Lipids in plant tissue cultures. VI. Effect of temperature on the lipids of Brassica napus and Tropaeolum majus cultures

The lipid contents of callus cultures of rape (Brassica napus) and nasturtium (Tropaeolum majus) increase in response to decreasing the temperature, though to different degree. Irrespective of the incubation temperature the lipids in cultures of both plants contain as predominant classes steryl glycosides, esterified steryl glycosides, sterols, steryl esters and fatty acids and, as minor constituents, various proportions of triacylglycerols, phospholipids and several unidentified fractions.The ratio of phospholipids to triacylglycerols as well as the ratio of diacylglycerophosphorylethanolamines to dicylglycerophosphorylcholines increase with time both in rape cultures incubated at 30°C and in those kept at 5°C.The lipids in rape and nasturtium cultures grown at 30°C contain smaller proportions of polyunsaturated fatty acids than the lipids in cultures incubated at 5°C. Erucic acid, the major constituent fatty acid of the seed lipids, in both plants, occurs only in trace amounts in the lipids of callus cultures. In contrast, linoleic and linolenic acids, which occur only in traces in the seed lipids of nasturtium, are major constituent fatty acids in the lipids of callus cultures derived from seedlings of this plant.The levels of constituent polyunsaturated fatty acids in the diacylglycerophosphorylethanolamines and the diacylglycerophosphorylcholines increase with time whereas in the triacylglycerols only linolenic acid is slightly increased.     

Lipids in plant tissue cultures. V. Effect of environmental conditions on the lipids of Glycine soja and Brassica napus cultures.

The environmental conditions prevailing during growth of plant tissue cultures affect the concentration of certain lipid classes and the fatty acid patterns of the total lipids. Irrespective of whether the cultures are grown under continuous illumination or in the dark, aerated cultures contain larger proportions of sterols, steryl esters, steryl glycosides and various sterylglycolipids and lower concentrations of squalene than non-aerated cultures. The lipids of the latter contain larger proportions of saturated very long-chain fatty acids than those of the former cultures.     

Dimorphism-associated variations in the lipid composition of Candida albicans

Yeast and mycelial forms of Candida albicans ATCC 10231, growing together in 12 h and in 96 h cultures, were separated and their lipids were extracted and characterized. The total lipid content of the yeast forms was always lower than that of the mycelial forms. In 12 h cultures the lipids from the two morphological forms consisted mainly of polar compounds, viz, phospholipids and glycolipids. In 96 h cultures both the yeast and mycelial forms accumulated substantial amounts of apolar compounds, mainly steryl esters and triacylglycerols. The mycelial forms were more active than the yeast forms in this respect. Major differences in the lipid composition between the two morphological forms involved the contents of sterols and complex lipids that contain sterols. As a rule, the yeast lipids contained much larger proportions of free sterols than the mycelial lipids. However, the mycelial lipids contained several times more sterols than the yeast forms but bound as steryl glycosides, esterified steryl glycosides and steryl esters. Steryl glycosides and esterified steryl glycosides occurred in yeast lipids only in traces, if at all. The major steryl glycoside in the mycelial forms was unequivocally identified as cholesteryl mannoside. At both phases of growth the apolar and polar lipid fractions from the mycelial forms contained higher levels of polyunsaturated fatty acids (18:2 and 18:3) but lower levels of oleic acid (18:1) than the corresponding fractions from the yeast forms. The lipid content and composition of 12 h and 96 h yeast and mycelial forms of C. albicans KCCC 14172, a clinical isolate, were almost identical with those of C. albicans ATCC 10231.     

