Free Sterol Composition Research Articles

Modulation of plant resistance to diseases by water-soluble chitosan

Low-molecular-weight water-soluble chitosan with a molecular weight of 5 kDa obtained after enzymatic hydrolysis of native crab chitosan was shown to display an elicitor activity by inducing the local and systemic resistance of Solanumi tuberosum potato and Lycopesicon esculentum tomato to Phytophthora infestans and nematodes, respectively. Chitosan induced the accumulation of phytoalexins in tissues of host plants, decreased the total content and changed the composition of free sterols producing adverse effects on infesters, activated chitinases, beta-glucanases, and lipoxygenases, and stimulated the generation of reactive oxygen species. The activation of protective mechanisms in plant tissues inhibited the growth of taxonomically different pathogens (parasitic fungus Phytophthora infestans and root knot nematode Meloidogyne incognita).

Effects of environmental factors and metals on selenastrum capricornutum lipids

Studies of the effects of illumination, temperature and heavy metal exposure (Cu 2, Zn 2, Cd 2) on Selenastrum capricornutum have shown that these factors alter both the total fatty acid and free sterol composition of cells in batch culture. Dark treatment resulted in a decrease in the relative proportions of oleate (18 : 1) and an increase linoleate (18 : 2), together with an increase in the relative proportion of the sterol 24-ethyl-5 α-cholest-7-en-3 β-ol (Δ 7-chondrillastenol) and a decrease in 24-ethyl-5 α-cholesta-7,22-dien-3 β-ol (chondrillasterol). A shift in temperature from 25° to 10° led to an increase in the relative proportion of oleate and a decrease in linoleate and parinate (18 : 4), together with a significant increase in the relative proportion of 24-methyl-5 α-cholest-7-en-3 β-ol (ergostenol). Generally, exposure to heavy metal ions led to an increase in oleate (with all three metals) and altered the relative proportions of linoleate and parinate (changes being metal specific). Metal (ion) treatment also significantly increase Δ 22 desaturation of the 24-ethyl sterol components. The changes in the composition of many of the individual lipid components in response to heavy metal ion exposure occurred at concentrations which did not significantly affect the organisms specific growth rate. For example, the relative proportion of oleate was affected with only 1 μM Cu 2 in solution ( P < 0.10).

Properties of plasma membranes of P hsp 70- ipt transformed tobacco ( Nicotiana tabacum)

Application of 10 successive daily heat shocks reduced the growth of control tobacco ( Nicotiana tabacumL. cv. Petit Havana SR1) plants by about 15%; for P hsp 70- ipt transformed plants this is about 48%. The shoot diameter of these ipt-transformed plants increased by about 75%. In addition, in heat shock treated ipt-plants (IPT-HS) the upper lateral buds grew out due to a reduction of apical dominance. The older leaves of IPT-HS plants had a higher chlorophyll content. In spite of the observed effects due to a higher endogenous cytokinin content in the IPT-HS plants, no significant changes were observed on the plasma membrane fatty acid composition, nor on its fluidity as determined from the steady-state fluorescence anisotropy of DPH. Only a minor change in the plasma membrane free sterol composition was found as evidenced by a 20% decrease in the stigmasterol to sitosterol ratio in IPT-HS, indicative for a possible anti-senescence effect of enhanced endogenous cytokinins, but without significant effects on the plasma membrane function. © 1997 Elsevier Science Ltd. All rights reserved

Content and Composition of Free Sterols and Free Fatty Alcohols in Jojoba Oil

A method was developed for the isolation and identification of phytosterols and fatty alcohols in jojoba oil. The method consists in the separation of these compounds from wax esters in the oil by means of an aluminum oxide column followed by further fractionation of the minor components by column chromatography on silica gel. The 4-demethylsterols, 4-methylsterols, triterpene alcohols (4,4-dimethylsterols), and fatty alcohols are identified by means of their gas chromatographic and mass spectrometric data. The present paper includes a method for the quantitation of the free sterols in jojoba oil. Keywords: Plant; alcohol; sterol; lipid; Simmondsia

