Skip to main content
Plants logoLink to Plants
. 2023 Jan 2;12(1):188. doi: 10.3390/plants12010188

Volatile Compounds in Norway Spruce (Picea abies) Significantly Vary with Season

Katja Schoss 1,*, Nina Kočevar Glavač 1, Samo Kreft 1
Editors: Alen Albreht1, Mitja Križman1, Katerina Naumoska1
PMCID: PMC9824094  PMID: 36616317

Abstract

Norway spruce (Picea abies) is one of the most important commercial conifer species naturally distributed in Europe. In this paper, the composition and abundance of essential oil and hydrosol from the needles and branches of P. abies were investigated with an additional evaluation of changes related to different times of the year, annual shoots and branches, and differences in composition under different microenvironments. Essential oils and hydrosols obtained via hydrodistillation were analyzed using gas chromatography–mass spectrometry (GC-MS), where 246 compounds in essential oil and 53 in hydrosols were identified. The relative amounts of monoterpenes, sesquiterpenes, and diterpenes in essential oil changed significantly during the year, with the highest peak of monoterpenes observed in April (72%), the highest abundance of sesquiterpenes observed in August (21%), and the highest abundance of diterpenes observed in June (27%). The individual compound with the highest variation was manool, with variation from 1.5% (April) to 18.7% (June). Our results also indicate that the essential oil with the lowest allergenic potential (lowest quantity of limonene and linalool) was obtained in late spring or summer. Location had no significant influence on composition, while the method of collection for distillation (whole branch or annual shoots) had a minor influence on the composition. All nine main compounds identified in the hydrosol samples were oxygenated monoterpenes. The composition of P. abies hydrosol was also significantly affected by season. The method of preparing the branches for distillation did not affect the composition of P. abies hydrosol, while the location had a minor effect on composition.

Keywords: essential oil, hydrosol, chemical investigation, Picea abies, GC-MS

1. Introduction

Since ancient times, plants have been a source of bioactive compounds that humans use for beneficial health effects and as a resource for drug development. At present, there is significant interest in the use of natural plant-based bioactive compounds, especially when the material is sustainably accessible and can work within the circular economy. An important example of such a material is the branches of Picea abies (L.) H. Karst (Pinaceae), commonly known as European or Norway spruce, which is a tall essential-oil-bearing evergreen conifer used mostly for its wood [1,2]. P. abies accumulates volatile compounds in all parts of the plant, with the highest abundance in needles [3], which is also the least used part in the wood industry and the part of the plant with the largest volume [4]. Thus, in terms of sustainability, it is logical to exploit these needles for beneficial purposes.

Essential oils or volatile compounds (terpenes, terpenoids, and phenylpropanoids) are formed in aromatic plants as secondary metabolites [5] and play an important role in plant defense. Essential oils are typically produced through a distillation process where two products are obtained: an essential oil and a hydrosol. In European Pharmacopoeia, essential oils are described as odorous products, usually of complex composition, which are obtained from a botanically defined plant raw material via steam distillation, dry distillation, or a suitable mechanical process without heating [6]. Hydrosols, also known as hydrolats or aromatic waters, are not yet described in the European Pharmacopoeia. Hydrosols are generally known as the aqueous phase obtained from distillation that separates them from the essential oil phase and are highly diluted, acidic, and scented solutions containing variable amounts of essential oil along with other volatile water-soluble compounds (up to approximately 0.1%) [7,8]. While the scientific literature on essential oils is relatively abundant, much less is known about hydrosols. The compositional profiles of hydrosols may or may not qualitatively overlap with those of the corresponding essential oils, although hydrosol profiles typically differ significantly in quantitative terms [7].

Essential oils are known for their variety of activities. For example, conifer essential oils are known for their antibacterial, antifungal, insect larvicidal, antioxidant, anti-inflammatory, cytotoxic, and antiproliferative activities [9,10,11]. The investigation of hydrosol biological activity has focused mainly on their antimicrobial, antioxidant, and anti-inflammatory activities [7,12].

The volatile compounds of coniferous species are monoterpenes, sesquiterpenes, and diterpenes [10]; however, the qualitative and quantitative composition of such compounds depends on several factors, such as the anatomical part of the tree (needles, branches, or cones), genetic factors, and the health condition of the plant, as well as the environment, location, light quality, seasonal variations, and isolation and determination techniques used for analysis [5,10,11,13,14,15]. Thus, the composition of essential oils and hydrosols varies even within the same species. Since biological activity is dependent on chemical composition, such activity is similarly subject to variation. Although hydrosols are produced through the same distillation process as essential oils, their analyses have been the subject of a limited number of publications, despite their current popularity in the aromatherapy, food, and cosmetic industries [8].

The aim of this study was to investigate the composition and abundance of essential oils and hydrosols from the needles of P. abies and evaluate changes during different times of the year, as well as comparing the composition of annual shoots and branches and differences in composition in different microenvironments. The effects of seasonal variations on the chemical and biological characteristics of some essential oils of the Pinaceae family were previously reported in the literature [3,16,17]. However, no detailed reports are available on the seasonal chemical composition of the essential oil of P. abies. To the best of our knowledge, this is also the first publication on the composition of P. abies hydrosol.

2. Results and Discussion

2.1. Essential Oil Yield

The yield of essential oil in the annual shoots and branches of P. abies ranged from 0.02 to 0.34% (mL relative to fresh material) (Table 1), which means that 10 to 170 µL of essential oil was obtained during the distillation process from 50 g of fresh material. In the whole branch samples, the minimum oil yield of 10 µL was found in location 4, while the maximum yield (170 µL) was found in location 1. The yield was significantly (p = 0.001) influenced by the time of harvesting, with the highest in the fall, the lowest in both spring harvests, and another slight increase over the summer. The location and difference between the whole branch and shoots had no statistical effect on the amount of essential oil. The yield results are in agreement with those of Baath et al. [17], who observed an average yield from fresh needles of 0.07%. In contrast, our results are lower than the results from Radulescu et al. [11] and Visan et al. [10], who recorded an average abundance of essential oils of 1.01% and 1.02%, respectively. However, the yield in those studies was determined for dried plant material.

Table 1.

Yield of essential oil (%) obtained from 50 g of each sample.

Date Preparation of Branches Location 1 Location 2 Location 3 Location 4 Average
5 November 2017 Annual shoots 0.22 0.08 0.06 0.12 0.12
Whole branch 0.34 0.12 0.08 0.08 0.16
4 April 2018 Annual shoots 0.03 0.10 0.03 0.03 0.05
Whole branch 0.08 0.10 0.03 0.02 0.06
28 May 2018 Annual shoots 0.03 0.04 0.04 0.06 0.04
Whole branch 0.05 0.06 0.09 0.06 0.07
Annual shoots—ground 0.09 0.07 0.08 0.16 0.10
13 August 2018 Annual shoots 0.10 0.07 0.12 0.06 0.08
Whole branch 0.07 0.05 0.08 0.08 0.07
Average - 0.11 0.07 0.07 0.07 0.08 *

* without the results for “annual shoots—ground” samples.

2.2. Essential Oil Composition

A total of 246 different compounds were found in 32 samples, as presented in Appendix A. The abundance of all compounds is expressed as a percentage of the relative peak areas (percentage of the area of the chromatographic peak of an individual compound compared to the sum of the areas of all chromatographic peaks). A total of 17 compounds had an average abundance greater than 1.5% and are presented in Table 2. Only 8 compounds were detected in all 32 samples: limonene, β-pinene, borneol, camphene, abienol, α-pinene, T-muurolol, and α-humulene (in Table 2 marked in bold). The compound with the highest average relative peak area percentage detected in the samples, despite not being present in all samples, was manool, with a maximum abundance of 40.88% and an average abundance of 10.96%. In some studies, no diterpenes were detected in essential oils, as GC temperatures were set below 250 °C [9,18], while in studies using GC methods above 250 °C, manool was detected in greater quantities (Visan et al. [10]: 9.4%; Radulescu et al. [11]: 2.6–6.2%). The same applies to α-cadinol, which was observed at a maximum of 6.41% in our study and a maximum of 16.56% in the study by Garzola et al. [9]. In the study by Kartnig et al. [18], this compound was not observed, and in Visan et al. [10] and Radulescu et al. [11], this compound was present in 3.8% and 11.2–25.3%, respectively.

