terpenes and their oxygenated derivatives. Other chemicals present in EOs are benzene derivatives which are responsible for the aroma of spices. Significant representatives include phenols or phenolic ethers [8,9,10]. Furthermore, alkaloids also contribute to the olfactory sensations, especially to pungency, of some spices. [11,12]. The extraction of EOs can be carried out using wide range of techniques while distillation (steam, water or combined) remains the most common method applied in industrial scale. Extracts obtained by solvent extraction are called oleoresins and contain flavor constituents of spices and other compounds soluble in organic solvents [2,4,13]. The most important shortcomings of distillation are high consumption of plant material, loss of thermo-sensitive compounds and long extraction time. The main disadvantages of solvent extraction are environmental and safety hazards associated with accumulation of organic solvents, high energy costs and oxidation of aroma and coloring compounds from spices [14]. To overcome these drawbacks, various green extraction techniques have recently been developed, while supercritical fluid extraction (SFE) belongs to one of them. This method provides multiple advantages associated with utilization of supercritical fluids as solvents, which possess different physicochemical properties such as lower viscosity and higher diffusivity and therefore results in higher extraction rates and overall faster process. Moreover, their density, which is related to the solvent capacity, can be modified by adjusting the extraction pressure and temperature [14,15] Although several solvents can potentially be used during SFE, carbon dioxide (CO2) remains the most employed supercritical fluid. Its easily obtainable moderate critical pressure and temperature ensure the preservation of labile compounds in the extract. In addition, CO2 is ubiquitous in the environment, non-toxic to human health, non-flammable and widely available at low cost [16,17]. All above-mentioned characteristics make supercritical CO2 highly attractive to be used as a “green solvent” and have led to multiple practical applications in different industries. Hop extract, decaffeinated tea and coffee, nicotine-free tobacco, and specialty oils are examples of commercial products obtained by CO2 extraction at industrial level [18,19]. Extensive research has occurred in the field of possible alternative use of SFE in extraction of bioactive components from spices during last decades and have led to the availability of wide variety of products on the market with CO2 extract from cinnamon, ginger, black and white pepper as examples [17]. Regarding to differences in chemical composition of EOs and supercritical CO2 extracts from various spices and aromatic plants, a plethora of studies have been conducted. Most commonly, results indicated significant differences in quantities of individual compounds. EOs obtained by distillation contained higher amounts of low molecular weight components like monoterpenoids and phenylpropanoids, while CO2 extracts were richer in constituents of higher molecular weight like sesquiterpenoids and diterpenoids [20,21,22,23].
Cambodia as well as many Southeast Asian countries has cuisine that is generally considered as healthy and delicious due to the abundant use of fish and incorporation of many vegetables, fruits, herbs, and spices into every meal. Fresh spices and herbs are essential ingredients in Cambodian dishes and approximately 42 g of condiments and spices are consumed daily per person [24,25]. Kampot pepper is one of the most popular spices grown in Cambodia with international recognition. It is a cultivar of Piper nigrum L produced in the Kampot province with unique climatic and soil conditions, giving the pepper distinctive flavor and aroma from other kinds of peppercorns. This commodity has been exported to Europe since 1870 during the French Protectorate due to its exceptional organoleptic quality. Currently, four different types of Kampot pepper can be found on the market: green, black, red, and white pepper. Although all Kampot peppers have an excellent reputation regarding to their sensory properties, red peppercorns are especially rare because of the unique process of their production. Red berries are harvested in full maturity, then blanched, sun-dried and manually sorted [26]. Leaves and fruits of numerous Citrus species (Rutaceae) are widely used to flavor foods and beverages. Citrus hystrix DC, known as kaffir lime, can be mentioned as an example of regionally used condiment. Leaves and fruit juice from this citrus are used for various flavoring purposes in Khmer cuisine and EO from the fruit pericarp is utilized in cosmetics and beauty products. Previous research has shown that kaffir lime EO possess various biological activities, namely antimicrobial, antioxidant, repellent and antiviral. Numerous studies analyzing chemical composition of C. hystrix EO have been conducted and revealed substantial number of monoterpene hydrocarbons and their oxygenated derivatives, with main compounds α/β-pinene, citronellal, limonene and sabinene. [28,29,30]. Plants belonging to the Zingiberaceae family are also widely used for their unique aroma as spices in Southeast Asia. Amomum kravanh Pierre ex Gagnep. (Zingiberaceae) is cultivated in Cambodia and other Southeast Asian countries for its fruits and leaves that are used to flavor curries. Fruit EO and SFE extract contain eucalyptol and β-pinene as main components [31,32,33,34]. With exception of single report on chemical composition of C. hystrix leaf CO2 extract [35], there is no study dealing with supercritical extraction of fruits of these species of spices that are frequently used in Southeast Asian cuisine. Therefore, the main objective of this study was to determine the chemical composition of EOs and CO2 extracts obtained from fruits of three traditional Cambodian spices, namely A. kravanh, C. hystrix and P. nigrum 'Kampot'.

2. Results

In this investigation, three EOs and three CO2 extracts have been isolated from Cambodian spice species with respective yield values ranging from 3.01 to 5.22 % (EOs) and from 0.57 to 8.35 % (CO2 extracts). In EOs obtained from P. nigrum 'Kampot', C. hystrix and A. kravanh, a total of 35, 38 and 21 individual constituents have been identified using HP-5 column, representing 99.38, 98.68 and 99.88 % of their respective total contents. Using DB-HeavyWAX column, a total of 41, 50 and 24 compounds have been detected constituting 99.15, 98.06 and 99.26 % of the total EOs, respectively. In CO2 extracts, a total number of 32, 36 and 31 components have been determined amounting to 98.65, 99.36 and 92.69 % of the total extracts. When analyzed with DB-HeavyWAX column, 40, 54 and 40 compounds have been identified which accounted for 96.74, 98.06 and 95.57 % of their total respective contents. Sesquiterpenes, monoterpenes and their oxygenated derivatives were the most predominant chemical groups in almost all tested EOs and CO2 extracts with exception of A. kravanh extract, where higher fatty acids and long-chain alkanes have been identified as the most abundant chemicals.
In A. kravanh EO, oxygenated monoterpene eucalyptol has been determined as prevailing ingredient comprising 78.8/ 72.6 % of the total sample. Other compounds occurring in significant amounts were monoterpenes β-pinene (7.68/7.49 %), α-pinene (2.3/ 2.2 %) and oxygenated derivative α-terpineol (4.31/ 4.67 %). In HP-5 analyses, L-terpinene-4-ol amounted to 1.19 % of the sample, however, this constituent has not been detected by DB-HeavyWAX column, where monoterpene D-limonene was identified as the third most abundant component (5.12 %). On the contrary, chemical composition of CO2 extract differed substantially from the EO. In HP-5 investigation, long-chained alkane tricosane comprised 14.74 % of the total extract followed by monoterpenoid eugenol acetate accounting for 14.02 % of the sample. Oleic acid was the third most prevailing constituent comprising 12.21 % of the extract accompanied by oxygenated monoterpenoid eugenol (7.91 %) and long chained alkane pentacosane (5.19 %). In contrast with these findings, analyses with DB-HeavyWAX column differed considerably. Majority of the sample consisted of oleic and palmitic acids constituting 29.26 and 17.07 % of the total respective content followed by tricosane (5.26 %), eugenol acetate (5.24 %) and linoleic acid (5.17 %). Compared to the hydrodistillation, the yield of CO2 extract was much lower (0.6 %), and its physical properties were different as the extract had a waxy and semi-solid structure.
Investigation of C. hystrix EO revealed monoterpenes as the most prevalent class of chemical compounds. Monoterpenes β-pinene (29.95/29.45 %), D-limonene (24.54/23.24 %) and sabinene (9.94/10.23 %) accompanied by alcohols L-terpinene-4-ol (9.71/9.07 %) and α-terpineol (3.7/3.62 %) were the main constituents of EO. Similarly, β-pinene (30.2/28.9 %), D-limonene (23.99/23.74 %) and sabinene (13.36/19.55 %) followed by aldehyde citronellal (5.21/4.28 %) were predominant components of the CO2 extract. In HP-5 column analysis, furanocoumarin oxypeucedanin accounted for 2.96 % of the total extract, however, this compound was not detected by DB-HeavyWAX column.
In P. nigrum 'Kampot' EO, sesquiterpene β-caryophyllene has been identified as the dominant compound constituting 34.84/39.55 % of the total oil followed by monoterpenes 3-carene (18.72/18.48 %), D-limonene (11.18/10.93 %) and β-pinene (5.42/5.32 %) when detected by HP-5/ DB-HeavyWAX columns, respectively. Similarly, analysis of CO2 extract has revealed even higher content of β-caryophyllene (54.21/ 55.86 %) accompanied by 3-carene, D-limonene and β-selinene comprising 7.4/ 7.18 %, 6.26/ 6.03 %, and 5.24/ 4.76 % of the total extract. Complete chemical analyses of A. kravanh, C. hystrix and P. nigrum ‘Kampot’ EOs and CO2 extracts are provided in Table 1, Table 2 and Table 3. Chromatograms of EOs and CO2 extracts can be seen in Figure 1 and Figure 2.

