Free Sterols Research Articles

Characterization of sterol glucosyltransferase from Salinispora tropica CNB-440: Potential enzyme for the biosynthesis of sitosteryl glucoside

A sterol glucosyltransferase-encoded gene was isolated from Salinispora tropica CNB-440, a marine, sediment-dwelling, Gram positive bacterium that produces the potent anticancer compound, salinosporamide A. The full-length gene consists of 1284 nucleotides and encodes 427 amino acids with a calculated mass of 45.65kDa. The gene was then cloned and heterologously expressed in Escherichia coli BL21(DE3). The amino acid sequence shares 39% similarity with the glycosyltransferase from Withania somnifera, which belongs to glycosyltransferase family 1. Enzyme reactions were carried out with the various free sterols (acceptor) and NDP-sugars (donor). The purified protein only showed activity for glucosylation of β-sitosterol with UDP-D-glucose and TDP-D-glucose donors, and optimal activity at pH 7.5 and 37°C. Among these two donors, UDP-D-glucose was preferred.

A Comprehensive Characterization of Lipids in Wheat Straw

The chemical composition of the lipids in wheat straw was studied in detail by gas chromatography and mass spectrometry. Important discrepancies with the data reported in previous papers were found. The predominant lipids identified were series of long-chain free fatty acids (25% of total extract), followed by series of free fatty alcohols (ca. 20%). High molecular weight esters of long chain fatty acids esterified to long chain fatty alcohols were also found (11%), together with lower amounts of other aliphatic series, such as n-alkanes, n-aldehydes, and glycerides (mono-, di-, and triglycerides). Relatively high amounts of β-diketones (10%), particularly 14,16-hentriacontanedione, which is the second most abundant single compound among the lipids in wheat straw, were also identified. Finally, steroid compounds (steroid hydrocarbons, steroid ketones, free sterols, sterol esters, and sterol glycosides) were also found, with sterols accounting for nearly 14% of all identified compounds.

Plant Sterol Metabolism. Δ7-Sterol-C5-Desaturase (STE1/DWARF7), Δ5,7-Sterol-Δ7-Reductase (DWARF5) and Δ24-Sterol-Δ24-Reductase (DIMINUTO/DWARF1) Show Multiple Subcellular Localizations in Arabidopsis thaliana (Heynh) L

Sterols are crucial lipid components that regulate membrane permeability and fluidity and are the precursors of bioactive steroids. The plant sterols exist as three major forms, free sterols, steryl glycosides and steryl esters. The storage of steryl esters in lipid droplets has been shown to contribute to cellular sterol homeostasis. To further document cellular aspects of sterol biosynthesis in plants, we addressed the question of the subcellular localization of the enzymes implicated in the final steps of the post-squalene biosynthetic pathway. In order to create a clear localization map of steroidogenic enzymes in cells, the coding regions of Δ7-sterol-C5-desaturase (STE1/DWARF7), Δ24-sterol-Δ24-reductase (DIMINUTO/DWARF1) and Δ5,7-sterol-Δ7-reductase (DWARF5) were fused to the yellow fluorescent protein (YFP) and transformed into Arabidopsis thaliana mutant lines deficient in the corresponding enzymes. All fusion proteins were found to localize in the endoplasmic reticulum in functionally complemented plants. The results show that both Δ5,7-sterol-Δ7-reductase and Δ24-sterol-Δ24-reductase are in addition localized to the plasma membrane, whereas Δ7-sterol-C5-desaturase was clearly detected in lipid particles. These findings raise new challenging questions about the spatial and dynamic cellular organization of sterol biosynthesis in plants.