Table 2.

List of volatile compounds whose average abundance, according to their relative peak intensity (RPI) (Area %), was greater than 1.5% in 32 essential oil samples, with their minimum, maximum, and average abundance obtained at different season times and with different plant parts with included p-value as detected by gas chromatography-mass spectrometry. The compounds were identified based on their mass spectra and retention indices (RI; Db, database RI; Ms, measured RI).

Compound RI Minimum RPI (%) Maximum RPI (%) Average RPI (%) Standard Deviation No. of Samples without Compound 17 November 18 April 18 June 18 August Seasons Branch Annual Shoots Collection Method
Chemical Group Ms Db Average STD Average. STD Average. STD Average STD p Average STD Average STD p
α-pinene 930 933 0.10 9.18 3.25 3.12 0 5.47 3.30 4.84 2.74 0.56 0.48 2.14 2.51 0.00 3.87 3.35 2.64 2.84 0.27
monoterpene hydrocarbon
camphene 946 942 0.15 16.44 3.96 4.19 0 6.93 4.80 4.78 1.20 0.51 0.28 3.61 5.35 0.01 3.64 3.91 4.27 4.56 0.68
monoterpene hydrocarbon
β-pinene 974 978 0.11 28.51 5.59 6.63 0 7.68 5.99 10.56 9.02 1.15 1.34 2.99 3.49 0.01 7.77 7.97 3.41 4.14 0.06
monoterpene hydrocarbon
myrcene 987 991 0.00 10.95 3.23 3.09 2 4.04 2.90 5.04 2.22 0.61 0.49 3.23 4.04 0.02 3.20 2.92 3.26 3.34 0.96
monoterpene hydrocarbon
limonene 1027 1030 0.19 22.78 6.86 6.00 0 9.05 4.36 12.53 6.21 1.20 0.82 4.66 4.10 0.00 7.47 6.17 6.25 5.96 0.58
monoterpene hydrocarbon
eucalyptol 1030 1032 0.00 6.04 1.70 1.81 7 1.86 1.53 3.90 1.51 0.15 0.24 0.88 0.84 0.00 1.65 1.82 1.75 1.85 0.88
monoterpene ketone
camphor 1144 1149 0.00 3.81 1.79 1.20 2 2.16 0.96 3.12 0.40 0.80 0.58 1.07 1.02 0.00 1.83 1.12 1.75 1.30 0.85
monoterpene ketone
camphene hydrate 1156 1152 0.00 3.41 1.51 0.94 2 1.38 0.45 2.58 0.47 0.95 0.59 1.11 1.11 0.00 1.46 0.81 1.55 1.07 0.79
monoterpene alcohol
borneol 1169 1173 0.48 11.67 4.50 2.54 0 4.09 1.48 6.47 2.15 3.34 1.38 4.11 3.67 0.07 4.30 2.11 4.70 2.97 0.67
monoterpene alcohol
bornyl acetate 1282 1285 0.00 32.19 9.84 6.93 1 9.40 3.05 17.64 8.54 4.19 2.60 8.14 3.75 0.00 8.15 4.63 11.53 8.46 0.17
monoterpene ester
(E)-caryophyllene 1418 1424 0.00 9.97 2.64 2.11 1 2.92 2.17 2.18 1.83 3.24 3.12 2.23 1.00 0.71 3.18 2.39 2.11 1.70 0.16
sesquiterpene hydrocarbon
α-humulene 1453 1454 0.22 8.82 1.94 2.00 0 1.94 1.90 0.76 0.60 3.38 2.97 1.69 0.91 0.06 1.76 1.53 2.12 2.42 0.62
sesquiterpene hydrocarbon
δ-cadinene 1516 1518 0.00 6.25 2.03 1.75 4 2.60 1.67 0.34 0.27 2.16 1.86 3.01 1.57 0.01 1.82 1.69 2.23 1.84 0.52
sesquiterpene hydrocarbon
T-muurolol 1642 1645 0.16 10.58 2.38 1.99 0 1.77 1.23 1.16 1.08 3.45 3.09 3.15 1.06 0.05 2.36 2.60 2.40 1.17 0.96
sesquiterpene alcohol
α-cadinol 1653 1659 0.00 16.56 6.41 4.87 3 5.46 3.83 2.44 1.91 7.18 5.32 10.55 4.38 0.01 5.85 5.35 6.96 4.45 0.53
sesquiterpene alcohol
manool 2050 2062 0.00 40.88 10.96 11.83 4 10.21 9.49 1.51 1.59 18.73 11.43 13.38 14.76 0.02 11.88 12.94 10.03 10.95 0.67
diterpene alcohol
abienol 2141 2152 0.19 21.48 3.83 4.29 0 1.13 0.89 2.32 1.92 8.44 6.08 3.44 2.20 0.00 4.39 5.41 3.27 2.87 0.47
diterpene alcohol
17 November 18 April 18 June 18 August Branch Annual shoots
Monoterpene hydrocarbons 22.89 33.17 37.75 4.03 16.63 25.95 19.83
Oxygenated monoterpenes (alcohols, esters, ketones) 19.34 18.89 33.71 9.43 15.31 17.39 21.28
Sesquiterpene hydrocarbons 6.61 7.46 3.28 8.78 6.93 6.76 6.46
Oxygenated sesquiterpenes (alcohols, esters, ketones) 8.79 7.23 3.6 10.63 13.7 8.21 9.36
Diterpene alcohols 14.79 11.34 3.83 27.17 16.82 16.27 13.3

The main compound classes in the studied essential oils were monoterpenes (monoterpene hydrocarbons: 22.89%; oxygenated monoterpenes: 19.34%), followed by sesquiterpenes (sesquiterpene hydrocarbons: 6.61%; oxygenated sesquiterpenes: 8.79%) and diterpenes (diterpene alcohols: 14.79%).

These results are in partial agreement with four other studies. Baath et al. [17], who investigated the essential oil composition of 16 different conifer species, including P. abies, reported α-pinene, camphene, limonene, and bornyl acetate to be the four predominant compounds; in our study, these compounds ranked ninth, seventh, third, and second. In a study by Garzoli et al. [9], the main components analyzed by GC-MS were β-pinene (44.7%), α-pinene (20.2%), limonene (14.2%), and camphene (7.2%); in our study, these compounds ranked fifth, ninth, third, and seventh, respectively. When analyzed by GC-headspace [9], the results were as follows: β-pinene (43.8%), α-pinene (34.5%), camphene (10.5%), and limonene (8.0%). In a study by Visan et al. [10], the compounds with the highest abundance were limonene (21.1%), α-pinene (11.6%), bornyl acetate (11.08%), camphene (10.7%), and manool (9.4%). In a study by Radulescu [11], the composition of essential oil was also evaluated considering geographical aspects. Plants were collected from different parts of Romania, and the geographical influence was significant, but the highest- abundance compounds remained similar throughout (α-cadinol, 11.2–25.3%; bornyl acetate, 11.8–15.0%; camphene, 3.8–14.1%; and limonene, 6.3–13.0%. These compounds were ranked in our study as twentieth, second, seventh, and third, respectively (see Appendix A)).