3. Discussion

As a result of GC-MS analysis, eucalyptol has been detected as the dominant constituent of A. kravanh EO. Such finding is in accordance with previously published studies investigating the chemical composition of EO from this plant [31,32], or from other species of Amomum genus [39,40]. In correspondence with results of Zhang et al. [33], β-pinene and α-terpineol have been detected as abundant compounds in the analyzed sample of the EO. Contrastingly, Diao et al. [31] reported relatively lower amounts of α-pinene (5.71 %) and β-pinene (2.41 %), while terpinyl acetate (11.2 %) and dipentene (6.1 %) were abundant EO components. These slight differences can be attributed to different geographical origins of the samples. Correspondingly with study of Zhang et al. [33], D-limonene has been identified as third most prevalent compound of the EO when investigated by DB-HeavyWAX column. In the literature, only results for chemical analyses using the DB-HeavyWAX column are available, however, they differ from our results. According to Yothipitak et al. [34], eucalyptol (71. 45 %), β-pinene (8. 64 %) and limonene (4. 77 %) were three dominant constituents of extract obtained by SFE from A. kravanh. These variances can be caused by different extraction parameters (33 °C and 175 bars) used during the SFE process and by distinct geographical origin (Thailand) of the plant sample. Different main constituents in HP-5 column and DB-HeavyWAX column analyses could be caused by stronger detection sensitivity and ability of polar DB-HeavyWAX column to separate and quantify fatty acids and their methylesters from the rest of the sample compared to the non-polar HP-5. Furthermore, polar columns based on polyethylene glycol have more accurate results in identification of fatty acid saturation and therefore are commonly employed in analyses of complex fats and oils [41,42].
In C. hystrix EO, β-pinene has been determined as the most abundant constituent, which is in accordance with previously published analyses, where percentages of β-pinene ranged from 25.93 to 47.93 % [28,43,44,45]. Study carried out by Jantan et al. [28] and Tran et al. [44]also revealed limonene as the second most dominant compound, comprising almost 15 and 20 % of the sample, respectively. Sabinene has been detected as the third most abundant constituent and such finding is in agreement with research carried out by the above-mentioned study [44]. However, slight discrepancy can be observed in comparison to investigation conducted by Sato et al. [43], where this monoterpene accounted for more than 20 % of the sample and has been second most dominant compound of the total oil. Moreover, our sample was lower in citronellal in comparison to previously published data. Since the samples from previously published studies were collected in Malaysia [28], Vietnam [46] and Thailand [43], differences in chemical composition can be attributed their different geographical origin. Furthermore, in case of study conducted by Sato et al. [43], steam distillation has been used as extraction method. In addition, maturity of the fruit and processing of the sample before extraction are factors which can affect the chemical composition of the EO [29]. Although CO2 extraction was previously performed from the leaves of this species in investigation carried out by Norkaew et al. [35], to the best of our knowledge, this is the first report investigating the chemical composition of CO2 extract isolated from the peel of this species. Due to the existence of large oil sacs or oil glands in the Citrus spp. fruit rind, their EOs have traditionally been obtained by cold expression. Cold pressed EOs of citruses comprise of volatile fraction with mono and sesqui-terpenes and their oxygenated derivatives. However, non-volatile fraction represented by coumarins, psoralens and other oxygen heterocyclic compounds, is also present in cold-pressed oils [47,48]. Although investigation of C. hystrix cold-pressed EO is currently not available in the literature, several research teams compared cold-pressed and hydrodistilled EOs from more common Citrus species. The most common conclusion was higher recovery of terpene hydrocarbons in the cold-pressed oils, which are compounds responsible for the typical aroma of Citrus oils [46,49]. Therefore, comparison of cold pressed C. hystrix EO with other extraction methods is highly encouraged for future research related to chemical composition or bioactivity assessment.
The main difference between P. nigrum 'Kampot' EO and CO2 extract was presence of pellitorine, belonging to the piperamides, which amounted to more than 1 % of the total CO2 extract. Such nitrogen containing compound has also been detected in research conducted by Luca et al. [50] in much lower amounts (0.18 %), however, other piperamides like piperine, piperettine and guineesine have also been determined in their extracts. This slight dissimilarity can be attributed to the different P. nigrum cultivar assessed in our research and different extraction conditions of the SFE process, where higher pressure (up to 300 bars) has been used for selective recovery of piperazines. The predominant compound in EO has been determined as sesquiterpene β-caryophyllene, which is in consonance with numerous previously published studies [51,52,53] assessing the composition of P. nigrum EO. Such sesquiterpene has been present in amounts ranging from 29.9 to 62.3 % of the volatile oil. Other major constituents in our sample were 3-carene and D-limonene. This is corresponding well with research conducted by Li et al. [52], where 3-carene and D-limonene were present in maximal respective amounts of 26.84 and 25.83 % in the various samples of EOs obtained from black and white peppers of Chinese origin. However, slightly different components were discovered in investigation carried out by Andriana et al. [51], where β-thujene and β-selinene accounted for 20.58 and 5.59 % of the sample, respectively. Furthermore, Kapoor et al. and Bagheri et al. [53,54] reported limonene (13.2 %), β-pinene (7.9 %) and sabinene (5.9 %) as predominant compounds of the EO. These slight differences in the main components can be attributed to different cultivar of P. nigrum assessed in our study, together with different harvest and post-harvest handling of the peppercorns used to produce red pepper [27]. In CO2 extract, the amount of β-caryophyllene was even higher than in EO and such finding is in accordance with research executed by Bagheri et al. [54], where the recovery of this sesquiterpene was also higher than in the hydrodistillated EO. Moreover, higher recovery of sesquiterpenes and their oxygenated derivatives for CO2 extracts opposed to EOs has also previously been reported [50,55]. Following main components were monoterpenes 3-carene and D-limonene and such finding corresponds well with study conducted by Topal et al. [56]. Same compounds amounted to 10.32 and 5.4 % in P. nigrum CO2 extract, respectively. Slight discrepancy can be observed in research executed by Luca et al. [50], where sabinene was present in 8.61 % and limonene comprised 8.21 % of the total P. nigrum extract, which can be attributed again to the different cultivar researched in our study. In addition, to the best of our knowledge, the chemical composition of P. nigrum 'Kampot' EO and CO2 extract has been assessed for the first time in our report.