Effects of arbuscular mycorrhiza (AM) on health ofLinum usitatissimum L. infected by fungal pathogens

Effects of arbuscular mycorrhizal (AM) symbiosis on health ofLinum usitatissimum infected by fungal pathogens were investigated exemplarily. Physiological and biochemical analyses were done to explain the mechanisms underlying the AM effects. AM plants showed increased resistance against the wilt pathogen (Fusarium oxysporum f. sp.lini), the level of this effects depended on the plant cultivars which all showed the same level of root colonization by arbuscular mycorrhizal fungi (AMF). In contrary to that, AM plants were highly susceptible against the shoot pathogenOidium lini, but they suffered less than non-AM plants in terms of shoot fresh weight, CO2 assimilation and content of sucrose in shoot apex. This indicates that AM not only activates resistance mechanisms but also can induce tolerance against pathogens. The concentration of phytohormones such as auxin- and gibberellin-like substances were increased in shoots of AM plants. In roots the ethylene production was increased, too. Furthermore the content and composition of free sterols were highly altered in leaves of AM plants. Root infection by AMF caused an increased respiratory activity and a reduced degree of DNA methylation, but both modifications only occurred in infected root parts indicating an increasing gene activity. The presented results suggest that nearly all parts of a plant are influenced by AM but not in the same manner. In the case of mildewed linseed the effect of AM on plant health was impressing, it indicates that AM has an ability to induce tolerance.

Lipid constituents of oil bodies in the cultured shoot primordia of Matricaria chamomilla

The oil bodies in the cultured shoot primordia of Matricaria chamomilla comprised 80% triacylglycerol, 5% diacylglycerol, 6% monoacylglycerol and 9% free fatty acids. Free sterols and steryl esters were minor components of the oil bodies. The lipid components in the oil bodies from the cultured shoot primordia were similar to those from seeds, flowers and leaves in the intact plants. Linoleic acid (18:2) and linolenic acid (18:3) were major components of fatty acid residue in the triacylglycerols in the oil bodies from the shoot primordia, whereas the content of palmitic acid (16:0) and stearic acid (18:0) was lower than that in oil bodies from the seeds, flowers and leaves of M. chamomilla. The free sterol composition varied only slightly between the oil bodies from seed or shoot primordia, but a greater variation was observed in the composition of the steryl esters, in terms of both the sterol and fatty acid moieties. The seed oil bodies contained a higher proportion of precursor sterols than those of the shoot primordia. As in the acylglycerols, linoleic and linolenic acids were the major fatty acids in the esters from shoot primordia oil bodies, whereas the steryl esters of the seed oil bodies contained a much higher proportion of saturated fatty acids.

The effects of excess exogenous mevalonic acid on sterol and steryl ester biosynthesis in celery (Apium graveolens) cell suspension cultures

Cell suspension cultures of celery ( Apium graveolens) have been used to investigate the fate of excess added mevalonic acid and the effects on the amounts of free sterols and steryl esters in the cells. Radiolabelling studies using [2- 14C] MVA showed that the main effect of excess MVA (5 mM) was an increase in the synthesis of steryl esters. Further analysis revealed that esterified 4,4-dimethylsterols were more highly labelled than 4-mono- and 4-desmethylsterols in MVA-treated cells, suggesting a restriction in the flow of carbon downstream from cycloartenol and 24-methylenecycloartanol, and thus indicating the presence of a post-mevalonic acid control point in the sterol biosynthesis pathway. The total fatty acids recovered from the celery cell lipids showed a very low incorporation of radioactivity from the [2- 14C] mevalonate, indicating that a limited breakdown of excess mevalonate to acetate may occur in these cells, possibly by the operation of the mevalonate shunt. However, this does not appear to be a major route for the disposal of excess mevalonate. Analysis of the free sterols and steryl esters showed that the administration of excess exogenous MVA (5 mM) to Apium graveolens cells caused a two-fold increase in the steryl ester content when compared with control cells, although there was little effect on the free sterol content or composition. When excess MVA was administered in conjunction with the sterol biosynthesis inhibitor paclobutrazol (50 μM), the steryl ester content increased almost three-fold. It is possible that, as well as providing a means of storage of excess sterols, the esterification of sterols may have a regulatory role in sterol biosynthesis, by controlling the rate of C-4 demethylation of precursor 4,4-dimethylsterols.