The lipids in photoautotrophic and heterotrophic cell suspension cultures of Chenopodium rubrum

Resembling the lipids in the leaves and other green organs of intact plants, the lipids in photoautotrophic cell cultures of Chenopodium rubrum were found to contain high proportions of monogalactosyldiacylglycerols and digalactosyldiacylglycerols, as well as fair amounts of sulfoquinovosyldiacylglycerols and diacylglycerophosphoglycerols. Conversely, the heterotrophic cell cultures, from which the photoautotrophic cultures had been derived, contained only traces of these compounds. The heterotrophic cultures were rich in sterols, sterol esters, sterol glycosides, and esterified sterol glycosides. The lipids of photoautotrophic cell cultures contained higher proportions of constituent linolenic acid, but lower concentrations of linoleic acid than those of heterotrophic cultures. In the photoautotrophic cultures, as in green leaves, linolenic acid was predominantly estrified in monogalactosyldiacylglycerols and digalactosyldiacylglycerols. This investigation shows that it is possible to select strains of cell cultures, which are capable of grosing photoautotrophically, with the aim of activating the biosynthesis of specific metabolites.     

Lipids in plant tissue cultures. II. Unusual fatty acids in lipids of Hydnocarpus anthelminthica cultures

The lipids in callus cultures of Hydnocarpus anthelminthica were studied after 60, 160 and 460 days of growth. In each of the cultures the lipid classes usually found in plant tissue cultures were detected. With increasing age of the cultures the total lipid content as well as the proportions of triglycerides decreased. The major constituent fatty acids of the total lipids were palmitic and linoleic acids. Small amounts of cyclopentenyl fatty acids were also present. The proportions of saturated straight-chain fatty acids increased with the age of the cultures whereas the proportions of monounsaturated straight-chain fatty acids decreased. Only small changes were observed with polyunsaturated fatty acids. The content of cyclopentenyl fatty acids rose with the age of the cultures. The monounsaturated straight-chain fatty acids consisted of mixtures of isomers whose composition changed with the age of the cultures. In contrast, the polyunsaturated straight-chain fatty acids belonged exclusively to the Δ9 series, regardless of the age of the cultures.     

Intracellular distribution and origin of sterols in Calendula officinalis leaves

In Calendula officinalis leaves 66% of all steryl forms are present in the ‘microsomal fraction’ (IV), 24% in the mitochondrial and Golgi membranes (III), 5% in the ‘chloroplast’ (II), 4% in the ‘cell wall and membrane’ (I) fraction and 1%. in the cytosol. Free sterols, their esters, glycosides and acylated glycosides are present in varying proportions in all cellular subtractions. Mevalonate-[2¹⁴C] labelling of sterols derived from various steryl forms showed that free sterols and all their derivatives, i.e. steryl esters and glucosides, are formed in fraction IV and are then translocated to other organelles. Fraction III is the main site of glycosylation of transported sterols as well as of acylation of steryl glycosides.     

Sterols and fatty acids from three species of mangrove

The hydrolysis of steryl esters on thin-layer chromatographic plates by porcine pancreatic lipase is described. The sterols and fatty acids produced were separated on the same plate, recovered, and analysed by gas-liquid chromatography for their compositions. Synthetic cholesteryl esters containing various saturated and unsaturated fatty acids and synthetic steryl oleates with various sterols were lipolysed along with steryl esters of Acanthus ilicifolius, Bruguiera gymnorhiza and Rhizophora mucronata mangrove leaves. The major sterol was sitosterol which was accompanied by cholesterol, campesterol, stigmasterol and 28-isofucosterol. In addition, stigmast-7-en-3β-ol was present in R. mucronata leaves. The component fatty acids found in all three species were 16:0, 18:0, 18:1, 18:2 and 18:3. The relative proportions of the sterols and fatty acids were significantly different from the chemotaxonomic standpoint. The results obtained by carrying out plate lipolysis for 45 min at 40° compared well with those produced by conventional chemical hydrolysis.     

Localization of lipids containing (Z)-11-eicosenoic acid and (Z)-13-docosenoic acid in Tropaeolum majus

(Z)-11-Eicosenoic (gondoic) and (Z)-13-docosenoic (erucic) acids were found in large proportions as constituent fatty acids of the triglycerides and polar lipids in seeds and petals of Tropaeolum majus. In the lipids of the other floral organs as well as in those of vegetative organs, only traces of these fatty acids were detected. During seed germination, the proportions of the two fatty acids did not change. (Z)-9,12- Octadecadienoic (linoleic) and (Z)-9,12,15-octadecatrienoic (linolenic) acids, which occurred only in traces in lipids of the seeds, were major constituent fatty acids of lipids in floral and vegetative organs as well as those of callus cultures.     

