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Article

Comparative Analysis of the Fatty Acid Profiles of Selected Representatives of Chlorella-Clade to Evaluate Their Biotechnological Potential

by
Elena Krivina
1,
Evgeny Degtyaryov
2,
Elizaveta Tebina
2,
Anna Temraleeva
1 and
Tatyana Savchenko
2,*
1
All-Russian Collection of Microorganisms (VKM), G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms, Pushchino Scientific Center for Biological Research, Russian Academy of Sciences, 142290 Pushchino, Russia
2
Institute of Basic Biological Problems, Pushchino Scientific Center for Biological Research, Russian Academy of Sciences, 142290 Pushchino, Russia
*
Author to whom correspondence should be addressed.
Int. J. Plant Biol. 2024, 15(3), 837-854; https://doi.org/10.3390/ijpb15030060
Submission received: 16 July 2024 / Revised: 5 August 2024 / Accepted: 19 August 2024 / Published: 21 August 2024
(This article belongs to the Special Issue Microalgae as a Powerful Tool for Biopharming Development)
Browse Figures
Versions Notes

Abstract

:
The objective of this study was to analyze the fatty acid composition of five strains from the genera Chlorella, Micractinium, and Meyerella and conduct an initial assessment of their biotechnological potential. It was found that the strain C. vulgaris VKM Al-335 is a good producer of palmitic acid, the Micractinium strains VKM Al-332 and VKM Al-343 are rich in ω-3 fatty acids, whereas the Meyerella strains VKM Al-346 and VKM Al-428 are producers of ω-6 fatty acids. A comparison of the biotechnological potential of algae with that of higher plant leaves (wheat) demonstrates that algal fatty acids exhibit greater diversity, although it is inferior to wheat leaves in terms of polyunsaturated and ω-3 fatty acids. Correlation analysis showed that when only straight-chain fatty acids were considered, the strains were distributed on the principal component analysis plot in accordance with their genetic relationships. However, when the entire fatty acid profile, inclusive of minor branched-chain and cyclic fatty acids, was analyzed, the algae distribution was in accordance with the environmental conditions in the original habitat, suggesting a possible connection between branched-chain and cyclic fatty acids and microalgae adaptability to environmental temperature conditions.

1. Introduction

Due to the increasing scarcity of natural resources and the environmental crisis, there is a growing interest in using new biological resources for producing environmentally friendly products and biofuels. Currently, algae are receiving more and more attention from biotechnology researchers. They are being considered as a raw material for the production of third-generation biofuels [1,2] and wastewater treatment [3]. Some consortia of algae are used for oil biodegradation and detoxification. A promising area of agricultural biotechnology is the use of algae metabolites to produce biological products that can stimulate plant metabolism, accelerate growth, and increase resistance to insect pests and phytopathogens [4,5,6]. Algae are used for the production of food, feed, dietary supplements, pharmaceutical and cosmetic products. Of particular interest are algae strains that produce valuable polyunsaturated fatty acids (PUFAs) and oxylipins [4,5].
The mass cultivation of algae offers several advantages over the cultivation of traditional crops. Selected strains can achieve amazingly high growth rates due to a very short doubling period (1–2 days) [7]. Algae can grow in a wide range of environments, including brackish, marine or hypersaline water bodies, and wastewater. This means that they do not compete with food crops for arable land. Importantly, algae have a much higher species diversity than terrestrial plants. This implies a greater variety of metabolites that can be useful for biotechnology [4]. To date, more than 2000 secondary metabolites produced by algae have been identified [8]. At the same time, due to the high plasticity of algal metabolism, the biochemical composition of their biomass can be adjusted to meet the specific needs of the application through appropriate cultivation techniques. An important biotechnological characteristic of several algae is the ability to grow autotrophically, heterotrophically, or mixotrophically, which significantly expands the possibilities of their cultivation and allows the modification of the metabolic profile by changing the growth conditions or method of cultivation [9,10]. Several bioactive compounds, such as the phytohormone methyl jasmonate, can be also used as effective modulators of the fatty acid composition of microalgae cells [11]. However, the main challenge to widespread industrial production of algae is that it is more expensive than traditional feedstocks. One of the possible ways to solve this problem is to search for new effective producers of valuable metabolites.
The so-called “small green balls” from the Chlorella-clade have been very actively used in biotechnology for many decades both as model objects and as raw materials for the production of various food products, dietary supplements, fertilizers, etc. According to various estimates, between 2000 and 5000 tons of biomass are produced annually from Chlorella-like algae, and experts predict that production volumes will continue to increase in the future, since a number of representatives of this group are characterized by the ability to quickly accumulate biomass with a relatively high content of valuable metabolites, including fatty acids (FAs), and are not demanding to cultivation conditions [1,4,5,12]. In this regard, the study of the biotechnological potential of various representatives of the Chlorella-clade has not only scientific but also practical significance.
In order to consider algae as an alternative to agricultural plants and to develop a strategy to unlock the potential of algae in general, and Chlorella representatives in particular, we compared their fatty acid profiles with traditional raw material sources, including traditional crops. For example, wheat is one of the most common agricultural crops. It is grown on a larger area than any other food crop (220.7 million hectares or 545 million acres in 2021) [13]. Furthermore, wheatgrass is beginning to be widely used as a dietary supplement to improve overall well-being, activate the immune system, enhance tissue regeneration, etc. [13,14].
Accurate identification and quantification of FAs and their derivatives is a prerequisite for appropriate strain selection. A significant obstacle is a substantial discrepancy in the data on the content of individual FAs in microalgae, which can be explained by the real variability of the FA profile associated with differences in algal growth conditions, physiological age of culture [9,10], or arise from differences in methods of FA extraction and analysis. The most reliable and comparable results are obtained by analyzing organisms grown under similar conditions using the same analytical methods.
This study presents the results of a comparative analysis of the major and minor fatty acids and oxylipins in representatives of the three different genera from the Chlorella-clade, as well as an attempt to assess their biotechnological potential. It is worth emphasizing that several analyzed strains are representatives of new, recently described species, and the presented study is the first comprehensive estimation of their biotechnological value. To reveal the studied algae potential as an alternative to using agricultural plants, we compared their fatty acid profiles with the fatty acid profile of wheat leaves, which are one of the dominant cereal cultures in many countries and are actively used as a dietary supplement and «superfood».