4. Materials and Methods

4.1. Plant material and sample preparation

Fruits of A. kravanh and C. hystrix were purchased in local markets (Orussey Market, Phnom Penh, KH and Stung Treng Market, KH) and P. nigrum 'Kampot' fruits (red peppercorns) were obtained in a pepper farm store (La Plantation, Kampot, KH). C. hystrix was peeled and pericarp was used for further analyses. Dried material was homogenized by Grindomix apparatus (GM 100 Retsch, Haan, DE). The residual moisture contents of samples were determined gravimetrically at 130 °C for 1 h by Scaltec SMO 01 analyzer (Scaltec Instruments, Gottingen, DE) in triplicate according to the Official Methods of Analysis of the Association of Official Agricultural Chemists and expressed as arithmetic averages (15.79 %, 22.51 % and 14.39 % for A. kravanh, C. hystrix and P. nigrum, respectively).

4.2. Hydrodistillation of EOs

EOs were extracted by hydrodistillation of 100 g of ground plant materials in one litter of distilled water for 3 h using Clevenger-type apparatus (Merci, Brno, CZ) according to the procedure described in the [37]. Since hydrodistillation belongs to the most utilized methods for commercial production of EOs from C. hystrix spp. [29] the properties of samples prepared in our investigation should be alike to those commercially available. All EOs have been stored in 2 ml sealed glass vials at 4 °C until further use.

4.3. Supercritical CO2 extracts preparation

Supercritical CO2 extraction was carried out using Spe-ed SFE helix system (Applied Separations, Allentown, PA). Initially, 10 g of ground material have been filled into the 100 ml stainless steel extraction vessel between a glass wool bilayer. Subsequently, the filled vessel was installed into the extraction module and the extraction process was carried out using following parameters: isocratic pressure 200 Ba, temperature 40 °C and flow rate 5 LPM. The extracts have been captured into 60 ml glass collection vials (Applied Separations, Allentown, PA) and stored in 2 ml sealed glass vials at 4 °C until further utilization.

4.4. Gas chromatography-mass spectrometry analysis (GC-MS)

For determination of chemical composition of EOs and supercritical CO2 extracts, GC-MS analysis has been performed using the dual-column/dual-detector gas chromatograph Agilent GC-7890B. System is equipped with auto sampler Agilent 7,693, two columns, a fused-silica HP-5MS column (30 m × 0.25 mm, film thickness 0.25 μm, Agilent 19091s-433) and a DB-HeavyWAX (30 m × 0.25 mm, film thickness 0.25 μm, Agilent 122–7132), and a flame ionization detector (FID) coupled with single quadrupole mass selective detector Agilent MSD-5977B (Agilent Technologies, Santa Clara, CA). Helium has been utilized as a carrier gas at a flow rate 1 ml/min and the injector temperature was set 250 °C for both columns. The oven temperature was raised for both columns after 3 min from 50 to 280 °C. Initially, the heating velocity was 3 °C/min until the system reached temperature 120 °C. Subsequently the velocity increased to 5 °C/min until temperature 250 °C and after 5 min holding time the heating speed reached 15 °C/min until the obtained temperature 280 °C. Heating was followed by 20 min isothermal period. Samples of EOs and supercritical CO2 extracts were diluted in n-hexane for GC–MS (Merck KGaA, Darmstadt, DE) at the concentration 20 μl/ml. 1 μl of the solution was injected in split mode in a split ratio 1:30. The mass detector was set to the following conditions: ionization energy 70 eV, ion source temperature 230 °C, scan time 1 s, mass range 40–600 m/z.

4.5. Identification of constituents, quantification, and statistical analysis

Identification of compounds was based on comparison of their retention indices (RI), retention time (RT) and mass spectra in the National Institute of Standards and Technology Library ver 2.0.f (NIST) as well as in the literature [38]. The certain identified compounds were confirmed by co-injection of authentic standards, namely camphene (97.5 %, CAS: 79-92-5), β-caryophyllene (80 %, CAS: 87-44-5), humulene (96 %, CAS: 6753-98-6), linalool (97 %, CAS: 78-70-6), α-phellandrene (95 %, CAS: 4221-98-1), α-pinene (99 %, CAS: 7785-70-8), β-pinene (99.0 %, CAS: 18172-67-3) and γ-terpinene (97 %, CAS: 99-85-4) (Sigma-Aldrich, Prague, CZ). The RI were calculated for constituents separated by HP-5 column using RT of n-alkanes series ranging from C8 to C40 (Sigma-Aldrich, Prague, CZ). For each analyzed EO and CO2 extract, the final number of individual constituents was computed as the sum of components simultaneously identified using both columns and the remaining compounds detected by individual columns only. Quantitative data are expressed as relative percentage content of constituents determined by FID. Chemical analysis of EO or CO2 extract was performed in triplicates and relative peak area percentages were expressed as mean average of these three independent measurements ± standard deviation.

5. Conclusions

In summary, this study reports determination of chemical composition of EOs and CO2 extracts from three Cambodian spices, namely A. kravanh, C. hystrix and P. nigrum 'Kampot', using GC-MS equipped with two columns of different polarity. Differences between chemical composition of EOs and CO2 extracts have been observed in all species, whereas the most significant difference was detected in A. kravanh fruits. Furthermore, the analyses of both columns also differed substantially in the CO2 extract of this species. In non-polar HP-5 column, long chained alkane tricosane was the main compound, while in DB-HeavyWAX analysis, oleic and palmitic acids were two main constituents of the extract. C. hystrix and P. nigrum 'Kampot' CO2 extracts were generally richer in sesquiterpenes and their oxygenated derivatives in comparison to EOs, where monoterpenes were more abundant. Furthermore, fatty acid derivatives and other higher molecular weight constituents were also more prevalent in CO2 extracts. In addition, to the best of our knowledge, EO and CO2 extract from P. nigrum 'Kampot' fruits and CO2 extract from the peel of C. hystrix have been isolated and analyzed for the first time in our study. Such finding suggests that supercritical CO2 can potentially be used for extraction of all three Cambodian spices. Nevertheless, further research determining the most efficient extraction parameters will be needed before its application in the spice processing practice.