Phospholipid and free sterol composition of hypocotyl plasma membranes of ageing mung bean seedlings

Plasma membrane fractions were prepared from different hypocotyl segments (‘top’, ‘middle’ and ‘lower’ segments) of ageing Mung bean seedlings (four-seven days old) and analysed for the composition of phospholipids (headgroups) and free sterols. Effects of progressing seedling age were most striking at the base of the hypocotyl. Net phospholipid content decreased from the top to the base of the hypocotyl, and demonstrated increased susceptibility to degradation in the physiologically oldest (lower) segments. Breakdown of phosphatidylethanolamine and phosphatidylcholine resulted in increased amounts of phosphatidic acid. In top segments the membrane phospholipid composition remained essentially unchanged, which could be expected since the physiological age of these segments was identical during the four days of the experiment. Similarly, top segments did not show marked sterol changes. In middle segments, the rapid increase (2.4 times) of stigmasterol between day 4 and 5 reflected a final step in the membrane maturation process. Lower segments demonstrated a 40% decrease of both stigmasterol and sitosterol. Cholesterol and campesterol remained unchanged in all three segments.

Effects of triadimefon and osmotic stress on plasma membrane composition and ATPase activity in white spruce (Picea glauca) needles

White spruce [Picea glauca (Moench) Voss.] seedlings were grown in solution culture and treated with 20 mg I‐1 triadimefon [1‐(chlorophenoxy)‐3,3‐dimethyl‐1‐(1,2,4‐triazol‐1‐yl)‐2‐butanol] for 4 weeks and then subjected to osmotic stress with polyethylene glycol 3350. Water potentials and electrolyte leakage were measured in control and triadimefon‐treated seedlings before and after the plants were subjected to osmotic stress. The plasma membranes were isolated from needles to study their lipid composition and the activity of plasma‐membrane bound ATPase. Triadimefon treatment reduced water potentials and increased leakage of electrolytes from seedlings. However, when the seedlings were exposed to osmotic stress, triadimefon‐treated plants maintained higher water potentials and leaked less electrolytes compared with the control plants. Both triadimefon and osmotic stress treatments inhibited the activity of plasma membrane‐bound ATPase and altered the composition of free sterols in the plasma membranes. Triadimefon‐treated plants contained traces of campesterol, which was not present in control. Osmotic stress drastically reduced phospholipid:protein and sterol:protein ratios and increased sterol:phospholipid ratios in the plasma membranes

Analysis of sterols in plants and cell cultures producing ecdysteroids: I. Chenopodium album

Cell cultures have been obtained in vitro from a plant producing ecdysteroids, Chenopodium album. The composition of free and esterified sterols was compared in plant and cell cultures after purification by TLC and identification by GC/MS. Free sterols were the most abundant. A selectivity of sterol esterification was noted for 4,4-dimethylsterols (except in roots) and Δ7-sterols. The 4-desmethylsterols from leaves, roots and cell cultures of Chenopodium album were a mixture of Δ7-sterols (>61%) and Δ5-sterols (>36%). The Δ7-sterols were 24-ethylcholesta-7,22-dien-3β-ol and 24-ethylcholest-7-en-3β-ol. The isolated Δ5-sterols were sitosterol and stigmasterol. Traces of cholesterol and Δ5,7-sterols were also detected. Our attention was also given to ecdysteroid contents, which showed much lower quantity in cell cultures than in plants, confirming however the presence of 20-hydroxyecdysone.