2. Materials and Methods

2.1. Isolation and Cultivation of Algal Strains

The strain VKM Al-335 (=ACSSI 335 (Algal Collection of Soil Science Institute, Russian Federation), =IPPAS C-21 (Collection of Microalgae and Cyanobacteria IPPAS of the K.A. Timiryazev Institute of Plant Physiology Russian Academy of Sciences (Moscow, Russian Federation), =DMMSU-4 (Department of Microbiology of Moscow State University (Moscow, Russian Federation)) was isolated from an unknown freshwater lake, in Japan in 1960. The authentic strain Micractinium thermotolerans VKM Al-332 (=ACCSI 332, =IPPAS C-16, =LARG-3 (Laboratory of Radiation Genetics, Russian Federation)) was found in a hot spring located on the Chukotka Peninsula (Russian Federation) in 1967. The two authentic strains Micractinium lacustre VKM Al-343 (=ACSSI 343) and Meyerella similis VKM Al-346 (=ACSSI 346) were isolated from small urban reservoirs of the Vasilyevsky Lakes system (Tolyatti, Samara region, Russian Federation) in 2019. The authentic strain Meyerella krienitzii VKM Al-428 (=ACSSI 428) was found in tundra soils of Pleistocene Park (Sakha Republic, Russian Federation) in 2020. All cultures were grown on solid BG-11 medium (1% agar) at 23–25 °C with a 12:12 h light: dark cycle, at a light intensity of 60–75 μmol photons·m–2·s–1 provided by cool white, fluorescent lamps.

2.2. Microscopy

The morphological characteristics and life cycle of the strains were studied using light microscopy with a Leica DM750 microscope (Leica Mikrosysteme Vertrieb GmbH, Wetzlar, Germany). Photos were taken with a color digital camera Leica Flexacam C3 (Leica Mikrosysteme Vertrieb GmbH, Wetzlar, Germany). The studied strains were observed for a period of 2 weeks to 6 months. Leica Application Suite X software (Version 3.7.5) was utilized for morphometric measurements. Microscopy of samples and measurement of cell sizes were carried out according to standard protocols [15]. At least 100 cells of each strain were analyzed for size measurements.

2.3. DNA Isolation, Amplification, Purification, and Sequencing

DNA isolation, amplification, purification, and sequencing were carried out according to standard methods described in detail in the articles [16,17].

2.4. Methods of Phylogenetic Analysis

Phylogenetic analysis was performed on a concatenated dataset of the 18S–ITS1–ITS2 sequences. The closely related sequences were searched using the BLASTn algorithm in the NCBI GenBank [18]. All sequences met the following requirements: reading quality (without degenerate and unknown nucleotides), reading length (sequences including 18S–ITS1–ITS2 and measuring at least 2200 bp), and maximum affinity (similarity ≥ 95%). The dataset included sequences from a collection of authentic strains. Thus, the dataset contained 51 sequences of representatives of Trebouxiophyceae and the authentic strain Edaphochlorella mirabilis SAG 38.88 as the outgroup. If there were introns in the 18S rRNA gene, they were removed from the alignment. Taxon names are listed according to the international electronic database AlgaeBase [19]. 18S rRNA gene alignment was performed separately using MAFFT software ver. 7 (Q-INS-i method), alignment of ITS1 and ITS2 was performed taking into account their secondary structure in the 4SALE program [20]. Based on the minimum values of the Bayesian information criterion (BIC), the TNe + I + G4 nucleotide substitution model was selected as the optimal model. Further phylogenetic analysis was carried out using the methods described in [21].

2.5. Fatty Acid Extraction and Analysis

Extraction of fatty acids from algae samples or frozen wheat leaf tissue was carried out by the method of Bligh and Dyer [22]. Fatty acid methyl esters were obtained by incubating the extracted fatty acids in 1 mL of 8% sulfuric acid in methanol. The samples were incubated at 90 °C for 90 min and then cooled to room temperature. One mL of 10% sodium chloride in water and 250 μL of hexane were added, and after 10 min, the upper phase containing fatty acid methyl esters was transferred to a vial and analyzed by gas chromatography-mass spectrometry (GC-MS). The conditions for GC-MS analysis have been described in detail previously [16,17]. Identification of the fatty acids was carried out by comparing their mass spectra with fatty acid standards Supelco 37 Component FAME Mix (Supelco, Bellefonte, PA, USA) and using the NIST’20 Mass Spectral Library (National Institute of Standards and Technology, Gaithersburg, MD, USA).
The unsaturation index was calculated according to the equation: UI = 0.01× [(mol % of monoenoic FA) + (2× mol % of dienoic FA) + (3× mol % of trienoic) + (4× mol % tetraenoic FA)].

2.6. Statistical Analysis

All experiments were performed in three biological replicas. To determine the statistically significant difference between species, a one-way analysis of variance (ANOVA) was performed. This was followed by the Tukey post hoc test (for multiple comparisons) when significant differences (p ≤ 0.05) were found. Different letters and stars on graphs indicate statistically significant differences.
Principal Component Analysis (PCA) Calculator (Statistics_calculators 2017) available from https://www.statskingdom.com/pca-calculator.html (accessed on 30 May 2024) was used to build a PCA plot. Pearson’s correlation coefficient was calculated to confirm significant correlations between the content of various fatty acids in algae samples.

3. Results

3.1. Morphological Observations by Light Microscopy

All the studied strains had a morphology that is typical of the so-called “small green balls” (Figure 1) [16,17]. The cells were solitary and did not produce bristles. They were predominantly spherical or broadly oval in shape. The smallest cell size was observed in the strain Mr. similis VKM Al-346, which was no more than 3 µm in diameter. The cell sizes of strains Chlorella vulgaris VKM Al-335, Mr. krienitzii VKM Al-428, and Mc. thermotolerans VKM Al-332 were larger but did not exceed 7 µm. The strain Mc. lacustre VKM Al-343 had the largest cell size among the studied strains, up to 10.5 µm. The chloroplasts in all strains were parietal, primarily cup-shaped. The pyrenoid was the only one in the strains C. vulgaris VKM Al-335, Mc. thermotolerans VKM Al-332, Mc. lacustre VKM Al-343, whereas it was expectedly absent in the strains Mr. similis VKM Al-346, Mr. krienitzii VKM Al-428. Reproduction was by autospores.