Plant phloem sterol content: forms, putative functions, and implications for phloem-feeding insects

All eukaryotes contain sterols, which serve as structural components in cell membranes, and as precursors for important hormones. Plant vegetative tissues are known to contain mixtures of sterols, but very little is known about the sterol composition of phloem. Plants are food for many animals, but plant-feeding arthropods (including phloem-feeding insets) are unique among animals in that they have lost the ability to synthesize sterols, and must therefore acquire these essential nutrients from their food, or via endosymbionts. Our paper starts by providing a very brief overview of variation in plant sterol content, and how different sterols can affect insect herbivores, including those specializing on phloem. We then describe an experiment, where we bulk collected phloem sap exudate from bean and tobacco, and analyzed its sterol content. This approach revealed two significant observations concerning phloem sterols. First, the phloem exudate from each plant was found to contain sterols in three different fractions – free sterols, sterols conjugated to lipids (acylated), and sterols conjugated to carbohydrates (glycosylated). Second, for both plants, cholesterol was identified as the dominant sterol in each phloem exudate fraction; the remaining sterols in each fraction were a mixture of common phytosterols. We discuss our phloem exudate sterol profiles in a plant physiology/biochemistry context, and how it relates to the nutritional physiology/ecology of phloem-feeding insects. We close by proposing important next steps that will advance our knowledge concerning plant phloem sterol biology, and how phloem-sterol content might affect phloem-feeding insects.

Involvement of Bicarbonate-Induced Radical Signaling in Oxysterol Formation and Sterol Depletion of Capacitating Mammalian Sperm During In Vitro Fertilization1

This study demonstrates for the first time that porcine and mouse sperm incubated in capacitation media supplemented with bicarbonate produce oxysterols. The production is dependent on a reactive oxygen species (ROS) signaling pathway that is activated by bicarbonate and can be inhibited or blocked by addition of vitamin E or vitamin A or induced in absence of bicarbonate with pro-oxidants. The oxysterol formation was required to initiate albumin dependent depletion of 30% of the total free sterol and >50% of the formed oxysterols. Incubation of bicarbonate treated sperm with oxysterol-binding proteins (ORP-1 or ORP-2) caused a reduction of >70% of the formed oxysterols in the sperm pellet but no free sterol depletion. Interestingly, both ORP and albumin treatments led to similar signs of sperm capacitation: hyperactivated motility, tyrosin phosphorylation, and aggregation of flotillin in the apical ridge area of the sperm head. However, only albumin incubations led to high in vitro fertilization rates of the oocytes, whereas the ORP-1 and ORP-2 incubations did not. A pretreatment of sperm with vitamin E or A caused reduced in vitro fertilization rates with 47% and 100%, respectively. Artificial depletion of sterols mediated by methyl-beta cyclodextrin bypasses the bicarbonate ROS oxysterol signaling pathway but resulted only in low in vitro fertilization rates and oocyte degeneration. Thus, bicarbonate-induced ROS formation causes at the sperm surface oxysterol formation and a simultaneous activation of reverse sterol transport from the sperm surface, which appears to be required for efficient oocyte fertilization.

Development of a Rapid and Simple Non-Derivatization Method to Determine Constituents and Antioxidative Capacity of Camellia Oils by HPTLC

Camellia oil is an edible vegetable oil with high value of nutrition and health protection function such as antioxidant and adjusting blood fat. In this study, a simple, rapid and effective HPTLC method was developed for analyzing the composition and antioxidant constituents of camellia oil. The HPTLC was performed on G60 plate with n-hexane-diethyl ether-acetic acid (6:4:0.1, v/v/v) as mobile phase combined with two coloration methods (ethanol containing 10% phosphomolybdic acid, ethanol containing 0.03% DPPH) and scanning densitometry technique. The unsaturated fatty glyceride, free fatty acids, sterols and lipids including triolein, oleic acid, ergosterin, β-sitosterol, tocopherol and phospholipids in camellia oils were determined and performed densitometrically at λs1 = 620 nm and λs2 = 517 nm. The results show that the main components of different samples of camellia oil are similar, however the contents are diverse. The antioxidative test shows that camellia oil has obvious antioxidant capability as olive oil, especially the pressed virgin oil. Therefore, this non-derivatization HPTLC method can be used for composition and antioxidative capacity determination of camellia oils.