Author Contributions

Coordination and performing of extraction procedures and GC-MS data analyzes and manuscript drafting, K.V, coordination and performing of GC-MS analyzes and participation in manuscript preparation, K.U, coordination of plant material obtaining and participation in manuscript preparation, S.N, conceptualization and coordination of the whole study and manuscript finalization, L.K, all authors have read and agreed to the final version of the manuscript.

Funding

This research has been financially supported by Czech University of Life Sciences Prague (project IGA 20233109).

Institutional Review Board Statement

Not applicable.

Data Availability Statement

All is contained within the article.

Conflicts of Interest

The authors declare no conflict of interest.

Sample Availability

Samples of the EOs and CO2 extracts tested are available from the authors.

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Figure 1. GC-MS chromatograms of EOs of a) A. kravanh, b) C. hystrix and c) P. nigrum ‘Kampot’ (analyzed with HP-5 column). Peak numbers and constituents’ names: 1. β-pinene, 2. eucalyptol, 3. α-terpineol, 4. D-limonene, 5. L-terpinene-4-ol, 6. 3-carene, 7-β-caryophyllene and 8. β-selinene.
Figure 1. GC-MS chromatograms of EOs of a) A. kravanh, b) C. hystrix and c) P. nigrum ‘Kampot’ (analyzed with HP-5 column). Peak numbers and constituents’ names: 1. β-pinene, 2. eucalyptol, 3. α-terpineol, 4. D-limonene, 5. L-terpinene-4-ol, 6. 3-carene, 7-β-caryophyllene and 8. β-selinene.
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Figure 2. GC-MS chromatograms of CO2 extracts of a) A. kravanh, b) C. hystrix (analyzed on DB-HeavyWax column) and c) P. nigrum ‘Kampot’ (analyzed with HP-5 column). Peak number and compound names: 1. eugenol acetate, 2. tricosane, 3. palmitic acidt, 4. oleic acid, 5. β-pinene, 6. sabinene, 7. D-limonene, 8. 3-carene, 9. β-caryophyllene and 10. Pellitorine.
Figure 2. GC-MS chromatograms of CO2 extracts of a) A. kravanh, b) C. hystrix (analyzed on DB-HeavyWax column) and c) P. nigrum ‘Kampot’ (analyzed with HP-5 column). Peak number and compound names: 1. eugenol acetate, 2. tricosane, 3. palmitic acidt, 4. oleic acid, 5. β-pinene, 6. sabinene, 7. D-limonene, 8. 3-carene, 9. β-caryophyllene and 10. Pellitorine.
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Table 1. Chemical composition of A. kravanh EO and CO2 extract.
Table 1. Chemical composition of A. kravanh EO and CO2 extract.
RIa Compoundb Cc Extraction type/Column type/Peak area [%] Column type/Identification methodd
Essential oil CO2 extract
Obs. Lit. HP-5 MS DB-Wax HP-5 MS DB-Wax HP-5 MS DB-Wax
923 931 α-Thujene MH 0.09 ± 0.01 - - - - - - - - - RI, GC-MS -
929 932 α-Pinene MH 2.3 ± 0.05 2.20 ± 0.03 - - - - - - RI, GC-MS, Std GC-MS, Std
- 945 α-Fenchene MH - - - - - - - - - 0.01 ± 0.01 - GC-MS
944 946 Camphene MH 0.07 ± 0.01 0.08 ± 0.00 - - - - - - RI, GC-MS, Std GC-MS, Std
970 969 Sabinene MH 0.2 ± 0.04 0.23 ± 0.00 - - - - - - RI, GC-MS GC-MS
973 974 β-Pinene MH 7.68 ± 0.08 7.49 ± 0.09 - - - - - - RI, GC-MS, Std GC-MS, Std
989 988 β-Myrcene MH 0.78 ± 0.03 0.86 ± 0.08 - - - - - - RI, GC-MS GC-MS
1003 1002 α-Phellandrene MH 0.08 ± 0.01 - - - - - - - - - RI, GC-MS, Std GC-MS, Std
1015 1009 4-Carene MH 0.24 ± 0.03 0.17 ± 0.12 - - - - - - RI, GC-MS GC-MS
- 1014 α-Terpinene MH - - - 0.22 ± 0.01 - - - - - - - GC-MS
1025 1022 o-Cymene MH 0.69 ± 0.04 0.87 ± 0.02 - - - - - - RI, GC-MS GC-MS
1031 1026 Eucalyptol MO 78.89 ± 0.42 72.60 ± 0.89 - - - 0.08 ± 0.01 RI, GC-MS GC-MS
- 1031 D-Limonene MH - - - 5.12 ± 0.09 - - - 0.01 ± 0.01 - GC-MS
1058 1054 γ-Terpinene MH 1.05 ± 0.06 1.06 ± 0.02 - - - - - - RI, GC-MS GC-MS
- 1083 Fenchone MO - - - 0.18 ± 0.02 - - - - - - - GC-MS
1087 1086 Isoterpinolene MO 0.43 ± 0.02 - - - - - - - - - GC-MS -
1105 1095 Linalool MO 0.45 ± 0.01 0.50 ± 0.01 - - - - - - RI, GC-MS, Std GC-MS, Std
1174 1162 δ-Terpineol MO 0.39 ± 0.05 0.43 ± 0.01 - - - - - - RI, GC-MS GC-MS
1182 1174 L-terpinen-4-ol MO 1.19 ± 0.12 1.32 ± 0.02 - - - - - - RI, GC-MS GC-MS
1196 1186 α-Terpineol MO 4.31 ± 0.17 4.67 ± 0.08 3.68 ± 0.16 1.77 ± 0.15 RI, GC-MS GC-MS
1350 1346 α-Terpinyl acetate MO - - - - - - 0.17 ± 0.01 - - - RI, GC-MS -
1368 1356 Eugenol MO - - - - - - 7.91 ± 0.18 5.06 ± 0.18 RI, GC-MS GC-MS
- 1416 α-Santalene SH - - - - - - - - 0.08 ± 0.01 - GC-MS