3.2. Phylogenetic Analysis

The topology of the Chlorella-clade when using a fragment 18S–ITS1–ITS2 that takes into account the secondary structure of each marker coincides in general with the results of the analysis performed using 18S–ITS1–ITS2 without taking into account the secondary structure of gene 18S rRNA and variable spacers ITS1 and ITS2 [16,17]. All the studied strains were representatives of the Chlorella-clade (Figure 2). The strain VKM Al-335 was clustered with an authentic strain C. vulgaris SAG 211-11b (SH-aLRT—100%, BP—100%, PP—1.00). The genetic distances were less than 0.1% and corresponded to the intraspecific level in the Chlorella-clade. For comparison, the lowest levels of interspecific variation were observed between species Mc. variable and Mc. singularis, and they amounted to no more than 0.4%. CBCs and differences in the secondary structure of ITS1 and ITS2 were absent between the studied strain and the authentic strain SAG 211-11b. Thus, the studied strain was identified as C. vulgaris.
In the present study, phylogenetic positions of strains Mc. lacustre VKM Al-343 and Mc. thermotolerans VKM Al-332 were consistent with the results of a previous phylogenetic analysis [16], which was conducted using the fragment 18S–ITS1–5.8S–ITS2 sequence, but without considering the secondary structure of gene 18S rRNA and variable spacers ITS1 and ITS2. The strain VKM Al-343 also was clustered with Chlorella-like species Mc. inermum, Mc. simplicissimum, Mc. singularis, and also Mc. variabile, which sometimes shows the classic Micractinium-like morphotype (SH-aLRT—100%, BP—100%, PP—1.00). In this case, the genetic distances between strain VKM Al-343 and strains of sister species were 0.5–0.7% (interspecific level). The strain Mc. thermotolerans VKM Al-332 was clustered with the sister strain Mc. tetrahymenae SAG 2587 (SH-aLRT—100%, BP—100%, PP—1.00). The genetic distance between them was 0.7%, which corresponded to the interspecific level. The strains Mr. similis VKM Al-346 and Mr. krienitzii VKM Al-428 were representatives of the genus Meyerella (SH-aLRT—100%, BP—100%, PP—1.00). Their phylogenetic positions are also consistent with a previous study [17]. The genetic distances between representatives of three different species within the genus were 1.6–2.5%, which corresponds to the interspecific level.

3.3. Fatty Acid Composition of the Studied Organisms

In Figure 3, the results of the fatty acid analyses of five studied strains of three different genera from Chlorella-clade are presented.
The figure shows that the fatty acid composition of the species examined varies noticeably. The largest differences are observed between members of different genera. The smallest differences in the profile are found in representatives of the genus Meyerella, and the fatty acid profiles of Mc. thermotolerans VKM Al-332 and Mc. lacustre VKM Al-343 are also similar in many respects. One striking characteristic of the fatty acid composition of C. vulgaris is the high content of saturated hexadecanoic fatty acid. Also noticeable is the presence of polyunsaturated hexadecatetraenoic acid (16:4Δ4,7,10,13) in Mc. lacustre VKM Al-343. A prominent characteristic of the fatty acid composition of Meyerella representatives is the dominance of octadecadienoic acid (18:2Δ9,12). The relative content of α-linolenic acid in Meyerella strains is significantly lower than in C. vulgaris VKM Al-335, while in Micractinium strains it is significantly higher. In general, the representatives of the genus Meyerella have more unsaturated fatty acids with two double bonds, both 16-carbon and 18-carbon fatty acids. Also of interest is the presence of γ-linolenic acid (18:3Δ6.9,12) in strain C. vulgaris VKM Al-335 and Micractinium strains.
For comparison, we investigated the fatty acid profile of leaves of hexaploid wheat Triticum aestivum L., variety Saratovskaya-60. The fatty acid profile of the leaves of hexaploid wheat is noticeably less diverse and represented by four fatty acids, 16:0 (18.52 ± 4.05%), 16:1Δ7 (6.07 ± 1.13%), 18:2Δ9,12 (7.46 ± 0.93%), and 18:3Δ9,12,15 (67.75 ± 1.17%). Based on the generated data, several qualitative indexes were calculated for the microalgae fatty acid profiles and compared with the indexes of the wheat leaf samples (Table 1).
Strains Mr. krienitzii VKM Al-428 and Mc. lacustre VKM Al-343 contain the highest amount of polyunsaturated fatty acids, with Mr. krienitzii VKM Al-428 PUFA being mainly represented by 18-carbon, and Mc. lacustre VKM Al-343 by 16-carbon. UI is highest in Mc. lacustre VKM Al-343; this alga is also characterized by the highest ratio ω-3/ω-6, which is an important characteristic of dietary fats. Interestingly, wheat leaves are markedly more abundant in both PUFA and ω-3 FA content than the studied algae, mainly because of the very high content of α-linolenic acid. However, the total UI is still lower in wheat leaves than in the strain VKM Al-343.
In addition to the major FA described, several minor fatty acids were detected in the algal samples including straight-chain, branched-chain, and cyclic fatty acids (Table 2). The content of each minor FA does not exceed 3% of total fatty acids, and they are not presented on the graph in Figure 3. Because of the more complex structure, the structural formulae of the cyclic FAs are presented in Table 2.
All algae contain tetradecanoic acid: in strain C. vulgaris VKM Al-335 and Micractinium strains, this FA content is about 1%; in Meyerella strains, the amount of this FA is much lower. Odd-numbered chain pentadecanoic acid was also found in Micractinium strains. Algae samples except Mc. thermotolerans VKM Al-332 also contain branched-chain fatty acids (BCFAs) with chain lengths from 14 to 18 carbons; three of these FAs (15-methyl-hexadecanoic acid, 16-methyl-heptadecanoic acid (iso-stearic), and 17-methyl-octadecanoic acid) are iso-fatty acids, containing the branch point on the penultimate carbon atom, including one FA with an odd-numbered chain (16-methyl-heptadecanoic acid). BCFAs are particularly numerous in Mr. similis VKM Al-346 and Mr. krienitzii VKM Al-428. Some cyclic fatty acids were also found in studied samples, but it is important to remember that they can be formed by modification of polyunsaturated fatty acids during extraction. Double bonds of unsaturated fatty acids can undergo changes during extraction and esterification procedures and form cyclic groups. The identification of these minor fatty acids relies only on Mass Spectral Library predictions, as branched-chain and cyclic FA standards were not used in the study. Although the content of minor fatty acids is not large, they can fulfill important biological functions. The presence of side branching and cyclic groups changes the physicochemical properties of FAs significantly (Supplementary Table S1).
It has been previously demonstrated that the data on fatty acid content can be considered compositional data, and statistical tools for compositional data analysis can be applied to study the fatty acid profiles [23]. We have used fatty acid content data obtained for the five microalgae to construct the Covariance matrix (Table 3) and then visualized the results using a Principal Component Analysis (PCA) biplot (Figure 4). The analysis of the content of straight-chain fatty acids revealed significant correlations.
The visualization of the data by PCA plot can provide deeper insight into data and reveals the major factor shaping the fatty acid profile (genetic relationship or natural habitat). We generated two PCA plots, using only straight-chain FAs in one case (Figure 4a) and all major and minor FAs (Figure 4b). The first two principal components in the first plot explained more than 75% of the total variance (51.02% by PC1 and 24% by PC2) in the first case, and 72.5%-(46.66% by PC1 and 25.8% by PC2) in the second case. The first PCA plot shows that based on the fatty acid profile, different genera are distributed in different quarters of the plot, with representatives of the genus Meyerella being closest to each other. The second PCA plot shows that two strains of different Meyerella species are still closely spaced on the plot, but Mc. thermotolerance VKM Al-332 and Mc. lacustre VKM Al-346 are separated, with Mc. lacustre VKM Al-346 spaced closer to C. vulgaris VKM Al-335.
Besides FAs, several fatty acid derivatives and related compounds were identified in studied microalgae samples, including three oxylipins: cis-9,10-epoxyoctadecanoic acid (or oxiraneoctanoic acid) and 7-methyl-Z-tetradecen-1-ol acetate found in C. vulgaris VKM Al-335, and hydroxy-derivative of dodecanoic fatty acid (3-hydroxy-dodecanoic acid) present in both Meyerella representatives. Structural formulae of identified oxylipins are presented in Table 4. These oxylipins were not found in Micractinium strains.
In addition, all studied samples contained phthalic acid derivatives, such as methyl tetradecyl ester of phthalic acid, which can be biosynthesized by living organisms, including algae, or can be introduced into the samples from the external environment since these compounds are currently extensively produced and utilized [24].