Lipids in a sulfur-rich lacustrine sediment from the N^|^ouml;rdlinger Ries (southern Germany) with a focus on free and bound sterols

The distribution of free sterols in a sulfur-rich lacustrine sediment of Miocene age deposited in the ancient crater lake of the Nordlinger Ries (southern Germany) was investigated and compared with the corresponding distributions of esterified and kerogen-bound sterols. The three fractions exhibited the same suite of principal sterols, with 4α,24-dimethyl-5α- cholestan-3β-ol, dinosterol and dinostanol displaying the highest concentrations and relative abundances. The distributions of sterols, steroidal ketones, n-alkanols and isoprenoid alcohols suggest a prevalent deposition of autochthonous aquatic organic matter under saline conditions. In particular, the high abundance of 4α,24-dimethyl-5α-cholestan-3-one, dinosterone and dinostanone and the corresponding 4-methyl sterols indicates the importance of dinoflagellate productivity in this former crater lake and its significant contribution to the sedimentary lipids in the Nordlinger Ries sediment. The similarity in structures and relative abundance of the major 4-methyl sterols and the corresponding steroidal ketones suggest that they are biosynthetically linked.

Effects of Echinostoma caproni miracidia dose on the neutral and polar lipids of Biomphalaria glabrata as determined by high-performance thin-layer chromatography

The effects of a 5 versus 25 miracidia exposure of Echinostoma caproni on the lipid composition of Biomphalaria glabrata was studied using high performance thin layer chromatography (HPTLC)-densitometry. A 50 miracidia dose was not used because such a high level of exposure caused severe snail mortality by 3 weeks post-exposure (PE). Lipids were determined in the digestive-gland gonad complex (DGG) of the exposed snails and in the uninfected matched controls at 2 and 4 weeks PE. Extraction of lipids from DGGs was carried out by the Folch method with chloroform-methanol (2:1), and extracts were analyzed on Analtech HPTLC-HLF pre-adsorbent silica gel plates with measurement of separated bands using a CAMAG Scanner 3. For neutral lipids the mobile phase was petroleum ether-diethyl ether-glacial acetic acid (80:20:1) and the detection reagent was 5% ethanolic phosphoric acid, and for polar lipids chloroform-methanol-deionized water (65:25:4) mobile phase and 10% cupric sulfate in 8% phosphoric acid detection reagent were used. No significant differences in the concentrations of free sterols, free fatty acids, triacylglycerols, phosphatidylcholine, and phosphatidylethanolamine were seen at 2 weeks PE in any of the groups. At 4 weeks PE, the free fatty acid concentration increased significantly in the snails exposed to 25 miracidia compared to that of the 5 miracidia/snail group or the controls. Elevation of the free fatty acid fraction in the high dose snail group suggested that some changes occurred in the lipid metabolism of the snails in that group as a function of miracidia dose.

Influence of baking and in vitro digestion on steryl ferulates from wheat

So far, data on absorption and metabolism of steryl ferulates from edible sources is scarce. Therefore, the impact of enzyme-aided baking and in vitro digestion was examined in this study. Wheat flours and wheat breads were subjected to a static in vitro digestion model and changes in the contents of steryl ferulates and free sterols (possible hydrolysis products of steryl ferulates) were monitored. Baking degraded steryl ferulates at a similar rate in all types of breads (43–47%) compared to the corresponding flours, while baking induced changes in free sterols showed no clear pattern. In vitro digestion provoked five folds lower content of steryl ferulates in flours than in breads and it also resulted in significant free sterol accumulation. Interestingly, bioaccessibility (0.01–0.25%) was not influenced by the cereal matrix. The four steryl ferulate species, which were detected in wheat, showed similar hydrolysis rates during digestion. As baking had a significant impact on the steryl ferulate content of wheat, we suggest that both raw and processed sources should be considered further in vitro, animal or human trials, when studying the metabolic fate of steryl ferulates.