1421 1419 β-Caryophyllene SH - - - - - - 1.37 ± 0.25 0.66 ± 0.02 RI, GC-MS, Std GC-MS, Std
1457 1452 Humulene SH - - - - - - 0.41 ± 0.03 0.12 ± 0.10 RI, GC-MS, Std GC-MS, Std
1486 1465 (Z)-muurola-4(14),5-diene SH 0.16 ± 0.01 - - - - - - - - - RI, GC-MS -
- 1478 γ-Muurolene SH - - - 0.16 ± 0.02 - - - - - - - GC-MS
1484 1484 Germacrene D SH - - - - - - 1.38 ± 0.02 - - - RI, GC-MS -
1490 1489 β-Selinene SH 0.35 ± 0.03 0.25 ± 0.01 2.06 ± 0.04 0.88 ± 0.16 RI, GC-MS GC-MS
1497 1496 Valencene SH - - - - - - 0.69 ± 0.10 0.09 ± 0.01 RI, GC-MS GC-MS
1508 1505 β-Bisabolene SH 0.36 ± 0.03 0.23 ± 0.01 3.59 ± 0.06 1.12 ± 0.10 RI, GC-MS GC-MS
1518 1513 γ-Cadinene SH - - - - - - 1.34 ± 0.09 0.73 ± 0.11 RI, GC-MS GC-MS
- 1514 Cubebol SO - - - - - - - - - 0.39 ± 0.01 - GC-MS
1525 1521 β-Sesquiphellandrene SH 0.12 ± 0.02 0.15 ± 0.00 1.60 ± 0.62 0.65 ± 0.02 RI, GC-MS GC-MS
1531 1521 Eugenol acetate MO - - - - - - 14.02 ± 0.74 5.23 ± 0.11 RI, GC-MS GC-MS
1558 1542 (Z)-Sesquisabinene hydrate SO 0.12 ± 0.02 0.15 ± 0.00 0.51 ± 0.11 0.20 ± 0.01 RI, GC-MS GC-MS
1566 1561 (E)-Nerolidol SO - - - 0.12 ± 0.00 1.71 ± 0.03 0.61 ± 0.04 RI, GC-MS GC-MS
1595 1577 (E)-Sesquisabinene hydrate SO - - - - - - 1.11 ± 0.30 0.44 ± 0.01 RI, GC-MS GC-MS
- 1577 Spathulenol SO - - - - - - - - - 0.15 ± 0.00 - GC-MS
1591 1582 Caryophyllene oxide SO - - - - - - 0.39 ± 0.10 0.25 ± 0.02 RI, GC-MS GC-MS
1675 1674 β-Bisabolol SO 0.04 ± 0.04 0.12 ± 0.04 0.21 ± 0.00 0.07 ± 0.00 RI, GC-MS GC-MS
1691 1685 α-Bisabolol SO - - - - - - 0.21 ± 0.00 0.07 ± 0.00 RI, GC-MS GC-MS
1714 1715 β-Santalol SO - - - - - - 0.56 ± 0.04 0.90 ± 0.01 RI, GC-MS GC-MS
- 1959 Palmitic acid FAD - - - - - - - - - 17.07 ± 0.23 - GC-MS
2086 2100 Heneicosane AH - - - - - - 2.12 ± 0.03 0.76 ± 0.00 RI, GC-MS GC-MS
- 2113 Linoleic acid FAD - - - - - - - - 5.17 ± 0.36 - GC-MS
2166 2141 Oleic Acid FAD - - - - - - 12.21 ± 0.25 29.26 ± 0.42 RI, GC-MS GC-MS
- 2172 Stearic acid FAD - - - - - - - - - 2.16 ± 0.09 - GC-MS
2186 2200 Docosane AH - - - - - - 1.57 ± 0.09 0.51 ± 0.10 RI, GC-MS GC-MS
2286 2300 Tricosane AH - - - - - - 14.74 ± 0.60 5.24 ± 0.09 RI, GC-MS GC-MS
2383 2400 Tetracosane AH - - - - - - 1.94 ± 0.03 0.69 ± 0.05 RI, GC-MS GC-MS
2482 2500 Pentacosane AH - - - - - - 5.19 ± 0.39 1.88 ± 0.08 RI, GC-MS GC-MS
- 2700 Heptacosane AH - - - - - - - - - 0.46 ± 0.02 - GC-MS
2096 NA Nonadecan-2-one K - - - - - - 0.54 ± 0.02 - - - GC-MS -
2261 NA Tetradec-9-enal A - - - - - - 0.24 ± 0.01 - - - GC-MS -
2267 NA Palmitoleic acid FAD - - - - - - 0.80 ± 0.06 1.94 ± 0.03 GC-MS GC-MS
2464 NA Hexadec-7-enal A - - - - - - 1.50 ± 0.13 0.11 ± 0.02 GC-MS GC-MS
2691 NA Azelaic acid bis(2-ethylhexyl) ester E - - - - - - 4.99 ± 0.12 1.96 ± 0.20 GC-MS GC-MS
2810 NA β-Monoolein E - - - - - - 2.88 ± 0.23 - - - GC-MS -
- NA Tricosanol A - - - - - - - - - 0.38 ± 0.01 - GC-MS
- NA Cyclopentadecanone K - - - - - - - - - 0.46 ± 0.01 - GC-MS
- NA Pentacos-1-ene AH - - - - - - - - - 0.48 ± 0.04 - GC-MS
- NA Heptacos-1-ene AH - - - - - - - - - 1.31 ± 0.04 - GC-MS
- NA Hexadec-9-enoic acid FAD - - - - - - - - - 0.30 ± 0.04 - GC-MS
- NA Octacosanol A - - - - - - - - - 3.16 ± 0.06 - GC-MS
- NA Squalene TH - - - - - - - - - 1.17 ± 0.05 - GC-MS
- NA Glyceryl linolenate E - - - - - - - - - 1.43 ± 0.06 - GC-MS
- NA β-Sitosterol O - - - 0.17 ± 0.14 - - - - - - - GC-MS
Total identified [%] 99.88 ± 0.05 99.29 ± 0.16 91.64 ± 0.88 95.41 ± 0.46
Footnotes 1: a) RI = retention indices for HP-5 column; Obs = retention indices determined relative to a homologous series of n-alkanes (C8-C40) on a HP-5MS column, Lit = literature RI values (Adams, 2007), NA = RI values were not available in the literature. b) C = Class; A - Aldehydes, DH - Diteprene hydrocarbons, E - Esters, FAD - Fatty acid and fatty acid derivatives, MH - Monoterpene hydrocarbons, MO - Oxygenated monoterpenes, O - Others, SH - Sesquiterpene hydrocarbons, SO - Oxygenated sesquiterpenes, d)Identification method: GC-MS = Mass spectrum was identical to that of National Institute of Standards and Technology Library (ver. 2.0.f), RI = the retention index was matching literature database; Std = constituent identity confirmed by co-injection of authentic standards. e) Retention indices were not calculated for compounds calculated only by DB-HeavyWAX column.
Table 2. Chemical composition of C. hystrix EO and CO2 extract.
Table 2. Chemical composition of C. hystrix EO and CO2 extract.