4. Discussion

Despite the long history of research, members of the Chlorella-clade continue to be of interest, both from a strictly scientific perspective and from a biotechnological point of view [2,4,9,16,17,25]. In this study, a detailed analysis was conducted on the fatty acid profiles of representatives of three genera within Chlorella-clade: Chlorella (C. vulgaris VKM Al-335), a recently described species of the genus Micractinium (Mc. lacustre VKM Al-343, Mc. thermotolerans VKM Al-332) [13], and Meyerella (Mr. similis VKM Al-346, Mr. krienitzii VKM Al-428) [17]. Since sparse data on the FA composition of cells of selected representatives of the Chlorella-clade have been reported [2,16,17], this study, by comparing them to each other and with leaves of higher plants, expands the understanding of the biotechnological potential of these organisms. As a result of the simultaneous analysis of fatty acids and their derivatives in representatives of three different genera and five different species of “small green balls”, significant differences were revealed both in the relative content of fatty acids with different carbon chain lengths and the degree of unsaturation (Table 1). In C. vulgaris VKM Al-335 and both Micractinium strains, sixteen-carbon chain FAs represent the majority of FAs, with a maximum value of 62.3% in the strain Mc. lacustre VKM Al-343, whereas in Meyerella strains the majority of FAs is represented by eighteen-carbon FAs. The highest content of unsaturated FAs, including PUFAs, is found in Mr. krienitzii VKM Al-428 (more than 80%) and the lowest in C. vulgaris VKM Al-335 (less than 50%). A very high content of unsaturated FAs is also found in Mc. lacustre VKM Al-343, whose FA profile is characterized by the highest unsaturation index (2.39) and the highest content of ω-3 FAs. These characteristics are largely determined by the presence of high levels of hexadecatetraenoic acid. The high content of unsaturated FAs and the high value of the unsaturation index found in Mc. thermotolerans VKM Al-332 growing under conditions of elevated temperatures are unexpected. It is known that a high content of unsaturated fatty acids leads to an increased fluidity of lipid membranes, which is an advantage for organisms growing at low temperatures [26].
Among all the strains studied, the strain C. vulgaris VKM Al-335 was the most active producer of saturated palmitic acid (up to 44.3%). Such an intensive accumulation of this particular fatty acid, as a rule, is not typical for representatives of this species (on average they accumulate not more than 24%) [27,28,29], although the very high content of palmitic acid has been previously detected in this alga [30]. Within the Chlorella-clade, such a high content of palmitic acid in strains with the confirmed taxonomic status was previously reported for the strain Hegewaldia parvula SAG 7.93 (46.4%) [27]. A high level of palmitic acid was also found in the strain of Neochlorella semenenkoi IPPAS C-1210 (35.9%) [28], but it is still lower than that of the studied strain. Palmitic acid is used in the production of soap, cosmetics, industrial lubricants, and in the food industry [31]. The strain C. vulgaris VKM Al-335, even when cultivated under standard conditions, is only slightly inferior in the content of this acid to the leaders such as palm oil (45.1%) and karuka nut (44.9%), and significantly superior to cotton oil (24.7%), corn (12.2%), soybean (11%) [32], and wheat Triticum aestivum (in this study). In this strain, α-linolenic acid is a predominant unsaturated FA (22.7%), but it is not the maximum detected within the species C. vulgaris (16.1–39.1%) [27,28,29,30,31].
Both studied Micractinium strains were rich in valuable ω-3 α-linolenic acid, representing about a third of the total FAs, although they are not super-producers. This FA, a component of cell membranes and blood vessels, is not synthesized in sufficient amounts in the human body and is one of the necessary components of a healthy diet. In medicine, it is used as part of parenteral nutrition products, dietary supplements, dermatoprotective, and lipid-lowering drugs [4,27]. The ω-3/ω-6 ratio is very high in the strain Mc. lacustre VKM Al-343 (6.44). In the Chlorella-clade, this coefficient is only higher in the strain Mc. variabile KNUA034 (9.69) [4,13,25,33,34]. Such a high relative abundance of ω-3 FAs combined with a very high total unsaturated fatty acid content (almost 80 mol % of total FA) makes this strain a very valuable resource for PUFA production. A high ω-3/ω-6 ratio in the diet is very effective at reducing the risk of several chronic diseases [34]. In addition, these algae accumulate 7,10,13-hexadecatrienoic acid (Mc. thermotolerans VKM Al-332–21.6%, Mc. lacustre VKM Al-343—16%). Comparable levels of 7,10,13-hexadecatrienoic acid were found in another Chlorella-clade strain, Lewiniosphaera symbiontica SAG 211-40a (18%). Strain Mc. lacustre VKM Al-343 is able to produce significant amounts of 4,7,10,13-hexadecatetraenoic acid (19%). Hexadecatetraenoic acid is quite rare within this clade. The considerable quantity, which is greater than that of the studied strains, has only been identified in the strain Didymogenes palatina SAG 30.92 (24.7%) [27]. In general, however, this fatty acid is more characteristic of some representatives of the genera Undaria, Ulva [35,36], and Monoraphidium [37]. This FA suppresses the production of eicosanoids involved in various pathological immune responses [35]. It is also believed that this fatty acid may affect the development of mullet embryos and juveniles. Therefore, these algae could be considered as a potential biological supplement for fish farms [38].
As mentioned above, the biotechnological potential of the studied Meyerella strains is associated with eighteen-carbon FAs. Unlike other studied strains, they are promising producers of linoleic acid (Mr. similis VKM Al-346–26.5%, Mr. krienitzii VKM Al-428–39.1%). This is particularly evident in the case of strain VKM Al-428, which is the most efficient producer of this FA, not only among the strains under study but also within the Chlorella-clade. Previously, the highest values were observed in the strain Micractinium sp. KNUA032 (34.7%), which, like our strain, was also isolated from regions with harsh living conditions (low temperatures, specific conditions of solar radiation) [2,25,27]. α-Linoleic acid is one of the two essential fatty acids for humans, which should be obtained from food [39]. It is a component of quick-drying oils used in oil paints and varnishes. It is also used to reduce the risk of cardiovascular diseases, diabetes, and premature death [40,41]. γ-Linolenic acid has been found in C. vulgaris VKM Al-335 and both Micractinium strains. γ-Linolenic acid possesses anti-inflammatory properties. The total market for oils containing this FA is about USD 60–70 million per year [42], but sources are very limited [43].