Most compounds present in higher percentages were previously studied for their potential biological or therapeutic activities, which suggests possible usages of P. abies essential oil in conventional and complementary medicine, such as aromatherapy. Manool possesses antigenotoxic, anticarcinogenic, and anti-inflammatory potential [19], while an anti-melanoma effect was shown in vivo in mice [20]. Bornyl acetate possesses anti-inflammatory [21] properties; limonene possesses antihyperalgesic [22,23], antidiabetic [24], anti-inflammatory [25], and antioxidative [25] potential; α-cadinol has antifungal [26] properties; β-pinene antimicrobial [27] has antioxidative properties [28] and, in combination with linalool, antidepressant properties [29]; borneol has antiglycemical [30], antihyperlipidemic [30], antioxidative [30], and antinociceptive [31] properties; camphene has antitumor [32], antioxidative [33], and hypolipidemic [34] properties; abienol has antifungal, antimicrobial, and antineoplastic properties [35]; α-pinene has antimicrobial [27,36], antioxidative [28], anti-inflammatory [37], gastroprotective [38], and antinociceptive properties [39]; myrcene has antioxidative [40] and anti-inflammatory properties [41]; (E)-caryophyllene has anti-inflammatory [42], anticonvulsant [43], and antinociceptive properties [44]; T-muurolol has antifungal properties [26]; δ-cadinene has anticancer [45] and acaricidal properties [46]; α-humulene has anti-inflammatory properties [47]; and camphor has anti-inflammatory [48], eucalyptol antibacterial [49], anti-inflammatory [50,51], and antioxidative [51] properties.

P. abies essential oil is also widely used as a cosmetic ingredient, where it is desirable for the oil to contain monoterpene abundance that is as low as possible, since monoterpenes are generally responsible for allergic reactions. In P. abies essential oil, citronellol and limonene were recognized as typical allergens [52], and their use in cosmetic products is regulated [53]. Limonene was also found among the main compounds in the studied samples (average yield of 6.86%). In addition, some monoterpenes are highly prone to oxidation, such as α-pinene and β-pinene (in this study, they were detected at a yield of 3.25% and 5.59%, respectively), and air-oxidized products were shown to be potent skin allergens [54].

2.3. Variation of Essential Oil Composition

Comparing the composition of essential oils obtained from plant materials at different times of the year, we found that shares of monoterpenes, sesquiterpenes, and diterpenes greatly varied during the year, as shown in Table 2. The abundance of both monoterpene hydrocarbons and oxygenated monoterpenes was the highest in April (38% and 34%, respectively) and lowest in June (4% and 9%, respectively). The abundance of total sesquiterpenes was highest during August (20.63%) and lowest during April (6.88%), and the abundance of diterpenes was highest during June (27.17%) and lowest during April (3.83%). The individual compounds that varied most significantly (more than 5%) were manool (18.73% in June and 1.51% in April), bornyl acetate (17.64% in April and 4.19% in June), limonene (9.05% in November and 1.20% in June), α-cadinol (10.55% in August and 2.44% in April), and β-pinene (10.56% in April and 1.15% in June).

Organoleptically, in terms of odor, the most desirable component of P. abies essential oil is bornyl acetate, which provides the typical pleasant smell of needles, while the least desirable is myrcene, which produces an unpleasant smell of plastic. Both compounds were highest in early spring, fell sharply over the course of spring and then slowly increased again through summer and autumn.

The allergens limonene and linalool varied in the same way, i.e., with a peak in April, a sharp decrease in June, and a subsequent increase from August to November, while a minor amount of citronellol (0.01%) was detected only in April. Our results indicate that the essential oil with the least allergenic potential was obtained in late spring or summer.

A study by Kamaitytė-Bukelskienė et al. [3] studied the content variation of α-pinene and β-pinene from May to September and observed the highest abundance in May, which corresponds well with our results.

We conclude that season has the strongest influence on the composition of P. abies essential oil, generally providing the largest change (increase or decrease) from April to June. Of the 17 compounds with the highest abundance, differences between seasons were significant (p < 0.05) for 13 compounds (Table 2). The abundance of all seven sesquiterpenes and diterpenes increased sharply in spring, while at the same time, the abundance of all ten monoterpenes dropped. The largest change was observed for the diterpene manool and the monoterpenes limonene and eucalyptol. Manool increased by more than a factor of 10, and limonene and eucalyptol decreased by a factor of more than 10.

The location had no significant influence on the composition of spruce essential oil, which was expected since the distances between the locations were smaller than 20 km. Nevertheless, the sampling on four locations additionally supported the dependence on seasonal variation.

The method of branch collection had little effect on the composition of P. abies essential oil (Table 2). The differences between the essential oil in annual shoots and whole branches were significant for none of the 17 compounds with the highest abundance. The largest difference was observed in the abundance of β-pinene, which was 2× more common in the essential oil from whole branches than in the essential oil from annual shoots. Conversely, the abundance of bornyl acetate in essential oil from whole branches was 30% lower than that in essential oil from annual shoots. Since bornyl acetate is a desired component of P. abies essential oil and β-pinene is not, better quality essential oil can be obtained using only annual shoots, but this requires more manual labor.

The method used for preparing the branches before distillation (the annual shoots of the third harvesting term were prepared by grinding in addition to the standard method) did not significantly affect the composition of the essential oil (data not shown), but the yield was 2.5× higher in the distillation of grinded samples (p = 0.017) (see Table 1).

Based on the data from this study, we conclude that the composition of P. abies essential oil varies greatly with season but not with short-distance geographic origin or the preparation of distilled material.

2.4. Hydrosol Composition

Although hydrosols are produced through the same distillation process as essential oils, their analyses have been subject to a limited number of publications. To the best of our knowledge, this is the first publication on the composition of P. abies hydrosol. Similar to essential oils, it is believed that the composition of hydrosols produced from plant parts of the same species differs when collected in different countries, different geographical locations within a country, different seasons, or at different plant growth stages, consequently altering their biological activities [8]. The applied analysis technique, or the preparation of the sample for analysis (extraction with organic solvent or direct analysis), also has an important influence on the results. As shown by Kokalj Ladan et al. [55], direct hydrosol analysis should be used when the total composition of a hydrosol is the research focus because it allows for the identification of hydrophilic compounds that are not extracted into the organic solvent phase, and true ratios among compounds are obtained. However, it has to be emphasized that direct analysis has poorer repeatability. Based on the above-mentioned advantages of direct hydrosol analysis this approach was selected for compositional analysis in this study.

The first observation based on the results shown in Table 2 and Table 3 shows a large difference in the qualitative and quantitative composition of the essential oil and hydrosol. In the 32 hydrosol samples of P. abies, a total of 53 different compounds were found. The abundances of all compounds are expressed here as a percentage of the relative peak areas (percentage of the area of the chromatographic peak of an individual compound compared to the sum of the areas of all chromatographic peaks). This percentage, at the same time, corresponds to the weight percentage representation of this compound in relation to the non-aqueous (terpene) part of the hydrosol. Nine compounds had abundance greater than 1.2% (Table 3).

Table 3.

List of volatile compounds whose average abundance according to their relative peak intensity (RPI) (Area %) in 32 hydrosol samples was greater than 1.2% with their log P, minimum, maximum, and average abundance obtained at different season times and with different plant parts with included p-value as detected by gas chromatography-mass spectrometry. The compounds were identified based on their mass spectra and retention indices (RI; Db, database RI; Ms, measured RI).