Influence of Hydrocarbons Exposure on Survival, Growth and Condition of Juvenile Flatfish: A Mesocosm Experiment

Juveniles of numerous commercial marine flatfish species use coastal and estuarine habitats as nurseries. Hence, they are likely to be exposed to a number of anthropogenic stressors such as accidental and chronic exposure to chemical contaminants. Little is known about their response to such pollutants at the individual level and about the consequences on their population dynamics. Mesocosm experiments were conducted to determine whether short (24 h) but high exposure to petroleum hydrocarbons (1/1000 v: v water: fuel), similar to what happened after an oil spill on coastal areas, affects survival and biological (growth, body condition and lipid reserve) performances of juvenile common sole, which live on near shore and estuarine nursery grounds. Results demonstrated that this type of exposure significantly reduce survival, growth (size, recent otolith increment and body condition), and especially energy storage (triacylglycerol to free sterol ratio) of the juvenile fish on the medium-term (three months after the exposure). These medium-term consequences affect future recruitment of this long- lived species.

Novel Steroidal Components from the Underground Parts of Ruscus aculeatus L.

Two new furostanol saponins 1–2 and three new sulphated glycosides 3a,b and 4 were isolated from the underground parts of Ruscus aculeatus L., along with four known furostanol and one spirostanol saponins 5–9 and three free sterols. All of the structures have been elucidated on the basis of spectroscopic data 1D and 2D NMR experiments, MS spectra and GC analyses.

Tonoplast of Beta vulgaris L. contains detergent-resistant membrane microdomains

The experiments conducted on tonoplast of Beta vulgaris L. roots were performed to identify detergent-resistant lipid-protein microdomains (DRMs, interpreted as lipid rafts).The presence of DRMs can be found when dynamic clustering of sphingolipids, sterols, saturated fatty acids is registered, and the insolubility of these microdomains in nonionic detergents at low temperatures is proven. The elucidation of tonoplast microdomains has been based on results obtained with the aid of high-speed centrifuging in the sucrose gradient. The experiments have shown that tonoplast microdomains are rich in sphingolipids, free sterols and saturated fatty acids (such a lipid content is also typical of lipid-protein microdomains of other membranes), while only few phospholipids are present in tonoplast microdomains. The presence of microdomains has been confirmed by fluorescence and confocal microscopy using filipin and Laurdan as fluorescent probes. The experiments with Laurdan have shown that tonoplast microdomains are characterized by a high order compared to characteristics of the rest of the tonoplast. Thus, the presence of detergent-resistant lipid-protein microdomains in the tonoplast has been demonstrated.

Impact of morphogenetic effectors on the growth pattern and the lipid composition of the mycelium and the yeastlike cells of the fungus Mucor hiemalis

We investigated the growth and the cell lipid composition of the mycelium and of the yeast-like form of Mucor hiemalis VKMF-1431 obtained under aerobic conditions by treatment with the morphogenetic agents itraconazole, exogenous triacylglycerols (TAGs), and trehalose. The sporangiospores of a 20-day culture were inoculated on the medium with glucose. Under these conditions, the fungus produced both mycelium and yeast-like cells. It was established that, upon the germination of old (20-day) sporangiospores, the fungus predominantly used the mycelium development strategy in the presence of trehalose and TAGs. It was characterized by a low ratio between the two bulk membrane lipids (PEA/PC) and increased levels of PC and polyunsaturated fatty acids (FA). Compared to the mycelium, the yeast cell morphotype obtained on the medium with glucose was distinguished by an elevated PEA/PC ratio, lowered TAG, free sterol (FS) and esterified sterol (ES) levels, a decreased ES/FS ratio that correlated with the reserve sterol pool size, and a lowered content of unsaturated fatty acids (the linoleic and the γ-linolenic acid). These peculiarities of the lipid composition of yeastlike cells correlated with the intensity of yeastlike growth. Light and electron microscopy revealed differences between the above cell morphotypes. With itraconazole, yeast-like cells were characterized by the destruction of the endoplasmic reticulum membranes and formation of a large number of vacuoles. The suggestion was confirmed that the state/age of inoculum sporangiospores exerts an influence on the capacity for dimorphism in mucorous fungi such as M. hiemalis. The data obtained testify to an involvement of lipids in the process of adaptation to environmental factors and to their regulatory role in morphogenetic processes associated with the formation of alternative morphotypes of the mucorous fungus.