For a true assessment of biotechnological potential, the content of valuable metabolites must be compared not only between closely related species but also with representatives of distant groups. In this study, we used hexaploid wheat as an example of a distant photosynthetic organism. A comparison of the FA profile of the microalgae strains studied and wheat leaves revealed unexpected results (Table 1): the lipids of wheat leaves exceed all studied strains in the content of PUFA and ω-3 FAs, with three-quarters of the FAs represented by 18-carbon chain FAs, mainly α-linolenic acid. This information is useful for understanding the biology of various photosynthetic organisms and should be regarded as a rationale for considering the biotechnological potential of leaves of higher plants for fatty acid production. It should be noted, however, that the use of higher plants for the production of fatty acids may be impractical for a number of reasons: major agricultural areas are used for the production of essential food, obtaining biomass requires long cultivation periods, and only seeds are thought to contain FAs in sufficient extractable quantities. Microalgae, which can be grown on marginal soils, in saline waters, and in compact bioreactors, are capable of accumulating considerable amounts of lipids, and under certain conditions, some species can produce up to 80% oil on a dry weight basis. There are strong indications that oil productivity of many microalgae exceeds that of better oil crops [13,44]. On the other hand, there is growing evidence that the quantitative production of microalgae oil is often overestimated [45]. Further accurate assessment of the biotechnological potential of different biological species is needed to identify the best and cheapest source of lipids and individual fatty acids.
It thus appears that with regard to major straight-chain FAs, the optimal sources remain a topic of debate. However, with regard to minor/unusual FAs, microalgae are unquestionably an excellent source. Importantly, the fatty acid composition of algae studied is characterized by greater diversity compared to wheat leaves. An important factor driving interest in algae is the presence of rare and unusual fatty acids and fatty acid derivatives that can be used as food, food additives, feed, and pharmaceuticals [46,47]. Rare fatty acids are of significant interest due to their notable biological activity, which makes them suitable for use as therapeutic compounds and as protectants against plant pathogens and pests [48]. Of particular interest is the identification of BCFAs in studied algae. Five BCFAs were detected; all of them are saturated FAs with branches formed by one methyl group (Table 2), and all of them with branches near the ω-end of the carbon chain, three iso-methyl FAs and two anteiso-methyl FAs. The highest content of BCFAs was observed in the representatives of the genus Meyerella. The cells of both Meyerella strains contain four BCFAs (three of them are found in both species), which in total comprise more than 3% of the total FAs, almost 4% in Mr. krienitzii VKM Al-428. C. vulgaris VKM Al-335 cells contain two BCFAs, Mc. lacustre VKM Al-343 one, and Mc. thermotolerans VKM-332 none.
Although BCFAs are present at low levels in cells, due to their physicochemical characteristics (Supplementary Table S1) they significantly affect the properties of membranes and their physiological functions [49]. The presence of BCFAs has been detected in bacteria, plants, and animals [50]. The biosynthesis and properties of BCFAs have been studied mainly in bacteria, where the role of non-linear FAs in increasing membrane fluidity has been demonstrated [51]. The fluidity of cell membranes in most organisms is usually determined by the presence of cis-unsaturated fatty acids; however, branched fatty acids can increase membrane fluidity and eliminate the organism’s need for unsaturated FAs. The formation of kinks at the branching point results in the reduction of the ordering of lipid molecules and the thickness of the lipid bilayer, thereby increasing the fluidity of the membranes, properties that are useful in low-temperature environments. It has been demonstrated that anteiso-branching is more effective at enhancing membrane fluidity than iso-branching [52]. The strain Mc. thermotolerans VKM Al-332, which inhabits a hot spring reservoir and can accumulate biomass even at 41 °C during the first 48 h of incubation [16], is devoid of BCFAs. At the same time, the FA profile of VKM Al-332 is characterized by a high PUFA content and a high unsaturation index (the second highest UI among the microorganisms analyzed after that of the closest relative strain Mc. lacustre VKM Al-343) (Table 1). The UI of Mc. thermotolerans VKM Al-332 FAs is higher than in strain C. vulgaris VKM Al-335, the species with moderate heat tolerance [53]. This suggests that the presence or absence of BCFAs can be used as an indicator of algae tolerance to low and high temperatures, respectively, while the value of the unsaturation index of FAs does not always reliably reflect the degree of thermotolerance. Certainly, our assumption based on the analysis of a limited number of algal strains should be tested on a larger number of species. In addition to their possible indicative functions as determinants of their phylogenetic relationship and temperature tolerance, BCFAs are of considerable biotechnological interest due to their health benefits [54], and the sources for their production, particularly from plants, are very limited [52].
Another group of non-oxygenated unusual fatty acids found in our experiments is represented by cyclic fatty acids. Three cyclic FAs have been identified in studied organisms; all of them are cyclopropane fatty acids: two–18-carbon chain FAs (2′-hexyl-[1,1′-bicyclopropyl]-2-octanoic acid and cyclopropaneoctanoic acid, 2-[[2-[(2-ethylcyclopropyl)methyl]cyclopropyl]methyl]-), and one-24-crabon chain (2-octyl-cyclopropanetetradecanoic acid). Interestingly, the highest number and total amount of cyclic FAs was found in the strain Mc. thermotolerans VKM Al-332, where the total content of three cyclic FAs was 5.44%. 2′-Hexyl-[1,1′-Bicyclopropyl]-2-octanoic acid was found in all studied organisms, while two others were only found in Mc. thermotolerans VKM Al-332. Cyclic fatty acids are a very poorly studied class of biological molecules, and the available information is mainly obtained from bacteria studies, although they have been found in protozoa, fungi, plants, as well as in food products such as milk, fish, and meat [55,56]. It is suggested that a cyclopropane ring can be formed at the double bond site of unsaturated fatty acids through the transfer of a methylene group from S-adenosyl methionine or through the intermediate steps associated with the formation of hydroperoxide groups [55,57]. Additionally, it cannot be ruled out that cyclic FAs are formed by modification of polyunsaturated fatty acids during extraction, although the high concentrations of antioxidant used in this work should prevent this process. The cyclopropane groups in the molecules of cyclic FAs we have found in our study are located at typical sites where double bonds are usually present. The presence of cyclic groups significantly alters the physicochemical properties of FAs by increasing their lipophilicity (Supplemental Table S1) [57]. Found cyclic FAs are characterized by higher calculated values of the octanol–water partition coefficient, which reflects lipophilicity [58]. According to the PubChem database (https://pubchem.ncbi.nlm.nih.gov/; accessed in May 2024), this coefficient ranges from 4.3 to 6.8 for the major FAs found in studied algae (Supplementary Table S1), increasing with the length of the carbon chain and decreasing with the degree of unsaturation. For cyclic FAs, these values are noticeably higher: the smallest value 7.8 was calculated for 2′-hexyl-[1,1′-bicyclopropyl]-2-octanoic