Sterol and ecdysteroid profiles of Serratula tinctoria (L.): Plant and cell cultures producing steroids

Cell suspension cultures have been obtained from Serratula tinctoria, a plant producing ecdysteroids. Sterol profiles and ecdysteroid contents have been analysed and compared in plants and cell cultures. In particular, the composition of free and esterified sterols was investigated using appropriate analytical techniques. In plants, esterified sterols were more abundant (50–70% of the total sterol) than in cell cultures (13–36%). A selectivity for sterol esterification was noted: in plants, the triterpenes (as amyrins) were esterified, whereas it was the 4-desmethylsterols (sitosterol and cholesterol) in cell cultures. Ecdysteroids were present in higher quantities in plant (0.1–1.2% dry wt) than in cell cultures (0.01–0.03%). Analysis confirmed the presence of 20-hydroxyecdysone, 20-hydroxyecdysone-3-acetate and polypodine B as the main compounds. Roots were not only the richest organ in ecdysteroids, but also in cholesterol, lathosterol and 24-methylene-cholesterol. The results obtained suggest interesting relationships between free or esterified sterols and ecdysteroid contents.

Effects of triazole inhibitors of sterol biosynthesis on the free and esterified sterol composition of celery cell suspension cultures

AbstractTreatment of celery cell suspension cultures with paclobutrazol and three other triazoles resulted in decreased growth and an accumulation of 14α‐methylsterols in both the free sterol and the steryl ester pools, thus indicating that the triazoles were inhibiting the action of the plant obtusifoliol 14α‐demethylase system. Obtusifoliol, 14α‐methylcampesta‐8,24(241)‐dien‐3β‐ol and 14α‐methylcampest‐8‐en‐3β‐ol were the main 14α‐methylsterols to increase in the free sterol pool. In the presence of the triazoles the steryl esters became virtually depleted of sitosterol, campesterol, stigmasterol and isofucosterol, which were replaced by 14α‐methylsterols with obtusifoliol becoming the dominant esterified sterol. Treatment of the celery culture with a fourth triazole which did not have a noticeable effect on growth and caused negligible accumulation of 14α‐methylsterols in the free sterol pool nevertheless produced some enrichment of the steryl esters in obtusifoliol. The results indicate that, following triazole treatment, a rapid esterification of the accumulating 14α‐methylsterol intermediates occurs before they start to build up in significant amount in the free sterol pool.

A comparison of the effects of ?-ketotriazole, MCPA and carbaryl on the plasma membrane Mg2+-ATPase activity and lipid composition of rice shoots (Oryza sativa)

The effect of γ-ketotriazole, MCPA and carbaryl on the lipid composition and ATPase activity associated with plasma membrane fractions from rice (Oryza sativa cv Bahia) shoots was investigated. With γ-ketotriazole and MCPA treatments the relative amount of δ5-avenasterol (%) was reduced in the plasma membrane, whereas with γ-ketotriazole a reduction was also found in the sitosterol content, expressed as a percentage of the total free sterol composition. The fatty acyl chain length of phosphatidylcholine fractions from MCPA-treated plants was also reduced. The plasma membrane Mg2+-ATPase activity was only stimulated in MCPA-treated plants. No changes were observed in lipid composition or ATPase activity with carbaryl treatment.

The lipid composition, fluidity, and Mg2+-ATPase activity of rice (Oryza sativa L. cv. Bahia) shoot plasma membranes: effects of ABA and GA3