Compound RI logP Minimum RPI (%) Maximum RPI (%) Average RPI (%) Standard Deviation No. of Samples without Compound 17 November 18 April 18 June 18 August Seasons Branch Annual Shoot Collection Method
Chemical Group Ms Db Average STD Average STD Average STD Average STD p Average STD Average STD p
(2E)-2-hexenal 841 850 2.38 0 19.9 2.65 5.25 22 8.7 7 0 0 0 0 1.89 3.79 0 2.92 5.25 2.38 5.40 0.78
artefact
(3Z)-3-hexen-1-ol 850 853 2.38 0 19.83 5.46 5.80 11 8.78 6.4 0 0 6.58 4.06 6.47 6.51 0.01 4.98 6.43 5.94 5.27 0.65
artefact
eucalyptol 1030 1032 2.82 0 26.15 8.40 7.80 10 8.27 4.25 17.21 4.84 0.42 1.18 7.7 8.02 0.00 8.14 7.51 8.66 8.31 0.86
monoterpene ether
camphor 1144 1149 2.13 0 25.04 13.06 6.58 3 13.52 4.54 18.92 4.56 9.02 6.9 10.77 6.21 0.01 14.09 6.73 12.02 6.47 0.38
monoterpene ketone
camphene hydrate 1152 1156 2.77 2.86 18.11 12.05 4.15 0 9.85 1.93 15.27 1.44 10.99 5.09 12.08 5.1 0.05 12.35 4.54 11.75 3.85 0.69
monoterpene alcohol
borneol 1169 1173 2.71 0 49.62 27.97 11.85 2 21.65 4.52 29.05 7.33 34.76 11.07 26.43 17.98 0.16 29.86 9.79 26.08 13.67 0.38
monoterpene alcohol
isoborneol 1169 1165 2.71 0 19.96 1.23 4.85 2 0 0 0 0 0 0 4.93 9.12 0.096 0.00 0.00 2.46 6.73 0.15
monoterpene alcohol
terpinen-4-ol 1184 1178 2.99 0 40.32 5.55 10.07 6 1.87 0.62 2.52 0.38 11.08 14 6.71 13.78 0.23 6.06 11.88 5.03 8.23 0.78
monoterpene alcohol
α-terpineol 1193 1195 2.79 0 35.82 12.35 8.53 4 8.36 2.34 12.42 2.15 10.97 11.32 17.63 11.46 0.17 10.87 6.54 13.82 10.14 0.34
monoterpene alcohol
Average 17 November 18 April 18 June 18 August / Branch Annual shoots /
Oxigenated monoterpenes 88.72 81.00 95.39 83.82 94.61 / 89.27 88.14 /

All of these compounds were oxygenated monoterpenes, with the exception of hexenol and hexenal, which were likely artefacts. The dominant abundance of oxygenated monoterpenes in hydrosols is not unusual and also appeared in other studies on the composition of hydrosols of conifers [56,57]. Only one compound (camphene hydrate) was detected in all 32 samples.

Among the most abundant compounds in hydrosols, only four were also the most abundant in the essential oil: borneol, camphor, camphene hydrate, and eucalyptol. All the most commonly represented compounds in hydrosols had a logP value (octanol/water) of less than 3, indicating that they are hydrophilic to moderately lipophilic. Compounds that were strongly represented in the essential oil but not in the hydrosol all had a logP greater than 4. Bornyl acetate, which has a logP of 3.6 (i.e., between 3 and 4), was present in the hydrosol with less than an average of 1%, ranked 14th and present in the essential oil with an average of 32% (ranked second). It should also be taken noted that the solubility of a compound in water at a neutral pH may be different than that at a pH between 3.5 and 4, which is the typical pH of a hydrosol [56,57].

The composition of P. abies hydrosol was, as is common for essential oils, significantly affected by the season. Of the nine compounds with the highest abundance, differences between seasons were significant for five compounds. However, the pattern of changes was not as uniform as that with essential oils. Only with eucalyptol and camphor did we observe the same pattern as that seen in essential oils, i.e., with the maximum abundance appearing in early spring (April), followed by a sharp fall in June and then a slow increase again by November.

The method of preparing the branches for distillation did not affect the composition (Table 3). The differences between collections were significant for none of the nine compounds with the highest abundance.

The location, moreover, had little effect on the composition of spruce hydrosol. The differences between sites were significant (data not shown) for none of the nine compounds with the highest abundance.

3. Materials and Methods

3.1. Plant Material and Sample Preparation

Branches of Picea abies (L.) Karst. were collected at four locations in the municipality of Loški Potok, Slovenia, in the settlements of Blošček (location 1; N: 45°43′59″, E: 14°33′13″; 809 m), Bela Voda (location 2; S: 45°41′20″, E: 14°37′09″; 924 m), Hrib (location 3; S: 45°41′49″, E: 14°37′16″; 868 m), and Lazec (location 4; S: 45°39′28″, E: 14°37′36″; 802 m). Plants were identified by Samo Kreft, and the voucher specimen of the plant was deposited in the Herbarium of the Department of Pharmaceutical Biology, Faculty of Pharmacy, University of Ljubljana, Slovenia (voucher No. BF00159/2017). Branches were collected four times (5 November 2017; 4 April 2018; 28 May 2018; and 13 August 2018) from growing (uncut) trees, up to a height of 5 m and cut at a branch thickness of approximately 1 cm.

Fresh whole branch samples, including annual shoots, were collected from every location and cut into 5 to 10 cm long pieces. Additionally, annual shoots (approximately 10 cm long unbranched thin twigs with needles) were collected separately.

In total, 50 g of each branch sample was put into a 1000 mL round flask with 500 mL of demineralized water and distilled for 4 h using a Clevenger-type apparatus according to general instructions of European Pharmacopoeia, “2.8.12. Determination of essential oils in herbal drugs” (6). No solvent was used during hydro-distillation. All samples (hydrosols and essential oils) were stored after distillation until analysis in glass tubes sealed with a glass stopper and frozen at −20 °C.

For additional comparison, distillation from ground annual shoots obtained from all locations at one time period (28 May 2018) was also performed (the particle size of the branches and needles was approximately 3 mm).

3.2. Gas Chromatography Coupled with Mass Spectrometry (GC-MS)

Essential oil (diluted in n-hexane (Suprasolv, Merck, Germany) (1 mg/mL)) and hydrosol (unprocessed) samples were analyzed on a Shimadzu GC-MS system GCMS-QP2010 Ultra equipped with a MS detector and Rxi-5Sil MS (Restek, Bellefonte, PA, USA) capillary column (30 m × 0.25 mm, film thickness 0.25 μm). The injector and ion source temperatures were set to 250 and 200 °C, respectively. The column temperature was programmed from 50 to 250 °C at a rate of 3 °C/min, and the lower and upper temperatures were held for 5 min. Helium (99.99%) was used as carrier gas at a flow rate of 1 mL/min. A sample of 1.0 μL was injected with an autosampler using the split mode with a split ratio of 1:100. For MS detection, an electron ionization mode with an ionization energy of 70 eV was used. The MS transfer line temperature was set to 250 °C. The m/z range was from 40 to 400 with a scanning frequency of 5 Hz.

Identification of the compounds was done based on a comparison of their mass spectra and retention indices with those of the synthetic compounds in the spectral library of the National Institute of Standards and Technology (NIST11), as well as the Flavors and Fragrances of Natural and Synthetic Compounds spectral library (FFNSC2). The linear retention indices were determined in relation to a homologous series of n-alkanes (C6-C24). Component relative concentrations were calculated from the GC peaks without using correction factors.

3.3. Statistical Analysis

All of the GC-MS analyses were conducted in duplicates. Statistical analysis of the data was performed by calculating the average, standard deviation (STD) and analysis of variance (ANOVA) using the SPSS 26 software. No variable transformation was applied. A probability value at p ≤ 0.05 was considered statistically significant.

4. Conclusions

The main compounds of P. abies essential oil analyzed in the present study were manool, bornyl acetate, limonene, α-cadinol, and β-pinene, while those of hydrosol were borneol, camphor, and α-terpineol. The comparison of essential oil composition between seasons showed significant variation in the abundance of monoterpenes and sesquiterpenes, as well as significant variation of monoterpenes in hydrosols.

Since the essential oil and hydrosol of P. abies have a wide option of possible applications in the pharmaceutical, cosmetic, and food industries, it is of great importance to define the composition of P. abies and consider variations in composition during the year. The information observed on seasonal variation may be useful in selecting the best season for essential oil or hydrosol distillation, as well as for choosing the product with the most desirable properties, such as the essential oil with the fewest allergens.