Modulation of lipid metabolism with the overexpression of NPC1L1 in mouse liver

Niemann-Pick C1-like 1 protein (NPC1L1), a transporter crucial in intestinal cholesterol absorption, is expressed in human liver but not in murine liver. To elucidate the role of hepatic NPC1L1 on lipid metabolism, we overexpressed NPC1L1 in murine liver utilizing adenovirus-mediated gene transfer. C57BL/6 mice, fed on normal chow with or without ezetimibe, were injected with NPC1L1 adenovirus (L1-mice) or control virus (Null-mice), and lipid analyses were performed five days after the injection. The plasma cholesterol levels increased in L1-mice, and FPLC analyses revealed increased cholesterol contents in large HDL lipoprotein fractions. These fractions, which showed α-mobility on agarose electrophoresis, were rich in apoE and free cholesterol. These lipoprotein changes were partially inhibited by ezetimibe treatment and were not observed in apoE-deficient mice. In addition, plasma and VLDL triglyceride (TG) levels decreased in L1-mice. The expression of microsomal triglyceride transfer protein (MTP) was markedly decreased in L1-mice, accompanied by the reduced protein levels of forkhead box protein O1 (FoxO1). These changes were not observed in mice with increased hepatic de novo cholesterol synthesis. These data demonstrate that cholesterol absorbed through NPC1L1 plays a distinct role in cellular and plasma lipid metabolism, such as the appearance of apoE-rich lipoproteins and the diminished VLDL-TG secretion.

Triacylglyceride (TAG) profiles of integumentary lipids isolated from three bat species determined by matrix-assisted laser desorption–ionization time-of-flight mass spectrometry (MALDI–TOF MS)

Lipids secreted from sebaceous glands in bat integument may play a role in determining host pathogenicity by the fungus Geomyces destructans in white-nose syndrome (WNS). To investigate this, we have determined the triacylglycerol (TAG) profiles for three bat species: eastern red bats ( Lasiurus borealis (Müller, 1776)), evening bats ( Nycticeius humeralis (Rafinesque, 1818)), and big brown bats ( Eptesicus fuscus (Beauvois, 1796)). Neutral lipids extracted from the hair and wing tissue were fractionated by preparative thin-layer chromatography (TLC) into four major lipid bands corresponding to cholesterol, free fatty acids (FFAs), TAGs, and sterol–wax esters. Densitometry showed higher proportions of TAGs in hair than from wing tissue. TAG bands were recovered and analyzed by matrix-assisted laser desorption–ionization time-of-flight mass spectrometry (MALDI–TOF MS). Mass spectra showed sodiated TAG species with variable fatty acyl (FA) moieties range from m/z 715.6–911.8. High intensity ion peaks were consistent with 16:0 and 18:1 as dominant FA moieties, and these were identified as palmitic and oleic acids, respectively, by liquid chromatography electrospray ionization mass spectrometry. We determined significant differences in TAG profiles between three bat species by MALDI–TOF MS, providing the first description of integumentary lipids in bats. In this study, we performed the first TAG profiling of bats, which suggest such profiles may be species-specific in bats.