Phytosterols in tobacco leaves at various stages of physiological maturity

Leaves of varying maturity from 84-day-old tobacco plants were harvested and analyzed for total sterol and their individual sterol components. The mature leaves had a significant higher sterol content than the immature leaves. Separation into free sterols, steryl esters, steryl glycosides, and acylated steryl glycosides showed that the free sterols accounted for most of the sterol increase, and stimgasterol was principally responsible for this increase.     

Sterols in relation to ageing of seeds of Helianthus annuus and Cicer arietinum

Under accelerated ageing at high relative humidity and high temperature for 4 days germination and membrane permeability remained unaffected both in sunflower and chick pea seeds. However, the steryl glycoside concentration in the pooled leachate increased progressively with ageing. Total sterols, as well as steryl glycosides and free sterols of the seeds, increased with a concomitant decline in steryl esters under accelerated ageing. Pretreatment with the sterol biosynthesis inhibitor SK & F 7997A₃ prevented the increase of total sterols under accelerated ageing conditions but there were increases in the amounts of steryl glycosides and free sterols and a decrease in steryl ester after such treatment, therefore, indicating interconversions of the various sterol types. Accelerated ageing also caused increases in free amino acids and soluble carbohydrate. Low relative humidity-high temperature and high relative humidity-low temperature failed to produce such effects.     

The Neutral Lipids of Paramecium tetraurelia: Changes with Culture Age and the Detection of Steryl Esters in Ciliary Membranes1

Neutral lipids, particularly triglycerides, accounted for the major decrease in the total lipid content in Paramecium cells that occurs with culture age. Sterols, triglycerides, and steryl esters were the major classes of neutral lipids in cells and isolated cilia. Free as well as high concentrations of esterified sterols were detected in purified ciliary membrane preparations. Stigmasterol and 7-dehydrostigmasterol were the major components of both free and esterified sterols of cells and cilia; however, when cholesterol was present in the growth medium, it was desaturated to 7-dehydrocholesterol and incorporated into cellular and ciliary lipids. Free fatty acids from cells and triglycerides from cells and cilia were low in polyunsaturated fatty acids and reflected the composition of fatty acids in the culture medium. An exception was the reduced concentration of stearate in triglycerides from whole cells. Greater than 50% of triglyceride fatty acids from cilia were saturated. The fatty acid compositions of cellular triglycerides and ciliary steryl esters did not change with culture age, but those of cellular steryl esters and ciliary triglycerides did change. In comparison with phospholipids, these neutral lipid fatty acid compositional changes were smaller. The sensitivity of these stigmasterol-containing cells to polyene antibiotics indicated that they were killed by nystatin > filipin > amphotericin B. The unexpected finding of high concentrations of steryl esters in ciliary membrane preparations is discussed.     

Occurrence of steryl glycosides and acylated steryl glycosides in some marine algae

There is some controversy concerning the presence of steryl glycosides and acylated steryl glycosides in eucaryotic algae. These two classes of sterol compounds were investigated in species belonging to the three major groups of eucaryotic algae: green algae (Ulva gigantea, Cladophora rupestris), brown algae (Fucus vesiculosus, Ascophyllum nodosum), and red algae (Rhodymenia palmata, Porphyridium sp.). All these algae contain both steryl glycosides and acylated steryl glycosides. The sterol components of these compounds vary according to the alga but they are always the same as the free sterols of the alga in question. The most common sugar moiety is glucose. In the acylated steryl glycosides, the fatty acid is mainly palmitic acid. The percentage of these compounds (as a percentage of the total sterol content) is often low.     