Six-day-old rice plants (Oryza sativa L., cv Bahia) were grown for 5 days more in nutrient solution culture containing 10−5 M abscisic acid (ABA) or gibberellic acid (GA3) (treated plants). Plasma membrane (PM) vesicles were isolated from the shoots of treated or control plants, and ATPase hydrolytic and proton-pumping activity, fluidity, and free sterol and phospholipid composition were determined. Both treatments resulted in modified plant growth and increases in PM fluidity. The ATPase hydrolytic activity was decreased by 25% of control values with ABA treatment and by 35% with GA3. Both treatments reduced proton-pumping by 23%. GA3 treatment reduced the relative amount (%) of stigmasterol in the PM from 38–34%, and increased Δ5-avenasterol from 4–7%. ABA and GA3 did affect the percentage composition of the fatty acyl chains of phosphatidylinositol (PI). GA3 treatment also resulted in a reduction of the total amount of PI from 73–37 mg mg−1 protein, as well as a decrease in the overall PM C16/C18 ratio and an increase in the degree of unsaturation. Our results show that there is no common link between a specific change in lipid composition and fluidity in these membranes. On the other hand, the reduction in Mg2+-ATPase activity found with both hormones suggests that there is no relationship between membrane-bound enzyme activity and membrane fluidity.

Free sterols, steryl esters, and lipid phosphorus in needles of scot's pine (Pinus sylvestris L.)

AbstractFree sterols, steryl esters, and lipid phosphorus were measured in new (current year) needles of Scot's pine during an annual cycle, and also in one‐, two‐, and three‐year‐old needles collected shortly after bud break. Sterols were identified and quantified by capillary gas chromatography and gas chromatography/mass spectrometry. Steryl esters were hydrolyzed enzymatically. Newly emerged needles contained highest amounts of free sterols and lipid phosphorus, probably reflecting increased membrane and organelle production, but low levels of steryl esters. Mature needles contained approximately equal amounts of free and esterified sterols. The molar phospholipid/free sterol ratio was 3∶1 at all the time periods studied. A dramatic increase of steryl esters was observed in the one‐, two‐, and three‐year‐old needles at times when new needles emerged. The individual free and esterified sterols were sitosterol, campesterol (presumably together with its C‐24 epimer), and cholesterol, at approximately 88, 10, and 2%, respectively. Isofucosterol, an intermediate in sitosterol biosynthesis, was present almost exclusively in newly emerged needles. Esterified sterols contained only trace amounts of isofucosterol. Shifts in favor of cholesterol and 24ζ‐methyl cholesterol occurred in the steryl esters during needle differentiation, and saturation grade of esterified fatty acids decreased. In mature needles, the composition of free sterols and steryl esters remained constant throughout the year.

Effects of terbinafine on growth, squalene, and steryl ester content of a celery cell suspension culture

The allylamine terbinafine inhibited growth of a celery ( Apium graveolens) cell suspension culture ( I 50 = 90 μM) and blocked the action of squalene epoxidase, resulting in an accumulation of squalene and a decrease in the sterol content of the cells. Celery cells were tolerant to squalene accumulation; inhibition of growth of cultures was associated with a fall in the free sterol content below about 1 μg sterol/mg dry wt of cells. At Day 14, untreated celery cells contained about 60% of the total sterol in the esterified form. However, the steryl ester pool was considerably lower in terbinafine-treated cells which may reflect an attempt to maintain the free sterol content above a threshold value. The composition of free sterols of terbinafine-treated cells was different from control cultures, suggesting that terbinafine has a second site of inhibition on the pathway to major sterols in plants.

Senescence of oat leaves: changes in the free sterol composition and enzyme activities of the plasma membrane

Plasma membrane fractions were prepared from young (10-day-old), mature (17-day-old) and senescent (10 + 7-day-old) oat leaves ( Avena sativa L.) through aqueous two-phase partitioning and analyzed for their free sterol composition. Continued leaf growth resulted in a strong increase in overall sterol content from 40 nmol · mg −1 lipid to about 120 nmol · mg −1 lipid (1.5% and 5% by wt. of total lipids respectively). This increase could be completely ascribed to increased cholesterol and stigmasterol concentrations. Kinetic parameters of vanadate-sensitive ATPase and NADH-FeCN reductase activities did not change significantly with leaf maturation. The sterol concentration increased to a lesser extent (4.2% and 4.4% by weight) in the plasma membrane of 10-day-old leaf segments that were cut and kept in light or dark for 7 more days. However, the increase in this case was completely due to a raise in Δ 7-avenasterol and 28-isofucosterol concentrations. The K m and V max values for the ATPase activity decreased significantly in these membranes. The characteristics of the FeCN 2 reductase and of glucan synthetase II activity were apparently not affected. The effect of senescence on sterol biosynthesis and on plasma membrane enzyme activities is discussed.