RIa Compoundb Cc Extraction type/Column type/Peak area [%] Column type/Identification methodd
Essential oil CO2 extract
Obs. Lit. HP-5 MS DB-Wax HP-5 MS DB-Wax HP-5 MS DB-Wax
923 931 α-Thujene MH 0.31 ± 0.01 0.34 ± 0.02 0.32 ± 0.02 0.31 ± 0 RI, GC-MS GC-MS
930 937 α-Pinene MH 2.93 ± 0.16 2.96 ± 0.09 2.54 ± 0.25 2.52 ± 0.09 RI, GC-MS, Std GC-MS, Std
- 945 Fenchene MH - - - - - - - - - 0.02 ± 0.02 - GC-MS
945 946 Camphene MH 0.16 ± 0.01 0.19 ± 0.01 0.14 ± 0.01 0.15 ± 0 RI, GC-MS, Std GC-MS, Std
971 976 Sabinene MH 9.94 ± 0.22 10.2 ± 0.15 19.36 ± 0.97 19.55 ± 0.33 RI, GC-MS GC-MS
974 980 β-Pinene MH 29.95 ± 0.54 29.5 ± 0.45 30.2 ± 1.84 28.9 ± 0.55 RI, GC-MS, Std GC-MS, Std
990 991 β-Myrcene MH 1.20 ± 0.05 1.38 ± 0.02 1.43 ± 0.06 1.58 ± 0.02 RI, GC-MS GC-MS
- 1004 Pseudolimonene MH - - - 0.97 ± 0.07 - - - 0.97 ± 0.04 - GC-MS
1003 1005 α-Phellandrene MH 0.07 ± 0.01 0.06 ± 0 - - - - - - RI, GC-MS, Std GC-MS, Std
- 1008 3-Carene MH - - - 0.01 ± 0.02 - - - - - - - GC-MS
1015 1009 4-Carene MH - - - - - - 0.02 ± 0.02 - ± - RI, GC-MS -
1015 1009 α-Terpinene MH 0.51 ± 0.04 0.61 ± 0.01 - - - - - - RI, GC-MS GC-MS
1025 1022 o-Cymene MH 1.91 ± 0.01 2.34 ± 0.01 0.1 ± 0.01 0.21 ± 0.02 RI, GC-MS GC-MS
1028 1031 D-Limonene MH 24.54 ± 0.16 23.2 ± 0.1 23.99 ± 0.45 23.74 ± 0.12 RI, GC-MS GC-MS
1058 1062 γ-Terpinene MH 1.67 ± 0.01 1.63 ± 0.02 0.08 ± 0 0.1 ± 0.01 RI, GC-MS GC-MS
1072 1065 (Z)-Sabinene hydrate MO - - - - - - 0.78 ± 0.06 1.06 ± 0.02 RI, GC-MS GC-MS
- 1071 β-Terpinene MH - - - - - - - - - 0.03 ± 0.01 - GC-MS
1074 1074 Linalool oxide MO 1.55 ± 0.02 1.57 ± 0.03 - - - 0.05 ± 0.01 RI, GC-MS GC-MS
1087 1086 Terpinolene MH 0.54 ± 0.03 0.52 ± 0.01 0.02 ± 0.01 0.05 ± 0 RI, GC-MS GC-MS
1105 1095 Linalool MO 0.72 ± 0.09 0.98 ± 0.02 0.52 ± 0.27 0.81 ± 0.07 RI, GC-MS, Std GC-MS, Std
- 1098 (E)-Sabinene hydrate MO - - - - - - - - - 0.35 ± 0.03 - GC-MS
- 1114 Fenchol MO - - - 0.02 ± 0.03 - - - - - - - GC-MS
1145 1137 Sabinol MO - - - - - - 0.08 ± 0.02 - ± - RI, GC-MS -
1150 1145 L-isopulegol MO 0.22 ± 0.04 0.28 ± 0.03 0.07 ± 0.01 0.11 ± 0 RI, GC-MS GC-MS
1154 1148 Citronellal MO 1.38 ± 0.07 1.05 ± 0.04 5.21 ± 0.31 4.28 ± 0.25 RI, GC-MS GC-MS
1174 1165 Borneol MO 0.09 ± 0.02 - - - - - - - - - RI, GC-MS -
1161 1167 dl-Isopulegol MO 0.12 ± 0.02 - - - - - - - - - RI, GC-MS -
1184 1174 L-terpinen-4-ol MO 9.71 ± 0.16 9.07 ± 0.22 0.36 ± 0.04 0.42 ± 0.02 RI, GC-MS GC-MS
- 1176 m-Cymen-8-ol MO - - - 0.09 ± 0 - - - - - - - GC-MS
- 1182 Pinocarveol MO - - - 0.03 ± 0.03 - - - 0.06 ± 0 - GC-MS
1199 1189 α-Terpineol MO 3.70 ± 0.09 3.62 ± 0.07 0.98 ± 0.17 1 ± 0.04 RI, GC-MS GC-MS
- 1194 Myrtenol MO - - - 0.05 ± 0 - - - - - - - GC-MS
1217 1205 (E)-Piperitol MO 0.02 ± 0.04 - - - - - - - - - RI, GC-MS -
1237 1228 Citronellol MO 0.75 ± 0.15 0.94 ± 0.03 0.43 ± 0.28 0.92 ± 0.04 RI, GC-MS GC-MS
- 1249 Geraniol MO - - - 0.1 ± 0 - - - - - - - GC-MS
1291 1273 (Z)-Ascaridole glycol O 0.40 ± 0.10 0.69 ± 0.02 - - - - - - RI, GC-MS GC-MS
- 1312 Citronellic acid MO - - - 0.3 ± 0.01 - - - 0.25 ± 0.01 - GC-MS
1351 1345 α-Cubebene SH - - - - - - 0.05 ± 0.02 0.07 ± 0 RI, GC-MS GC-MS
1355 1354 Citronellyl acetate MO 0.27 ± 0.03 0.32 ± 0.01 0.33 ± 0.11 0.44 ± 0.03 RI, GC-MS GC-MS
1384 1365 Neryl acetate MO 0.24 ± 0.04 - - - 0.22 ± 0.14 0.01 ± 0.02 RI, GC-MS GC-MS
1379 1374 α-Copaene SH 0.95 ± 0.03 0.75 ± 0.04 1.43 ± 0.13 1.21 ± 0.12 RI, GC-MS GC-MS
- 1379 Geranyl acetate MO - - - 0.36 ± 0.02 - - - 0.51 ± 0.03 - GC-MS
1391 1390 β-Cubebene SH 0.37 ± 0.01 - - - 1.04 ± 0.15 - ± - RI, GC-MS -
1395 1391 β-Elemene SH 0.05 ± 0.00 - - - - - - - - - RI, GC-MS -
1424 1419 β-Caryophyllene SH 0.72 ± 0.02 0.57 ± 0.01 1.12 ± 0.09 1.05 ± 0.09 RI, GC-MS, Std GC-MS, Std
- 1430 β-Copaene SH - - - 0.31 ± 0.01 - - - 0.96 ± 0.09 - GC-MS
1461 1454 Humulene SH 0.23 ± 0.01 0.2 ± 0.01 0.35 ± 0.03 0.32 ± 0.02 RI, GC-MS, Std GC-MS, Std
1488 1484 D-Germacrene SH 0.24 ± 0.01 0.07 ± 0 0.69 ± 0.05 0.49 ± 0.06 RI, GC-MS GC-MS
1502 1495 Bicyclogermacrene SH - - - - - - 0.14 ± 0.07 0.13 ± 0.01 RI, GC-MS GC-MS
1505 1499 α-Muurolene SH 0.10 ± 0.01 0.05 ± 0.02 0.1 ± 0.05 0.09 ± 0.01 RI, GC-MS GC-MS
- 1514 Cubebol SO - - - - - - - - - 0.24 ± 0.02 - GC-MS
1528 1524 β-Cadinene SH 1.49 ± 0.04 1.06 ± 0.1 1.75 ± 0.11 1.57 ± 0.07 RI, GC-MS GC-MS
- 1528 Calamenene SH - - - 0.01 ± 0.01 - - - - - - - GC-MS
- 1548 Elemol SO - - - 0.09 ± 0.02 - - - 0.01 ± 0.01 - GC-MS