Acknowledgments

The authors are grateful to Damjana Gregorič, M.Pharm. for providing the samples of fir branches, to Municipality of Loški Potok (Slovenia) for financing the GC analyses and to Irena Klančnik Mavec for skillful laboratory assistance.

Appendix A

List of volatile compounds whose average abundance, according to their relative peak intensity (RPI) (Area %), was greater than 0.1% in 32 essential oil samples, with their minimum (Min), maximum (Max), and average abundance as detected by gas chromatography-mass spectrometry. The compounds were identified based on their mass spectra and retention indices (RI; Db, database RI; Ms, measured RI). Unidentified compounds are presented as numbers according to their five most intensive mass ion peaks.

Compound RI Min Max Average RPI (%) Standard Deviation
Ms Db
manool 2050 2062 0.00 40.88 10.96 11.83
bornyl acetate 1282 1285 0.00 32.19 9.84 6.93
limonene 1027 1030 0.19 22.78 6.86 6.00
cadin-4-en-10-ol 1653 1659 0.00 16.56 6.41 4.87
β-pinene 974 978 0.11 28.51 5.59 6.63
borneol 1169 1173 0.48 11.67 4.50 2.54
camphene 946 942 0.15 16.44 3.96 4.19
abienol 2141 2152 0.19 21.48 3.83 4.29
α-pinene 930 933 0.10 9.18 3.25 3.12
myrcene 987 991 0.00 10.95 3.23 3.09
(e)-caryophyllene 1418 1424 0.00 9.97 2.64 2.11
t-muurolol 1642 1645 0.16 10.58 2.38 1.99
delta-cadinene 1516 1518 0.00 6.25 2.03 1.75
α-humulene 1453 1454 0.22 8.82 1.94 2.00
camphor 1144 1149 0.00 3.81 1.79 1.20
eucalyptol 1030 1032 0.00 6.04 1.70 1.81
camphene hydrate 1156 1152 0.00 3.41 1.51 0.94
243 (100) 271 (88) 43 (88) 41 (84) 286 (79) 0.00 43.00 1.43 7.59
α-terpineol 1192 1195 0.34 2.75 1.41 0.65
epi-α-cadinol 1640 1640 0.00 3.64 1.31 1.20
epimanool 2048 2057 0.00 20.45 1.24 4.04
α-bisabolol oxide a 1745 1748 0.00 34.96 1.23 6.20
germacrene d 1473 1480 0.00 4.20 1.01 1.04
longifolene 1407 1412 0.00 4.02 0.86 0.93
(e,e)-α-farnesene 1501 1504 0.00 8.01 0.81 1.68
41 (100) 93 (99) 81 (87) 91 (78) 105 (78) 0.00 12.00 0.80 2.28
α-terpinyl acetate 1344 1349 0.00 2.33 0.77 0.63
43 (100) 81 (79) 41 (70) 93 (66) 107 (64) 0.00 5.14 0.75 1.49
spathulenol 1574 1576 0.00 2.24 0.72 0.59
α-muurolol 1644 1651 0.00 1.48 0.64 0.44
beyerene 1921 1933 0.00 2.99 0.63 0.78
δ-3-carene 1007 1009 0.00 9.04 0.63 1.64
43 (100) 119 (46) 109 (42) 93 (37) 108 (27) 0.00 2.11 0.60 0.60
dodeca-(8e,10e)-dienyl acetate 1655 1665 0.00 3.06 0.55 0.72
caryophyllene oxide 1579 1587 0.00 2.29 0.48 0.56
α-muurolene 1496 1497 0.00 1.23 0.37 0.32
135 (100) 91 (20) 286 (20) 148 (20) 93 (17) 0.00 2.46 0.34 0.64
cyclosativene 1366 1367 0.00 10.54 0.33 1.86
manool oxide 1990 1989 0.00 1.24 0.32 0.36
terpinolene 1083 1086 0.00 1.43 0.31 0.29
santene 879 880 0.00 1.66 0.29 0.36
γ-cadinene 1511 1512 0.00 1.08 0.28 0.28
terpinen-4-ol 1178 1184 0.00 0.62 0.28 0.18
1,2,3,4,4a,7,8,8a-octahydro-4-isopropyl-1,6-dimethyl naphth-1-ol 1638 1641 0.00 2.44 0.27 0.62
(e)-nerolidol 1559 1561 0.00 0.91 0.25 0.22
tricyclene 920 923 0.00 1.27 0.25 0.33
α-bisabolol oxide b 1651 1655 0.00 7.94 0.25 1.40
43 (100) 79 (96) 80 (62) 67 (61) 41 (44) 0.00 1.25 0.24 0.33
273 (100) 105 (62) 148 (48) 43 (46) 107 (45) 0.00 5.00 0.23 0.91
hexahydro-benzene 678 682 0.00 1.49 0.22 0.29
α-bisabolone oxide a 1676 1682 0.00 5.02 0.22 0.90
43 (100) 81 (73) 41 (59) 55 (56) 93 (55) 0.00 2.55 0.21 0.48
abietadiene 2080 2089 0.00 0.95 0.21 0.30
109 (100) 105 (96) 119 (73) 41 (68) 91 (68) 0.00 0.97 0.20 0.26
131 (100) 187 (79) 286 (31) 105 (30) 43 (39) 0.00 2.30 0.19 0.51
humulene epoxide ii 1607 1613 0.00 0.78 0.19 0.22
α-cadinene 1535 1538 0.00 3.00 0.18 0.54
epicubenol 1626 1631 0.00 0.46 0.17 0.15
13-epi-manoyl oxide 2012 2022 0.00 0.75 0.17 0.24
isopulegyl acetate 1269 1273 0.00 5.32 0.17 0.94
43 (100) 79 (90) 80 (62) 67 (56) 41 (44) 0.00 0.71 0.16 0.21
spathulenol 1574 1576 0.00 1.42 0.16 0.43
carvotanacetone 1250 1249 0.00 1.61 0.16 0.38
tridecan-2-one 1491 1495 0.00 0.74 0.15 0.22
(100) 41 (88) 43 (84) 93 (81) 93 (81) 0.00 0.71 0.15 0.17
isopropyl-tetradecanoate 1821 1826 0.00 1.55 0.14 0.34
43 (100) 91 (89) 271 (87) 243 (87) 105 (86) 0.00 2.07 0.14 0.41
phytol 2106 2106 0.00 1.81 0.14 0.38
(e)-β-farnesene 1450 1452 0.00 0.81 0.13 0.20
n-hexadecanol 1877 1884 0.00 2.73 0.12 0.48
γ-terpinene 1056 1058 0.00 0.44 0.12 0.13
cis-muurol-5-en-4-alpha-ol 1573 1558 0.00 3.91 0.12 0.69
6z-pentadecen-2-one 1667 1670 0.00 0.67 0.12 0.20
t-muurolol 1642 1645 0.00 2.99 0.12 0.55
β-phellandrene 1028 1031 0.00 3.35 0.12 0.60
43 (100) 79 (88) 41 (54) 91 (51) 67 (50) 0.00 0.54 0.11 0.16
bicyclogermacrene 1492 1497 0.00 1.01 0.11 0.28
131 (100) 187 (69) 41 (57) 93 (56) 67 (55) 0.00 1.28 0.10 0.25
256 (100) 241 (77) 121 (72) 105 (67) 105 (67) 0.00 1.16 0.10 0.26

Appendix B

List of volatile compounds whose average abundance, according to their relative peak intensity (RPI) (Area %), was greater than 0.1% in 32 hydrosol samples, with their minimum (Min), maximum (Max), and average abundance as detected by gas chromatography-mass spectrometry. The compounds were identified based on their mass spectra and retention indices (RI; Db, database RI; Ms, measured RI). Unidentified compounds are presented as numbers according to their five most intensive mass ion peaks.