Lipid composition of plasma membranes isolated from sunflower seedlings grown under water-stress

Plasma membranes were isolated by aqueous two-phase-partitioning from sunflower ( Helianthus annuus cv. Isabel) seedlings grown both under field irrigation and dryland conditions. Water-stressed plants showed a decrease in the leaf water potential and in the osmotic potential at full turgor, with the turgor pressure remaining at positive values. Dryland conditions also induced a reduction in the bulk modulus of elasticity. Plasma membranes of irrigated plants were characterized by high contents of phospholipids (68% of total lipids), free sterols (15. 7%) and glycolipids (9. 1%), mainly glycosphingolipids and steryl glycosides. Diacylglycerols, triacylglycerols and free fatty acids were also present. The major phospholipids were phosphatidylcholine and phosphatidylethanolamine with smaller amounts of phosphatidylinositol and phosphatidylglycerol. Following water stress, the plasma membranes showed a reduction of about 24 and 31% in total lipids and phospholipids, respectively. Also the amounts of glycolipids and diacylglycerols decreased significantly upon water stress. There was no change in free fatty acids, however, and triacylglycerols and free sterols increased. As a consequence, the free sterol to phospholipid molar ratio increased from 0. 4 to 0. 7 under water deficit conditions. The ratio of phosphatidylcholine to phosphatidylethanolamine increased from 1. 1 (control plants) to 1. 6 (water-stressed plants), while phosphatidic acid rose to 4% of total phospholipids. Dehydration did not result in any substantial change in the unsaturation level of the individual lipid classes, however. The results show that dryland conditions resulted in a marked alteration in the lipid composition of the sunflower leaf plasma membrane     

Hexane soluble non-volatiles in flowering to fruiting stages of Fatsia japonica

The n-hexane soluble non-volatile fraction of the acetone extracts from the flower buds, the flowers and the immature and the mature fruits of Fatsia japonica were all found to contain fatty acids, fatty acid methyl esters, squalene, β-amyrin and sterols. At all the stages between budding to the mature fruit, the major fatty acids were palmitic and linoleic acids and the major phytosterol was stigmasterol. In addition steryl and β-amyrenyl esters were found in the flowers and the immature and the mature fruits, but these esters were not present in the flower buds. Sitosteryl ester was the major constituent of the steryl ester fraction in the fruiting stages. Phytol was found in only the flowering stage and triglycerides in only the mature fruits. The variations in the lipid constituents is discussed in relation to the stages from budding to the mature fruit.     

Lipid composition of the fat body and haemolymph and its relation to lipid release in Oncopeltus fasciatus

Lipid composition of the fat body and haemolymph of male milkweed bug, Oncopeltus fasciatus, was determined. Triglycerides were the predominant lipids of the fat body while diglycerides accounted for the major lipid in the haemolymph. Sterols, sterol esters, and non-esterified fatty acids were present in both fat body and haemolymph besides triglycerides and diglycerides. Only traces of monoglycerides were detected.Gas chromatographic analysis of the fatty acids revealed a difference in the fatty acid composition between fat body and haemolymph glycerides and sterol esters. Oleate and linoleate were the predominant unsaturated fatty acids in both fat body and haemolymph lipids and in the milkweed seeds as well.When fat body was labelled in vivo and in vitro with ¹⁴C-palmitate, the fatty acid was incorporated largely into the triglycerides. When the prelabelled fat body was incubated with a medium containing haemolymph the fat body released lipids mainly as diglycerides. Some radioactivity was observed in the triglycerides and non-esterified fatty acids also.Electrophoretic analysis of the incubation medium containing the haemolymph revealed that the released lipids were bound to three haemolymph lipoprotein bands. Lipid mobilization, release, and transport in Oncopeltus are discussed in relation to studies on other insects.     