acid and the highest value, greater than 11, was calculated for 2-octyl-cyclopropanetetradecanoic acid. In addition, these FAs are characterized by high molecular flexibility associated with the free rotation of atoms around single bonds. We believe that the lack of rigidity allows these highly lipophilic molecules to adopt different conformations and act as ‘molecular glue’ in membranes. Indeed, it has previously been reported that cyclic FAs increase the chemical and physical stability of bacterial membranes and protect them under adverse environmental conditions [59]. Based on the results of molecular dynamics simulations, it has been assumed that cyclopropane fatty acids in bacteria membranes play a dual function, stabilizing membranes against adverse conditions while promoting their fluidity [60], which allows microorganisms to survive under extreme environmental conditions [59]. Our finding of the presence of cyclic FAs in the strain Mc. thermotolerans VKM Al-332 probably, reveals a new potential aspect of the biological functions of these molecules in thermotolerant microalgae. Moreover, cyclic fatty acids are used as markers in food authentication, mainly in dairy products and meat [55]. The question of whether cyclic fatty acids could potentially serve as a marker for certain species or groups of algae at a higher taxonomic level, or be an indicator of certain properties, such as thermotolerance of microorganisms, needs to be explored in more detail.
Microalgae are known to be a valuable source not only of FA but also of FA derivatives [4]. In addition to their intrinsic value, PUFAs serve as a substrate for the formation of several valuable derivatives. Oxylipins—a large group of FA derivatives formed as a result of enzymatic or spontaneous oxygenation of FAs—play an important role in plants [61] and algae [62]. This group of FA derivatives is of particular interest due to their ecological importance [4] and therapeutic potential [48]. The most common are hydroxy, keto, and epoxy functional groups in the carbon chain; the carboxyl end group of FA can also undergo modifications. The potential of algae as producers of valuable oxylipins is almost unexplored, although individual studies indicate the diversity of this class of active compounds in algal cells [48,63]. The present study was not designed to extract and identify oxylipins but three oxylipins epoxy-, hydroxy-derivative, and fatty alcohol ester, were found to be co-extracted during fatty acid analysis. It should be noted that high concentrations of lipid-specific antioxidant were used in this work, which suggests that the oxylipins detected are not an oxidation product formed during fatty acid extraction.
Epoxy fatty acids are common in selected plant families [56]. cis-9,10-Epoxyoctadecanoic acid, identified in our study in C. vulgaris samples, has previously been found in humans [64], animals [65], seed oils [52], rust-infected wheat plants [66], and also in microalgae Scenedesmus acuminatus [67]. This oxylipin is most likely formed from oleic acid, and its biological functions are not yet known [41]. Hydroxy-FAs, containing a polar oxygenated functional group that would be incompatible with the hydrophobic environment of the membrane, have lubricant properties and show anti-inflammatory activity [4,48]. Such information may be important, as Chlorella spp. is currently the source of most microalgae-derived dietary supplements [47].
One of the most intriguing findings of this study is the significant correlation observed between the levels of different fatty acids in the organisms under investigation (Table 3). To the best of our knowledge, such an analysis has never been performed for green microalgae, although it may reveal new regulatory and phylogenetic relationships between biosynthetic pathways leading to the formation of different FAs. In our analysis, the modulus of the values of the correlation coefficients ranged from 0.036 to 0.973. The correlation coefficient equal to 0.973 and 0.953 for the presence of γ-linolenic acid and saturated FAs (14:0 and 16:0) is remarkable. The two saturated FAs, 14:0 and 16:0, were also correlated with each other (0.945). A strong correlation was observed between 16:2Δ7,10 and 18:2Δ9,12, and between 16:3Δ7,10,13 and 18:3Δ9,12,15, which may reflect the dependence of the content of these FAs on the activity of ω-3 FA desaturase in the studied microalgae [68]. It is important to exercise caution when interpreting the negative correlations in this analysis, as the content of individual FAs was estimated as a percentage of the total FA content. Consequently, an increase in one value is inevitably accompanied by a decrease in others, which contributes to the observed negative correlations. Moreover, in a living cell, some fatty acids can be used as substrates for the biosynthesis of others. This could explain the negative correlations observed in the present study between fatty acids with the same carbon chain length. However, this explanation is unlikely to be relevant to explain the significant negative correlations between 16:2Δ7,10 and 14:0, 16:2Δ7,10 and 16:0, and also between 16:2Δ7,10 and 18:3Δ6,9,12, which may have a different biological basis that we do not yet understand.
To identify the variables that account for most of the variance in the observed results, we performed a PCA (Figure 4). Upon analysis of only straight-chain FAs or the full range of detected FAs, including branched-chain and cyclic FAs, distinct graphical representations were obtained. When only straight-chain FAs were included, two components, PC1 and PC2, accounted for approximately 75% of the total variability (Figure 4a). Six fatty acids contributed positively to PC1 with the highest contribution from 16:0, 16:4, and α-linolenic acid. 16:4 and α-linolenic acid contributed positively and 16:0 negatively to PC2. 16:1 contributed negatively to PC1 and positively to PC2. In this PCA plot, three genera are distributed in three separate quarters of the plot area, and two Meyerella strains are most closely spaced. Thus, three genera were separated from each other, indicating a significant difference in the profile of major FA. When the analysis was performed using all detected FAs, including branched and cyclic FAs, the distribution of the studied objects on the PCA plot changed: both Meyerella strains were still closely spaced, but C. vulgaris VKM Al-335 and Mc. lacustre VKM Al-343 were distributed in the same quarter of the plot, while Mc. thermotolerance VKM Al-332 was distantly located in a separate quarter of the plot field. Thereby, strains of the species isolated from freshwater eutrophic reservoirs (C. vulgaris VKM Al-335 and Mc. lacustre VKM Al-343) turned out to be more similar in this case compared to the representatives of the same genera, living in different environmental conditions (Mc. lacustre VKM Al-343 and Mc. thermotolerans VKM Al-332 from a thermal spring). Therefore, it can be reasonably proposed that this distribution is better explained by habitat conditions than by genetic relationships, thereby suggesting the potential role of minor fatty acids in the adaptability of microalgae. If the reliability of the presented approach in FA analysis can be confirmed on a larger number of strains with different phylogenetic relatedness and habitat conditions, it will open new horizons for the characterization of algae chemotaxonomy and tolerance features.