Compound RI Mini Max Average RPI (%) Standard Deviation
Ms Db
borneol 1169 1173 0.00 49.62 27.97 11.85
camphor 1144 1149 0.00 25.04 13.06 6.58
α-terpineol 1193 1195 0.00 35.82 12.35 8.53
camphene hydrate 1152 1156 2.86 18.11 12.05 4.15
eucalyptol 1030 1032 0.00 26.15 8.40 7.80
terpinen-4-ol 1184 1178 0.00 40.32 5.55 10.07
hex-(3z)-enol 850 853 0.00 19.83 5.46 5.80
hex-(2e)-enal 841 850 0.00 19.90 2.65 5.25
isoborneol 1169 1165 0.00 19.96 1.23 4.85
α-bisabolol oxide a 1744 1748 0.00 33.45 1.16 5.93
n-octacosane 2161 2170 0.00 36.07 1.13 6.38
55 (100) 41 (72) 43 (53) 83 (43) 69 (37) 0.00 11.83 1.03 2.67
43 (100) 119 (47) 109 (39) 93 (39) 108 (25) 0.00 8.09 1.00 1.85
bornyl acetate 1282 1285 0.00 6.30 0.92 1.52
carvotanacetone 1250 1249 0.00 11.96 0.91 2.63
3-hydroxy-pentene 689 680 0.00 5.07 0.68 1.33
44 (100) 45 (35) 43 (20) 41 (18) 93 (17) 0.00 12.94 0.59 2.37
hex-(2e)-enal 824 830 0.00 11.09 0.56 2.17
131 (100) 187 (68) 105 (36) 41 (35) 286 (29) 0.00 9.67 0.37 1.73
43 (100) 72 (76) 108 (71) 71 (67) 93 (63) 0.00 3.72 0.34 0.77
pent-(2z)-enol 764 767 0.00 2.44 0.26 0.64
isobornyl acetate 1282 1285 0.00 3.33 0.24 0.71
135 (100) 91 (22) 286 (21) 41 (18) 93 (17) 0.00 4.86 0.19 0.87
43 (100) 243 (93) 271 (76) 41 (74) 91 (69) 0.00 5.39 0.18 0.95
pent-(2e)-enal 748 751 0.00 1.44 0.14 0.36
linalool 1132 1135 0.00 1.71 0.13 0.42
82 (100) 110 (64) 95 (47) 54 (28) 109 (25) 0.00 1.97 0.11 0.39
cis-hex-3-enal 797 797 0.00 1.35 0.10 0.34
77 (100) 45 (20) 78 (7) 59 (6) 62 (6) 0.00 0.87 0.10 0.25

Appendix C

Sample of GC-MS chromatogram of essential oil and hydrosol obtained from Picea Abies.

Figure A1.

Figure A1

GC-MS chromatogram of hydrosol obtained from Picea abies (Bele vode, avg. 2018).

Figure A2.

Figure A2

GC-MS chromatogram of essential oil obtained from Picea abies (Lazec, avg. 2018).

Author Contributions

Conceptualization, S.K. and N.K.G.; methodology, S.K.; validation, S.K., N.K.G. and K.S.; formal analysis, S.K.; writing—original draft preparation, K.S.; writing—review and editing, N.K.G. and S.K.; supervision, S.K. and N.K.G.; project administration, N.K.G.; funding acquisition, S.K. All authors have read and agreed to the published version of the manuscript.

Data Availability Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

Funding Statement

This research was funded by the municipality of Loški Potok.