Steryl Lipid Content and Composition in Bell Pepper Fruit at Three Stages of Ripening

Abstract The content and sterol composition of free sterols (FS), steryl esters (SE), acylated steryl glycosides (ASG), and steryl glycosides (SG) in pericarp tissue of bell pepper fruit from three cultivars (Capsicum annuum L. cv Gator Bell, Bell Tower, and Lady Bell) were determined at three stages of ripening (mature-green, turning, and red-ripe). In each cultivar, FS were predominant at all stages of ripening, with ≈5- to 20-fold lesser amounts of SE, ASG, and SG. The proportions of the sterol conjugates (SE, ASG, and SG) varied somewhat between cultivars, but there was a consistent increase in the ratio of SG to ASG with ripening. The sterol composition of steryl lipids was quite similar in the three cultivars, and only minor changes occurred with ripening. Sitosterol was always the major sterol, followed by campesterol. Sitosterol plus campesterol comprised 89% to 95% of the total sterols in FS, ASG, and SG, and 74% to 91% in SE. Cholesterol and stigmasterol were always present as minor (1% to 4%) constituents. In general, small increases in stigmasterol and several minor, unidentified sterols occurred with ripening, at the expense of sitosterol and campesterol. These changes were greatest in SE.

Cyclopropyl Sterol and Phospholipid Composition of Membrane Fractions from Maize Roots Treated with Fenpropimorph

Maize (Zea mays L.) caryopses were grown in the presence of fenpropimorph, a systemic fungicide, for 7 days in the dark. Membrane fractions enriched, respectively, in endoplasmic reticulum, plasma membrane, and mitochondria were isolated from control and treated maize roots and analyzed for their free sterol, phospholipid, and fatty acid composition. In treated plants, the intracellular distribution of free sterols was dramatically modified both qualitatively and quantitatively. The normally occurring Delta(5)-sterols disappeared almost completely and were replaced by 9beta, 19-cyclopropyl sterols, mainly cycloeucalenol and 24-methyl pollinastanol. These new compounds were found to accumulate in all the membrane fractions in such a way that the endoplasmic reticulum-rich fraction became the richest one in free sterols instead of the plasma membrane. In contrast, the fenpropimorph treatment of maize roots was shown not to affect either the relative proportions or the amounts of the individual phospholipids, but an increase in the unsaturation index of phospholipid-fatty acyl chains of the endoplasmic reticulum-rich fraction was observed. The present data suggest that, in higher plant membranes, cyclopropyl sterols could play a structural role similar to that of the bulk of Delta(5)-sterols.

Influence of sterol-inhibiting fungicides on the free sterol and free fatty acid composition of Red Delicious and Jonathan apple cultivars

SummaryApple cvs Red Delicious and Jonathan were treated with the sterol-inhibiting fungicides (SIFs) etaconazole, fenarimol and bitertanol or triadimefon. Treatments were applied to the foliage in two years as morning or evening applications. Free sterol and free fatty acid composition of the leaves were determined 24 and 72 h after application. SIFs had no consistent effects on the free sterol content of either apple cultivar whether applied in the morning or evening or whether measured 24 or 72 h after application. Evening applications generally resulted in a decrease in unsaturated free fatty acids while morning applications resulted in increased unsaturation. The possible effects of elevated levels of unsaturated fatty acids on sterol and growth regulator synthesis is discussed in relation to the observation that SIFs may be less detrimental to sterol and growth regulator synthesis if applied in the evenings.