5. Conclusions

Thus, our data show that each of the studied Chlorella-clade strains has a unique FA profile and provides an excellent source of fatty acids. Due to the considerable variability of the FA profile, the high content of unsaturated FAs, and the presence of rare valuable FAs, the studied organisms are useful for the production of food, food additives, feed, and pharmaceuticals. To fully exploit the biotechnological potential of microalgae, further comparative studies of different species should focus on identifying genetically determined traits of the fatty acid profile, as well as the norm of response and phenotypic plasticity of individual species. The results obtained expand our understanding of the biotechnological potential of representatives of the genera Micractinium and Meyerella and also suggest that in the future, as more data accumulate, the content of FA and their derivatives can be used as the chemotaxonomic characteristics. Since some strains have been isolated from ecosystems with extreme environmental conditions (hot springs and tundra soils in the Far North), a detailed analysis of the fatty acid profile of these organisms creates prerequisites for further investigation of the relationship between the fatty acid profile and algae adaptability to environmental conditions, such as extreme temperatures, salinity, and pH.
Furthermore, it is imperative to avoid a narrow perspective on the problem and to include species that are evolutionarily distant in the study. In the present study, this task was accomplished by including wheat, a representative of a higher plant that is cultivated in large agricultural areas. The research results showed that none of the studied strains surpass wheat leaves in content of unsaturated fatty acids, although each of them can be used to solve specific practical problems. In the future, additional studies are also needed to identify the optimal conditions for cultivating (temperature, light conditions, and the composition of nutrient media) the studied strains in order to obtain a maximum biomass enriched with target metabolites within the shortest possible time and at minimal cost.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/ijpb15030060/s1, Table S1: Physicochemical properties of FAs.

Author Contributions

E.K., E.D., E.T., A.T. and T.S. contributed equally to this work. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Russian Science Foundation, grant No. 22-16-00047.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

All authors confirm that the data supporting the findings of this study are available within the article and its Supplementary Material. In addition, raw data supporting this study’s findings are available from the corresponding author upon reasonable request.

Acknowledgments

To Shared Core Facilities of the Pushchino Scientific Center for Biological Research (http://www.ckp-rf.ru/ckp/670266/).

Conflicts of Interest

The authors declare no conflicts of interest.