Footnotes

Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

References

  • 1.Spinelli S., Costa C., Conte A., La Porta N., Padalino L., Del Nobile M.A. Bioactive Compounds from Norway Spruce Bark: Comparison Among Sustainable Extraction Techniques for Potential Food Applications. Foods. 2019;8:524. doi: 10.3390/foods8110524. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2.Co M., Fagerlund A., Engman L., Sunnerheim K., Sjöberg P.J.R., Turner C. Extraction of antioxidants from spruce (Picea abies) bark using eco-friendly solvents. Phytochem. Anal. 2012;23:1–11. doi: 10.1002/pca.1316. [DOI] [PubMed] [Google Scholar]
  • 3.Kamaitytė-Bukelskienė L., Ložienė K., Labokas J. Dynamics of isomeric and enantiomeric fractions of pinene in essential oil of picea abies annual needles during growing season. Molecules. 2021;26:2138. doi: 10.3390/molecules26082138. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Nastić N., Gavarić A., Vladić J., Vidović S., Aćimović M., Puvača N., Brkić I. Spruce (Picea abies (L.). H. Karst): Different Approaches for Extraction of Valuable Chemical Compounds. J. Agron. Technol. Eng. Manag. 2020 [Google Scholar]
  • 5.Sangwan N.S., Farooqi A.H.A., Shabih F., Sangwan R.S. Regulation of essential oil production in plants. Plant Growth Regul. 2001;34:3–21. doi: 10.1023/A:1013386921596. [DOI] [Google Scholar]
  • 6.European Directorate for the Quality of Medicines (EDQM) European Pharmacopoeia (Ph. Eur.) 10th ed. European Directorate for the Quality of Medicines (EDQM); Strasbourg, France: 2021. [Google Scholar]
  • 7.Aćimović M.G., Tešević V.V., Smiljanić K.T., Cvetković M.T., Stanković J.M., Kiprovski B.M., Sikora V.S. Hydrolates: By-products of essential oil distillation: Chemical composition, biological activity and potential uses. Adv. Technol. 2020;9:54–70. doi: 10.5937/savteh2002054A. [DOI] [Google Scholar]
  • 8.Rao R. Hydrosols and Water-Soluble Essential Oils: Medicinal and Biological Properties. In: Govil J.N., Bhattacharya S., editors. Recent Progress in Medicinal Plants Essential Oils 1. Stadium Press; New Delhi, India: 2013. pp. 120–140. [Google Scholar]
  • 9.Garzoli S., Masci V.L., Caradonna V., Tiezzi A., Giacomello P., Ovidi E. Liquid and Vapor Phase of Four Conifer-Derived Essential Oils: Comparison of Chemical Compositions and Antimicrobial and Antioxidant Properties. Pharmaceuticals. 2021;14:134. doi: 10.3390/ph14020134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Visan D.-C., Oprea E., Radulescu V., Voiculescu I., Biris I.-A., Cotar A.I., Saviuc C., Chifiriuc M.C., Marinas I.C. Original Contributions to the Chemical Composition, Microbicidal, Virulence-Arresting and Antibiotic-Enhancing Activity of Essential Oils from Four Coniferous Species. Pharmaceuticals. 2021;14:1159. doi: 10.3390/ph14111159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Radulescu V., Saviuc C., Chifiriuc C., Oprea E., Ilies D.C., Marutescu L., Lazar V. Chemical composition and antimicrobial activity of essential oil from shoots spruce (Picea abies L.) Rev. Chim. 2011;62:69–74. [Google Scholar]
  • 12.D’Amato S., Serio A., López C.C., Paparella A. Hydrosols: Biological activity and potential as antimicrobials for food applications. Food Control. 2018;86:126–137. doi: 10.1016/j.foodcont.2017.10.030. [DOI] [Google Scholar]
  • 13.Figueiredo A.C., Barroso J.G., Pedro L.G., Scheffer J.J.C. Factors affecting secondary metabolite production in plants: Volatile components and essential oils. Flavour Fragr. J. 2008;23:213–226. doi: 10.1002/ffj.1875. [DOI] [Google Scholar]
  • 14.Nikolić B., Ristić M., Bojović S., Marin P.D. Variability of the Needle Essential Oils of Pinus peuce from Different Populations in Montenegro and Serbia. Chem. Biodivers. 2008;5:1377–1388. doi: 10.1002/cbdv.200890126. [DOI] [PubMed] [Google Scholar]
  • 15.Nikolić B., Ristić M., Tešević V., Marin P.D., Bojović S. Terpene chemodiversity of relict conifers picea omorika, pinus heldreichii, and pinus peuce, endemic to Balkan. Chem. Biodivers. 2011;8:2247–2260. doi: 10.1002/cbdv.201100018. [DOI] [PubMed] [Google Scholar]
  • 16.von Rudloff E. Seasonal variation in the composition of the volatile oil of the leaves, buds, and twigs of white spruce (Picea glauca) Can. J. Bot. 1972;50:1595–1603. doi: 10.1139/b72-195. [DOI] [Google Scholar]
  • 17.Baath M.H., Burzo I. Quantitative and qualitative seasonal variation of volatile oil from 16 conifer species. Biol. Veg. 2009;55:103–110. [Google Scholar]
  • 18.Kartnig T., Still F., Reinthaler F. Antimicrobial activity of the essential oil of young pine shoots (Picea abies L.) J. Ethnopharmacol. 1991;35:155–157. doi: 10.1016/0378-8741(91)90067-N. [DOI] [PubMed] [Google Scholar]
  • 19.Nicolella H.D., Fernandes G., Ozelin S.D., Rinaldi-Neto F., Ribeiro A.B., Furtado R.A., Senedese J.M., Esperandim T.R., Veneziani R.C.S., Tavares D.C. Manool, a diterpene from Salvia officinalis, exerts preventive effects on chromosomal damage and preneoplastic lesions. Mutagenesis. 2021;36:177–185. doi: 10.1093/mutage/geab001. [DOI] [PubMed] [Google Scholar]
  • 20.Nicolella H.D., Ribeiro A.B., de Melo M.R.S., Ozelin S.D., da Silva L.H.D., Sola Veneziani R.C., Crispim Tavares D. Antitumor Effect of Manool in a Murine Melanoma Model. J. Nat. Prod. 2022;85:126–432. doi: 10.1021/acs.jnatprod.1c01128. [DOI] [PubMed] [Google Scholar]
  • 21.Yang H., Zhao R., Chen H., Jia P., Bao L., Tang H. Bornyl acetate has an anti-inflammatory effect in human chondrocytes via induction of IL-11. IUBMB Life. 2014;66:854–859. doi: 10.1002/iub.1338. [DOI] [PubMed] [Google Scholar]
  • 22.Piccinelli A.C., Morato P.N., dos Santos Barbosa M., Croda J., Sampson J., Kong X., Konkiewitz E.C., Ziff E.B., Amaya-Farfan J., Kassuya C.A.L. Limonene reduces hyperalgesia induced by gp120 and cytokines by modulation of IL-1 β and protein expression in spinal cord of mice. Life Sci. 2017;174:28–34. doi: 10.1016/j.lfs.2016.11.017. [DOI] [PubMed] [Google Scholar]
  • 23.Piccinelli A.C., Santos J.A., Konkiewitz E.C., Oesterreich S.A., Formagio A.S.N., Croda J., Ziff E.B., Kassuya C.A.L. Antihyperalgesic and antidepressive actions of (R)-(+)-limonene, α-phellandrene, and essential oil fromSchinus terebinthifoliusfruits in a neuropathic pain model. Nutr. Neurosci. 2015;18:217–224. doi: 10.1179/1476830514Y.0000000119. [DOI] [PubMed] [Google Scholar]
  • 24.Bacanlı M., Anlar H.G., Aydın S., Çal T., Arı N., Ündeğer Bucurgat Ü., Başaran A.A., Başaran N. D-limonene ameliorates diabetes and its complications in streptozotocin-induced diabetic rats. Food Chem. Toxicol. 2017;110:434–442. doi: 10.1016/j.fct.2017.09.020. [DOI] [PubMed] [Google Scholar]
  • 25.Suh K.S., Chon S., Choi E.M. Limonene protects osteoblasts against methylglyoxal-derived adduct formation by regulating glyoxalase, oxidative stress, and mitochondrial function. Chem.-Biol. Interact. 2017;278:15–21. doi: 10.1016/j.cbi.2017.10.001. [DOI] [PubMed] [Google Scholar]
  • 26.Cheng S.-S., Wu C.-L., Chang H.-T., Kao Y.-T., Chang S.-T. Antitermitic and Antifungal Activities of Essential Oil of Calocedrus formosana Leaf and Its Composition. J. Chem. Ecol. 2004;30:1957–1967. doi: 10.1023/B:JOEC.0000045588.67710.74. [DOI] [PubMed] [Google Scholar]
  • 27.Jaganathan S.K., Supriyanto E. Antiproliferative and molecular mechanism of eugenol-induced apoptosis in cancer cells. Molecules. 2012;17:6290–6304. doi: 10.3390/molecules17066290. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Wang W., Wu N., Zu Y.G., Fu Y.J. Antioxidative activity of Rosmarinus officinalis L. essential oil compared to its main components. Food Chem. 2008;108:1019–1022. doi: 10.1016/j.foodchem.2007.11.046. [DOI] [PubMed] [Google Scholar]
  • 29.Guzmán-Gutiérrez S.L., Bonilla-Jaime H., Gómez-Cansino R., Reyes-Chilpa R. Linalool and β-pinene exert their antidepressant-like activity through the monoaminergic pathway. Life Sci. 2015;128:24–29. doi: 10.1016/j.lfs.2015.02.021. [DOI] [PubMed] [Google Scholar]