Plant sterols: Diversity,biosynthesis, and physiological functions

Sterols, which are isoprenoid derivatives, are structural components of biological membranes. Special attention is now being given not only to their structure and function, but also to their regulatory roles in plants. Plant sterols have diverse composition; they exist as free sterols, sterol esters with higher fatty acids, sterol glycosides, and acylsterol glycosides, which are absent in animal cells. This diversity of types of phytosterols determines a wide spectrum of functions they play in plant life. Sterols are precursors of a group of plant hormones, the brassinosteroids, which regulate plant growth and development. Furthermore, sterols participate in transmembrane signal transduction by forming lipid microdomains. The predominant sterols in plants are β-sitosterol, campesterol, and stigmasterol. These sterols differ in the presence of a methyl or an ethyl group in the side chain at the 24th carbon atom and are named methylsterols or ethylsterols, respectively. The balance between 24-methylsterols and 24-ethylsterols is specific for individual plant species. The present review focuses on the key stages of plant sterol biosynthesis that determine the ratios between the different types of sterols, and the crosstalk between the sterol and sphingolipid pathways. The main enzymes involved in plant sterol biosynthesis are 3-hydroxy-3methylglutaryl-CoA reductase, C24-sterol methyltransferase, and C22-sterol desaturase. These enzymes are responsible for maintaining the optimal balance between sterols. Regulation of the ratios between the different types of sterols and sterols/sphingolipids can be of crucial importance in the responses of plants to stresses.     

The sterols of growth and stationary phases of Aspergillus nidulans cultures

The accumulation of 4-desmethyl and 4,4-dimethyl sterols, as well as the triterpenoid beta-amyrin, was analysed during both exponential and stationary phases of Aspergillus nidulans growth. Throughout growth, the amount of 4-desmethyl sterol was proportional to the cellular dry weight, while the dimethyl sterols and beta-amyrin stopped accumulating after day 2. The sterols were found primarily as the free alcohol and not as fatty acid esters, the glycosides, or acyl glycosides. The amount of beta-amyrin in stationary phase cultures was affected by the concentrations of Mg2+ and Cu2+.     

Steryl glycosides and acyl steryl glycosides from Musa paradisiaca

From peeled fruits of Musa paradisiaca (banana, vegetable variety), two new acyl steryl glycosides, sitoindoside-III and sitoindoside-IV, and two new steryl glycosides, sitosterol gentiobioside and sitosterol myo-inosityl-β-D-glucoside, have been isolated by gradient solvent extraction and extensive chromatography (CC, prep. TLC, GC and HPLC). The compounds have been characterized by comprehensive spectroscopic analyses (IR, ¹H NMR, GC, mass spectra, [α]_D) and crucial chemical transformation. Additionally, seasonal variations of the total sterols, free sterols, steryl esters, steryl glycosides and acyl steryl glycosides in the active samples of banana have been analysed. The results provide a basis for the observed fluctuations in the anti-ulcerogenic activity of the extracts, in different seasons, and the importance of appropriate formulation of the pure principles to optimize the activity.     

A survey of polar and nonpolar lipids from epidermis and epidermal appendages of the chicken (Gallus domesticus)

Total lipid was extracted from chicken (Gallus domesticus) epidermis, leg scale, claws, feathers and preen glands and analyzed by quantitative thin-layer chromatography. All of the tissue lipids contained large proportions of wax diesters, triglycerides, and free sterols and variable proportions of phospholipids, steryl esters and free fatty acids. All of the keratinized tissues, but not the preen gland, contained ceramides, acylceramides and cholesteryl sulfate. Acylglucosylceramides were found only in full thickness epidermis. Glucosylsterols and acylglucosylsterols were found in the keratinized tissues, and may be of significance in the evolutionary history of the epidermal water barrier.     

Occurrence of steryl and wax esters in Dicranum elongatum

The steryl and wax esters of the frozen subarctic moss Dicranum elongatum Schleich contained fatty acids 39.8 mg per gram dry green tissue. The content decreased with increasing age of the moss shoots, but no great changes were found in the fatty acid pattern of the esters. The major part of the steryl and wax ester fraction of the green shoots was made up of esterified sterols (85%), and the rest (15%) of esterified aliphatic alcohols. No great changes were found in their relative proportions with increased age of the shoots. Some changes were evident in the pattern of individual esterified sterols, however. The proportion of cycloartenol was lower in the older parts than in the green part, and the proportion of campesterol, stigmasterol, and β-sitosterol were lower in the green part. The major esterified aliphatic alcohols were 1-octadecanol, phytol and geranylgeraniol. The proportion of geranylgeraniol was highest in the green part and that of phytol in the older parts. The main alcohol of the surface lipids was 1-octadecanol.