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Ijpb 15 00060 g001
Figure 1. Microscopic images of strains (a) Chlorella vulgaris VKM Al-335, (b) Micractinium lacustre VKM Al-343, (c) Mc. thermotolerans VKM Al-332, (d) Meyerella similis VKM Al-346, (e) Mr. krienitzii VKM Al-428. Scale bar: 10 μm. The images were obtained at 100× magnification.
Figure 1. Microscopic images of strains (a) Chlorella vulgaris VKM Al-335, (b) Micractinium lacustre VKM Al-343, (c) Mc. thermotolerans VKM Al-332, (d) Meyerella similis VKM Al-346, (e) Mr. krienitzii VKM Al-428. Scale bar: 10 μm. The images were obtained at 100× magnification.
Ijpb 15 00060 g001
Ijpb 15 00060 g002
Figure 2. A rooted ultrametric phylogenetic tree of green microalgae from Chlorella-clade, constructed by the Bayes inference (BI), based on the 18S–ITS1–ITS2 sequences (2884 bp). As statistical support for the nodes of the tree, SH-aLRT support (SH-aLRT), posterior probabilities (PP), and bootstrap values (BP), respectively, are indicated; the values of SH-aLRT < 70%, BP < 70%, and PP < 0.7 are not shown. The model of nucleotide substitutions: TIM2e + I + G4. Note: studied VKM strains are highlighted in bold; *—authentic strains; (T)—type species.
Figure 2. A rooted ultrametric phylogenetic tree of green microalgae from Chlorella-clade, constructed by the Bayes inference (BI), based on the 18S–ITS1–ITS2 sequences (2884 bp). As statistical support for the nodes of the tree, SH-aLRT support (SH-aLRT), posterior probabilities (PP), and bootstrap values (BP), respectively, are indicated; the values of SH-aLRT < 70%, BP < 70%, and PP < 0.7 are not shown. The model of nucleotide substitutions: TIM2e + I + G4. Note: studied VKM strains are highlighted in bold; *—authentic strains; (T)—type species.
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Ijpb 15 00060 g003
Figure 3. Fatty acid composition of total lipids of studied representatives of the Chlorella-clade. Essential fatty acids are designated using delta nomenclature indicating the number of carbon atoms in the fatty acid chain, the number of double bonds after the colon, and the position of the double bonds after the “Δ” sign. Data presented as means ± standard deviation. Different letters indicate statistically significant differences between genotypes in the content of a given fatty acid according to one-way analysis of variance (ANOVA), p ≤ 0.05.
Figure 3. Fatty acid composition of total lipids of studied representatives of the Chlorella-clade. Essential fatty acids are designated using delta nomenclature indicating the number of carbon atoms in the fatty acid chain, the number of double bonds after the colon, and the position of the double bonds after the “Δ” sign. Data presented as means ± standard deviation. Different letters indicate statistically significant differences between genotypes in the content of a given fatty acid according to one-way analysis of variance (ANOVA), p ≤ 0.05.
Ijpb 15 00060 g003
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Figure 4. Principal component analysis (PCA) of fatty acid content in studied microalgae was conducted on the standardized values (standardized scaling), using only straight-chain (a) and all major and minor (b) fatty acids. X- and Y-axes show PC1 and PC2, respectively, with the amount of variance contained in each component. In plot (a), arrows indicate the contribution of individual FAs: 1—14:0, 2—15:0, 3—16:0, 4—16:1Δ7, 5—16:2Δ7,10, 6—16:3Δ7,10,13, 7—16:4Δ4,7,10,13, 8—18:1Δ9, 9—18:2Δ9,12, 10—18:3Δ9,12,15, 11—18:3Δ6,9,12.
Figure 4. Principal component analysis (PCA) of fatty acid content in studied microalgae was conducted on the standardized values (standardized scaling), using only straight-chain (a) and all major and minor (b) fatty acids. X- and Y-axes show PC1 and PC2, respectively, with the amount of variance contained in each component. In plot (a), arrows indicate the contribution of individual FAs: 1—14:0, 2—15:0, 3—16:0, 4—16:1Δ7, 5—16:2Δ7,10, 6—16:3Δ7,10,13, 7—16:4Δ4,7,10,13, 8—18:1Δ9, 9—18:2Δ9,12, 10—18:3Δ9,12,15, 11—18:3Δ6,9,12.
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Table 1. General characteristics of fatty acid profiles of studied microalgae in comparison with leaves of T. aestivum.
Table 1. General characteristics of fatty acid profiles of studied microalgae in comparison with leaves of T. aestivum.
C. vulgaris
VKM Al-335		Mc. lacustre VKM Al-343Mc. Thermotolerans
VKM Al-332		Mr. similis
VKM Al-346		Mr. krienitzii
VKM Al-428		T. aestivum
16-carbon FA, %59.1562.27 **54.4443.7635.5024.59
18-carbon FA, %34.0634.9841.8349.6659.36 **75.41
MUFA, %4.855.913.189.08 **6.066.07
PUFA, %44.0973.7768.4070.1175.37 **75.41
UFA, %48.9479.6871.5879.1981.43 **81.48
UI1.312.39 **1.951.841.852.25
ω-3 FA, %31.6963.85 **51.1634.8728.3267.75
ω-6 FA, %12.409.9117.2435.2447.04 **7.66
ω-3/ω-62.566.44 **2.970.990.608.84
“**” indicates the highest values among studied microalgae. UFA—unsaturated fatty acids; MUFA—monounsaturated fatty acids; PUFA—polyunsaturated fatty acids; UI—unsaturation index. The indexes of the fatty acid profile of wheat that are higher than those of the studied microalgae are given in bold and underlined.
Table 2. Minor fatty acids in studied algae cells.
Table 2. Minor fatty acids in studied algae cells.
C. vulgaris
VKM Al-335		Mc. lacustre VKM Al-343Mc. thermo-tolerans
VKM Al-332		Mr. similis
VKM Al-346		Mr. krienitzii
VKM Al-428
Minor straight-chain fatty acids
Tetradecanoic acid1.18 ± 0.200.99 ± 0.070.97 ± 0.040.45 ± 0.240.28 ± 0.02
Pentadecanoic acid 0.84 ± 0.021.69 ± 0.96
Branched-chain fatty acids
Tetradecanoic acid, 12-methyl-0.52 ± 0.04 1.07 ± 1.050.24 ± 0.21
Hexadecanoic acid, 14-methyl- 0.68 ± 0.050.83 ± 0.55
Hexadecanoic acid, 15-methyl-1.32 ± 0.031.20 ± 0.09
Heptadecanoic acid, 16-methyl- 1.45 ± 0.251.53 ± 0.31
Octadecanoic acid, 17-methyl- 1.18 ± 0.46
Cyclic fatty acids
[1,1′-Bicyclopropyl]-2-octanoic acid, 2′-hexyl-
Ijpb 15 00060 i001		 0.47 ± 0.192.04 ± 1.520.68 ± 0.470.84 ± 0.96
Cyclopropaneoctanoic acid, 2-[[2-[(2-ethylcyclopropyl)methyl] cyclopropyl]methyl]
Ijpb 15 00060 i002		 3.00 ± 0.13
Cyclopropanetetradecanoic acid, 2-octyl-
Ijpb 15 00060 i003		 0.40 ± 0.07
Table 3. Covariance matrix among the straight-chain fatty acid data set obtained from the analysis of five studied strains.
Table 3. Covariance matrix among the straight-chain fatty acid data set obtained from the analysis of five studied strains.
14:015:016:016:1Δ716:2Δ7,1016:3Δ7,10,1316:4Δ4,7,10,1318:1Δ918:2Δ9,1218:3Δ9,12,1518:3Δ6,9,12
14:01−0.3920.945−0.462−0.922−0.1450.084−0.348−0.8450.4070.973
15:0−0.3921−0.338−0.1290.6040.544−0.3740.7080.223−0.220−0.387
16:00.945 ***−0.3381−0.659−0.784−0.285−0.230−0.212−0.6470.1810.953
16:1Δ7−0.462−0.129−0.659 *10.252−0.1070.7470.0810.221−0.094−0.617
16:2Δ7,10−0.922 ***0.604−0.784 **0.25210.053−0.3780.6480.894−0.640−0.904
16:3Δ7,10,13−0.1450.544−0.285−0.1070.05310.176−0.191−0.3230.665−0.061
16:4Δ4,7,10,130.084−0.374−0.2300.747−0.3780.1761−0.443−0.4400.542−0.036
18:1Δ9−0.3480.708−0.2120.0810.648−0.191−0.44310.493−0.789−0.444
18:2Δ9,12−0.845 **0.223−0.6470.2210.894 **−0.323−0.4400.4931−0.773−0.815
18:3Δ9,12,150.407−0.2200.181−0.094−0.6400.665 *0.542−0.789 **−0.773 **10.470
18:3Δ6,9,120.973***−0.3870.953 ***−0.617−0.904 **−0.061−0.036−0.444−0.815 *0.4701
Statistically significant positive correlations are marked by green and negative by yellow. Stars indicate statistically confirmed significance: *, p < 0.1; **, p < 0.05; ***, p < 0.001.
Table 4. Oxylipins in studied algae strains.
Table 4. Oxylipins in studied algae strains.
C. vulgaris
VKM Al-335		Mc. lacustre VKM Al-343Mc.thermo-tolerans
VKM Al-332		Mr. similis
VKM Al-346		Mr. krienitzii
VKM Al-428
cis-9,10-epoxyoctadecanoic acid
Ijpb 15 00060 i004		+−−−−
dodecanoic acid, 3-hydroxy-
Ijpb 15 00060 i005		−−−++
7-methyl-Z-tetradecen-1-ol acetate
Ijpb 15 00060 i006		+−−−−
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Krivina, E.; Degtyaryov, E.; Tebina, E.; Temraleeva, A.; Savchenko, T. Comparative Analysis of the Fatty Acid Profiles of Selected Representatives of Chlorella-Clade to Evaluate Their Biotechnological Potential. Int. J. Plant Biol. 2024, 15, 837-854. https://doi.org/10.3390/ijpb15030060

AMA Style

Krivina E, Degtyaryov E, Tebina E, Temraleeva A, Savchenko T. Comparative Analysis of the Fatty Acid Profiles of Selected Representatives of Chlorella-Clade to Evaluate Their Biotechnological Potential. International Journal of Plant Biology. 2024; 15(3):837-854. https://doi.org/10.3390/ijpb15030060

Chicago/Turabian Style

Krivina, Elena, Evgeny Degtyaryov, Elizaveta Tebina, Anna Temraleeva, and Tatyana Savchenko. 2024. "Comparative Analysis of the Fatty Acid Profiles of Selected Representatives of Chlorella-Clade to Evaluate Their Biotechnological Potential" International Journal of Plant Biology 15, no. 3: 837-854. https://doi.org/10.3390/ijpb15030060

APA Style

Krivina, E., Degtyaryov, E., Tebina, E., Temraleeva, A., & Savchenko, T. (2024). Comparative Analysis of the Fatty Acid Profiles of Selected Representatives of Chlorella-Clade to Evaluate Their Biotechnological Potential. International Journal of Plant Biology, 15(3), 837-854. https://doi.org/10.3390/ijpb15030060

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