  • 30.Madhuri K., Naik P.R. Ameliorative effect of borneol, a natural bicyclic monoterpene against hyperglycemia, hyperlipidemia and oxidative stress in streptozotocin-induced diabetic Wistar rats. Biomed. Pharmacother. 2017;96:336–347. doi: 10.1016/j.biopha.2017.09.122. [DOI] [PubMed] [Google Scholar]
  • 31.Jiang J., Shen Y.Y., Li J., Lin Y.H., Luo C.X., Zhu D.Y. (+)-Borneol alleviates mechanical hyperalgesia in models of chronic inflammatory and neuropathic pain in mice. Eur. J. Pharmacol. 2015;757:53–58. doi: 10.1016/j.ejphar.2015.03.056. [DOI] [PubMed] [Google Scholar]
  • 32.Girola N., Figueiredo C.R., Farias C.F., Azevedo R.A., Ferreira A.K., Teixeira S.F., Capello T.M., Martins E.G.A., Matsuo A.L., Travassos L.R., et al. Camphene isolated from essential oil of Piper cernuum (Piperaceae) induces intrinsic apoptosis in melanoma cells and displays antitumor activity in vivo. Biochem. Biophys. Res. Commun. 2015;467:928–934. doi: 10.1016/j.bbrc.2015.10.041. [DOI] [PubMed] [Google Scholar]
  • 33.Tiwari M., Kakkar P. Plant derived antioxidants—Geraniol and camphene protect rat alveolar macrophages against t-BHP induced oxidative stress. Toxicol. Vitr. 2009;23:295–301. doi: 10.1016/j.tiv.2008.12.014. [DOI] [PubMed] [Google Scholar]
  • 34.Vallianou I., Peroulis N., Pantazis P., Hadzopoulou-Cladaras M. Camphene, a Plant-Derived Monoterpene, Reduces Plasma Cholesterol and Triglycerides in Hyperlipidemic Rats Independently of HMG-CoA Reductase Activity. PLoS ONE. 2011;6:e20516. doi: 10.1371/journal.pone.0020516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Kroumova A.B., Artiouchine I., Wagner G.J. Use of several natural products from selected nicotiana species to prevent black shank disease in Tobacco. Contrib. Tob. Nicotine Res. 2016;27:113–125. doi: 10.1515/cttr-2016-0013. [DOI] [Google Scholar]
  • 36.Bouzenna H., Hfaiedh N., Giroux-Metges M.-A., Elfeki A., Talarmin H. Potential protective effects of alpha-pinene against cytotoxicity caused by aspirin in the IEC-6 cells. Biomed. Pharmacother. 2017;93:961–968. doi: 10.1016/j.biopha.2017.06.031. [DOI] [PubMed] [Google Scholar]
  • 37.Zhou J.Y., Tang F.D., Mao G.G., Bian R.L. Effect of α-pinene on nuclear translocation of NF-κB in THP-1 cells. Acta Pharmacol. Sin. 2004;25:480–484. [PubMed] [Google Scholar]
  • 38.de Almeida Pinheiro M., Magalhães R.M., Torres D.M., Cavalcante R.C., Mota F.S.X., Coelho E.M.A.O., Moreira H.P., Lima G.C., da Costa Araújo P.C., Cardoso J.H.L., et al. Gastroprotective effect of alpha-pinene and its correlation with antiulcerogenic activity of essential oils obtained from Hyptis species. Pharmacogn. Mag. 2015;11:123–130. doi: 10.4103/0973-1296.149725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Him A., Ozbek H., Turel I., Oner A.C. Antinociceptive activity of alpha-pinene and fenchone. Pharmacologyonline. 2008;3:363–369. [Google Scholar]
  • 40.Donati M., Mondin A., Chen Z., Miranda F.M., do Nascimento B.B., Jr., Schirato G., Pastore P., Froldi G. Radical scavenging and antimicrobial activities of Croton zehntneri, Pterodon emarginatus and Schinopsis brasiliensis essential oils and their major constituents: Estragole, trans-anethole, β-caryophyllene and myrcene. Nat. Prod. Res. 2015;29:939–946. doi: 10.1080/14786419.2014.964709. [DOI] [PubMed] [Google Scholar]
  • 41.Rufino A.T., Ribeiro M., Sousa C., Judas F., Salgueiro L., Cavaleiro C., Mendes A.F. Evaluation of the anti-inflammatory, anti-catabolic and pro-anabolic effects of E-caryophyllene, myrcene and limonene in a cell model of osteoarthritis. Eur. J. Pharmacol. 2015;750:141–150. doi: 10.1016/j.ejphar.2015.01.018. [DOI] [PubMed] [Google Scholar]
  • 42.Zhang Z., Yang C., Dai X., Ao Y., Li Y. Inhibitory effect of trans-caryophyllene (TC) on leukocyte-endothelial attachment. Toxicol. Appl. Pharmacol. 2017;329:326–333. doi: 10.1016/j.taap.2017.06.016. [DOI] [PubMed] [Google Scholar]
  • 43.de Oliveira C.C., de Oliveira C.V., Grigoletto J., Ribeiro L.R., Funck V.R., Grauncke A.C.B., de Souza T.L., Souto N.S., Furian A.F., Menezes I.R.A., et al. Anticonvulsant activity of β-caryophyllene against pentylenetetrazol-induced seizures. Epilepsy Behav. 2016;56:26–31. doi: 10.1016/j.yebeh.2015.12.040. [DOI] [PubMed] [Google Scholar]
  • 44.Paula-Freire L.I.G., Andersen M.L., Gama V.S., Molska G.R., Carlini E.L.A. The oral administration of trans-caryophyllene attenuates acute and chronic pain in mice. Phytomedicine. 2014;21:356–362. doi: 10.1016/j.phymed.2013.08.006. [DOI] [PubMed] [Google Scholar]
  • 45.Hui L.-M., Zhao G.-D., Zhao J.-J. δ-Cadinene inhibits the growth of ovarian cancer cells via caspase-dependent apoptosis and cell cycle arrest. Int. J. Clin. Exp. Pathol. 2015;8:6046–6056. [PMC free article] [PubMed] [Google Scholar]
  • 46.Guo X., Shang X., Li B., Zhou X.Z., Wen H., Zhang J. Acaricidal activities of the essential oil from Rhododendron nivale Hook. f. and its main compund, δ-cadinene against Psoroptes cuniculi. Vet. Parasitol. 2017;236:51–54. doi: 10.1016/j.vetpar.2017.01.028. [DOI] [PubMed] [Google Scholar]
  • 47.Rogerio A.P., Andrade E.L., Leite D.F.P., Figueiredo C.P., Calixto J.B. Preventive and therapeutic anti-inflammatory properties of the sesquiterpene α-humulene in experimental airways allergic inflammation. Br. J. Pharmacol. 2009;158:1074–1087. doi: 10.1111/j.1476-5381.2009.00177.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 48.Chainy G.B.N., Manna S.K., Chaturvedi M.M., Aggarwal B.B. Anethole blocks both early and late cellular responses transduced by tumor necrosis factor: Effect on NF-κB, AP-1, JNK, MAPKK and apoptosis. Oncogene. 2000;19:2943–2950. doi: 10.1038/sj.onc.1203614. [DOI] [PubMed] [Google Scholar]
  • 49.Moghimi R., Aliahmadi A., Rafati H. Ultrasonic nanoemulsification of food grade trans-cinnamaldehyde: 1,8-Cineol and investigation of the mechanism of antibacterial activity. Ultrason. Sonochem. 2017;35:415–421. doi: 10.1016/j.ultsonch.2016.10.020. [DOI] [PubMed] [Google Scholar]
  • 50.Juergens U.R., Engelen T., Racké K., Stöber M., Gillissen A., Vetter H. Inhibitory activity of 1,8-cineol (eucalyptol) on cytokine production in cultured human lymphocytes and monocytes. Pulm. Pharmacol. Ther. 2004;17:281–287. doi: 10.1016/j.pupt.2004.06.002. [DOI] [PubMed] [Google Scholar]
  • 51.Kennedy-Feitosa E., Okuro R.T., Pinho Ribeiro V., Lanzetti M., Barroso M.V., Zin W.A., Porto L.C., Brito-Gitirana L., Valenca S.S. Eucalyptol attenuates cigarette smoke-induced acute lung inflammation and oxidative stress in the mouse. Pulm. Pharmacol. Ther. 2016;41:11–18. doi: 10.1016/j.pupt.2016.09.004. [DOI] [PubMed] [Google Scholar]
  • 52.The International Fragrance Association . The Complete IFRA Standards—50th Amendment. The International Fragrance Association; Geneva, Switzerland: 2022. [Google Scholar]
  • 53.European Commission Regulation No 544/2009 of the European Parliament and of the Council. Off. J. Eur. Union. 2009;284:1. [Google Scholar]
  • 54.Karlberg A.-T., Lepoittevin J.-P. One hundred years of allergic contact dermatitis due to oxidized terpenes: What we can learn from old research on turpentine allergy. Contact Dermat. 2021;85:627–636. doi: 10.1111/cod.13962. [DOI] [PubMed] [Google Scholar]
  • 55.Ladan M.K., Glavač N.K. GC–MS Analysis of A Helichrysum italicum Hydrosol: Sensitivity, Repeatability and Reliability of Solvent Extraction versus Direct Hydrosol Analysis. Appl. Sci. 2022;12:10040. doi: 10.3390/app121910040. [DOI] [Google Scholar]
  • 56.Garneau F.-X., Collin G., Gagnon H., Pichette A. Chemical Composition of the Hydrosol and the Essential Oil of Three Different Species of the Pinaceae Family: Picea glauca (Moench) Voss., Picea mariana (Mill.) B.S.P., and Abies balsamea (L.) Mill. J. Essent. Oil Bear. Plants. 2012;15:227–236. doi: 10.1080/0972060X.2012.10644040. [DOI] [Google Scholar]
  • 57.Francezon N., Stevanovic T. Chemical composition of essential oil and hydrosol from Picea mariana bark residue. BioResources. 2017;12:2635–2645. doi: 10.15376/biores.12.2.2635-2645. [DOI] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

Not applicable.


Articles from Plants are provided here courtesy of Multidisciplinary Digital Publishing Institute (MDPI)

RESOURCES