1588 1574 Germacrene D-4-ol SO - - - - - - 0.07 ± 0.01 0.17 ± 0.01 RI, GC-MS GC-MS
- 1577 Spathulenol SO - - - 0.09 ± 0 - - - 0.09 ± 0.01 - GC-MS
- 1582 Caryophyllene oxide SO - - - - - - - - - 0.05 ± 0 - GC-MS
- 1608 Humulene epoxide SO - - - 0.01 ± 0.01 - - - - - - - GC-MS
- 1619 Humulane-16-dien-3-ol SO - - - - - - - - - 0.04 ± 0 - GC-MS
1641 1627 Epicubenol SO 0.08 ± 0.01 0.08 ± 0.02 - - - - - - RI, GC-MS GC-MS
1647 1631 γ-Eudesmol SO 0.26 ± 0.01 0.41 ± 0.03 - - - - - - RI, GC-MS GC-MS
- 1645 Cubenol SO - - - 0.07 ± 0.02 - - - - - - - GC-MS
1656 1645 δ-Cadinol SO 0.10 ± 0.00 0 ± 0.01 - - - 0.02 ± 0.02 RI, GC-MS GC-MS
- 1649 β-Selinenol SO - - - 0.18 ± 0.02 - - - 0.04 ± 0 - GC-MS
1671 1652 α-Eudesmol SO 0.41 ± 0.01 0.06 ± 0.01 - - - - - - RI, GC-MS GC-MS
- 1656 Patchouli alcohol SO - - - - - - - - - 0.04 ± 0.01 - GC-MS
- 1942 Phytol DO - - - - - - - - - 0.06 ± 0 - GC-MS
- 1984 Palmitic acid FAD - - - - - - - - - 0.62 ± 0.01 - GC-MS
- 2132 Linoleic acid FAD - - - - - - - - - 0.28 ± 0.03 - GC-MS
2521 2501 Oxypeucedanin O - - - - - - 2.96 ± 0.64 - ± - RI, GC-MS -
2707 2707 β-Monolinolein E - - - - - - 0.18 ± 0.01 - ± - RI, GC-MS -
1562 NA Hedycaryol SO - - - - - - 0.17 ± 0.01 0.17 ± 0.02 GC-MS GC-MS
2009 NA Thunbergol DO - - - - - - 1.79 ± 0.12 1.4 ± 0 GC-MS GC-MS
2010 NA trans-Geranylgeraniol DO 0.79 ± 0.02 0.64 ± 0.01 0.31 ± 0.03 0.19 ± 0 GC-MS GC-MS
- NA 2-p-Menthen-1-ol MO - - - 0.15 ± 0.01 - - - - - - - GC-MS
- NA tau.-Muurolol SO - - - 0.05 ± 0.01 - - - - - - - GC-MS
- NA Tetradecanoic acid FAD - - - - - - - - - 0.16 ± 0.01 - GC-MS
- NA 17-Octadecynoic acid FAD - - - - - - - - - 0.12 ± 0.08 - GC-MS
- NA Ricinoleic acid FAD - - - - - - - - - 0.06 ± 0.01 - GC-MS
Total identified [%] 98.69 ± 0.22 98 ± 0.39 99.36 ± 0.07 98.06 ± 0.32
Footnotes 2: a) RI = retention indices for HP-5 column; Obs = retention indices determined relative to a homologous series of n-alkanes (C8-C40) on a HP-5MS column, Lit = literature RI values (Adams, 2007), NA = RI values were not available in the literature. b) C = Class; A - Aldehydes, DO -Oxygenated diterpenes, E - Esters, FAD - Fatty acid and fatty acid derivatives, MH - Monoterpene hydrocarbons, MO - Oxygenated monoterpenes, O - Others, SH - Sesquiterpene hydrocarbons, SO - Oxygenated sesquiterpenes, d)Identification method: GC-MS = Mass spectrum was identical to that of National Institute of Standards and Technology Library (ver. 2.0.f), RI = the retention index was matching literature database; Std = constituent identity confirmed by co-injection of authentic standards. e) Retention indices were not calculated for compounds calculated only by DB-HeavyWAX column.
Table 3. Chemical composition of P. nigrum 'Kampot' EO and CO2 extract.
Table 3. Chemical composition of P. nigrum 'Kampot' EO and CO2 extract.
RIa Compoundb Cc Extraction type/Column type/Peak area [%] Identificationd
Essential oil CO2 extract
Obs. Lit. HP-5 MS DB-Wax HP-5 MS DB-Wax HP-5 MS DB-Wax
923 924 α-Thujene MH 0.059 ± 0 - - - - - - - - - RI, GC-MS -
929 937 α-Pinene MH 2.806 ± 0.25 2.568 ± 0.09 0.649 ± 0.02 0.574 ± 0.04 RI, GC-MS, Std GC-MS, Std
944 946 Camphene MH 0.04 ± 0.01 0.05 ± 0 - - - - - - RI, GC-MS, Std GC-MS, Std
970 976 Sabinene MH 0.048 ± 0.03 0.091 ± 0 - - - - - - RI, GC-MS GC-MS
973 980 β-Pinene MH 5.424 ± 0.45 5.322 ± 0.14 2.039 ± 0.04 1.996 ± 0.14 RI, GC-MS, Std GC-MS, Std
989 991 β-Myrcene MH 1.477 ± 0.14 1.682 ± 0.12 0.681 ± 0.05 - - - RI, GC-MS GC-MS
- 1001 2-Carene MH - - - - - - - - - 0.076 ± 0.02 - GC-MS
1003 1005 α-Phellandrene MH 1.803 ± 0.14 1.481 ± 0.08 0.762 ± 0.03 0.681 ± 0.03 RI, GC-MS, Std GC-MS, Std
1008 1008 3-Carene MH 18.72 ± 1.46 18.49 ± 0.42 7.395 ± 0.17 7.181 ± 0.4 RI, GC-MS GC-MS
1025 1022 o-Cymene MH 1.399 ± 0.12 1.495 ± 0.04 0.636 ± 0.01 0.771 ± 0.05 RI, GC-MS GC-MS
1028 1031 D-Limonene MH 11.18 ± 0.79 10.93 ± 0.15 6.265 ± 0.11 6.034 ± 0.39 RI, GC-MS GC-MS
1058 1062 γ-Terpinene MH 0.056 ± 0.01 0.045 ± 0 - - - - - - RI, GC-MS GC-MS
1084 1086 Isoterpinolene MH 0.194 ± 0.04 0.4 ± 0.02 0.09 ± 0 - - - RI, GC-MS GC-MS
1087 1086 Terpinolene MH 0.428 ± 0.08 0.169 ± 0 0.156 ± 0.01 0.191 ± 0.01 RI, GC-MS GC-MS
1104 1095 Linalool MO 0.354 ± 0.03 0.453 ± 0 0.238 ± 0.05 0.386 ± 0.01 RI, GC-MS, Std GC-MS, Std
- 1140 Verbenol MO - - - 0.185 ± 0.02 - - - - - - - GC-MS
- 1179 p-Cymen-8-ol MO - - - 0.05 ± 0.05 - - - - - - - GC-MS
- 1318 2,3-Pinanediol MO - - - 0.254 ± 0.01 - - - - - - - GC-MS
- 1329 Piperonal O - - - - - - - - - 0.04 ± 0.01 - GC-MS
1339 1339 δ-EIemene SH 0.559 ± 0.02 0.588 ± 0.01 0.491 ± 0.01 0.516 ± 0.01 RI, GC-MS GC-MS
- 1340 Piperitenone MO - - - 0.063 ± 0.05 - - - - - - - GC-MS
1351 1351 α-Cubebene SH 0.097 ± 0 0.086 ± 0.02 0.144 ± 0.01 0.117 ± 0.01 RI, GC-MS GC-MS
- 1357 Octadecanal A - - - 0.57 ± 0.1 - - - - - - - GC-MS
1378 1374 α-Copaene SH 0.194 ± 0.01 0.17 ± 0.01 0.275 ± 0.01 0.247 ± 0.02 RI, GC-MS GC-MS
1394 1391 β-Elemene SH 1.483 ± 0.08 - - - 1.887 ± 0.03 1.303 ± 0.03 RI, GC-MS -
1410 1409 α-Gurjunene SH 0.164 ± 0.01 0.131 ± 0 0.257 ± 0.01 0.239 ± 0.03 RI, GC-MS GC-MS
1416 1411 α-Bergamotene SH 0.093 ± 0 0.01 ± 0.01 0.154 ± 0.01 0.018 ± 0 RI, GC-MS GC-MS
1425 1419 β-Caryophyllene SH 37.84 ± 2.05 39.55 ± 1.12 54.21 ± 0.85 55.86 ± 1.37 RI, GC-MS, Std GC-MS, Std
- 1434 γ-Elemene SH - - - 0.057 ± 0 - - - 0.12 ± 0.01 - GC-MS,
1440 1437 α-Guaiene SH 0.983 ± 0.07 - - - 1.363 ± 0.02 - - - RI, GC-MS -
1456 1454 β-Farnesene SH 0.101 ± 0.03 0.058 ± 0.05 0.143 ± 0 0.167 ± 0 RI, GC-MS GC-MS
1459 1454 Humulene SH 2.572 ± 0.22 2.52 ± 0.07 3.7 ± 0.02 3.465 ± 0.08 RI, GC-MS, Std GC-MS, Std
- 1475 γ-Gurjunene SH - - - 0.9 ± 0.03 - - - - - - - GC-MS
1493 1485 β-Selinene SH 3.653 ± 0.33 3.358 ± 0.15 5.242 ± 0.14 4.757 ± 0.11 RI, GC-MS GC-MS
1486 1492 Valencene SH 0.136 ± 0.01 - - - 0.224 ± 0.01 - - - RI, GC-MS -
1501 1494 α-Selinene SH 2.409 ± 0.23 1.972 ± 0.29 3.493 ± 0.08 3.009 ± 0.09 RI, GC-MS GC-MS
1510 1506 β-Bisabolene SH 1.131 ± 0.1 0.887 ± 0.1 1.711 ± 0.06 1.284 ± 0.05 RI, GC-MS GC-MS
1524 1520 7-epi-α-Selinene SH 0.114 ± 0.01 - - - 0.168 ± 0.03 - - - RI, GC-MS -
- 1528 Calamenene SH - - - - - - - - - 0.013 ± 0 - GC-MS
1533 1529 γ-Bisabolene SH 0.069 ± 0.01 - - - - - - - - - RI, GC-MS -
- 1561 Nerolidol SO - - - - - - - - - 0.076 ± 0 - GC-MS
- 1577 Spathulenol SO - - - 0.184 ± 0 - - - 0.118 ± 0.09 - GC-MS
- 1579 Isoaromadendrene epoxide SO - - - 0.086 ± 0 - - - - - - - GC-MS
1593 1582 Caryophylene oxide SO 2.941 ± 0.24 3.295 ± 0.26 2.036 ± 1.01 3.013 ± 0.2 RI, GC-MS GC-MS
1621 1608 Humulene epoxide II SO 0.154 ± 0.02 0.158 ± 0 0.128 ± 0.01 0.139 ± 0 RI, GC-MS GC-MS
1643 1638 Isospathulenol SO 0.492 ± 0.05 0.421 ± 0.32 0.463 ± 0.02 0.526 ± 0.08 RI, GC-MS GC-MS
1668 1651 Pogostole SO 0.154 ± 0.02 0.215 ± 0.1 - - - 0.09 ± 0 - GC-MS
- 1658 Neointermedeol SO - - - 0.054 ± 0.01 - - - - - - - GC-MS
1675 1665 Intermedeol SO 0.05 ± 0.02 0.07 ± 0.01 - - - - - - RI, GC-MS GC-MS
1950 1938 Pellitorine O - - - 0.006 ± 0.01 1.191 ± 0.09 1.669 ± 0.06 RI, GC-MS GC-MS
- 1953 Hexadec-9-enoic acid FAD - - - - - - - - - 0.103 ± 0 - GC-MS
- 1959 Palmitic acid FAD - - - - - - - - - 0.423 ± 0.04 - GC-MS
- 2141 Oleic Acid FAD - - - - - - - - - 0.267 ± 0.03 - GC-MS
2707 2707 β-Monolinolein E - - - - - - 1.84 ± 0.98 - - - RI, GC-MS -
2018 NA Heptadec-14-enal A - - - - - - 0.122 ± 0.04 - - - RI, GC-MS -
2815 NA β-Monoolein E - - - - - - 0.503 ± 0.6 0.52 ± 0.06 RI, GC-MS GC-MS
- NA Hexadec-9-en-1-ol O - - - - - - - - - 0.238 ± 0.01 - GC-MS
- NA Octadec-17-ynoic acid FAD - - - - - - - - - 0.025 ± 0.02 - GC-MS
- NA Tetradec-9-enal A - - - - - - - - - 0.073 ± 0.01 - GC-MS
- NA Kalecide O - - - - - - - - - 0.181 ± 0 - GC-MS
- NA Ricinoleic acid FAD - - - - - - - - - 0.231 ± 0.01 - GC-MS
- NA Sabinol SO - - - 0.072 ± 0.01 - - - - - - - GC-MS
Total identified [%] 99.4 ± 0.2 99.2 ± 0.1 98.7 ± 0.7 96.7 ± 0.1
Footnotes 3: a) RI = retention indices for HP-5 column; Obs = retention indices determined relative to a homologous series of n-alkanes (C8-C40) on a HP-5MS column, Lit = literature RI values (Adams, 1995), NA = RI values were not available in the literature. b) C = Class; A - Aldehydes, DH - Diteprene hydrocarbons, E - Esters, FAD - Fatty acid and fatty acid derivatives, MH - Monoterpene hydrocarbons, MO - Oxygenated monoterpenes, O - Others, SH - Sesquiterpene hydrocarbons, SO - Oxygenated sesquiterpenes, d)Identification method: GC-MS = Mass spectrum was identical to that of National Institute of Standards and Technology Library (ver. 2.0.f), RI = the retention index was matching literature database; Std = constituent identity confirmed by co-injection of authentic standards. e) Retention indices were not calculated for compounds calculated only by DB-HeavyWAX colu.mn.
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