Influence of harvest year, cultivar and geographical origin on Greek extra virgin olive oils composition: A study by NMR spectroscopy and biometric analysis

Two hundred twenty-one extra virgin olive oils (EVOO) were extracted from four olive mono-cultivars (Koroneiki, Tsounati, Adramitini, and Throubolia) originated from four divisions of Greece (Peloponnesus, Crete, Zakynthos, and Lesvos) and collected in five harvesting periods (2002–2006 and 2007–2008). All samples were chemically analysed by means of 1H and 31P NMR spectroscopy and characterised according to their content in fatty acids, phenolic compounds, diacylglycerols, total free sterols, free acidity, and iodine number. The influence of cultivars on the compositional data of the EVOO samples according to harvest year and geographical origin was examined by means of the forward stepwise canonical discriminant analysis (CDA) and classification binary trees (CBT). The CDA, when the a priori grouping was in accordance with harvest, was high (94%), whereas the classification in terms of groups formed by inclusions of geographical origin was reduced to 85%. Inclusion of both the harvesting year and geographical origin in the CDA analysis resulted in a high classification (90%) for the EVOO samples grouped into the four cultivars. The variables that most satisfactorily classified the Greek olive oils were the phenolics p-coumaric acid, pinoresinol, 1-acetoxypinoresinol, syringaresinol, luteolin, apigenin, and the hydrolysis products of oleuropein expressed collectively by the concentration of total hydroxytyrosol. Amongst the fatty acids, linoleic acid was the predictor with the highest discriminatory power. Finally, the phylogenetic significance of the olive oil compounds as determined by NMR was investigated by estimating their support to monophyly of cultivars.

Synthesis and biological evaluation of some novel 1-indanone thiazolylhydrazone derivatives as anti-Trypanosoma cruzi agents

A series of novel 4-arylthiazolylhydrazones (TZHs) derived from 1-indanones were synthesized in good yields (66–92%) in a simple procedure using microwave irradiation and then characterized by spectroscopy studies. The compounds were evaluated for their in vitro anti-Trypanosoma cruzi activity against the epimastigote, trypomastigote and amastigote forms of the parasite. Most TZHs displayed excellent activity, and were more potent and selective than the reference drug Benznidazole, used in the current chemotherapy. Analysis of the free sterols from parasite incubated with the compounds showed that inhibition of ergosterol biosynthesis is a possible target for the action of these new TZHs. In particular, TZH 9 emerged as a promising antichagasic compound to be evaluated in animal models.

Free sterols and steryl glycosides in sunflower seeds with high phytosterol contents

AbstractIn this paper, the distribution of sterol compounds has been analyzed in 10 samples of new sunflower seeds with high contents of phytosterols. The sunflower seeds were first extracted with hexane and the residual oil was then extracted with a mixture of chloroform:methanol (2:1). In addition, the lipids from the defatted seeds and from the total seeds were analyzed after acid hydrolysis and subsequent extraction. The evaluation of the sterol compounds was performed by high temperature GC directly in the sylanized unsaponifiable matter using α‐cholestanol as internal standard. This procedure allows for the quantification of free sterols and steryl glycosides in the oils extracted by hexane and chloroform:methanol (2:1) while in defatted meal and total seeds, they are determined as free sterols. The main characteristics of the fractions extracted by hexane were a variable oil content (19.8–38.7%) with medium unsaponifiable matter (1.4–4.8%), The major sterol compounds in the oil extracted by hexane were free sterols (4034–8507 mg/kg) while steryl glycosides were in low concentrations (80–280 mg/kg). On the contrary, the fraction extracted by chloroform:methanol had low lipid content (2.2–6.7%) with a high unsaponifiable matter (12.6–28.2%). The major sterol compounds in this fraction were steryl glycosides (9711–22 514 mg/kg). Finally, a third of the total sterols remained in the meal after the extraction of edible oil by hexane.

Phytosterols in pumpkin seed oil extracted by organic solvents and supercritical CO2

AbstractPhytosterols in Cucurbita pepo convar. citrullina seed oil were analyzed by GC–MS after extraction by the organic solvents, hexane and petroleum ether, and by supercritical carbon dioxide at 400 bar and 40°C. Desmosterol, campesterol, stigmasterol, β‐sitosterol, spinasterol, Δ7,22,25‐stigmastatrienol, Δ7‐stigmastenol, Δ7,25‐stigmastadienol, and Δ7‐avenasterol were identified mass spectrometrically via detection of the parent molecular ions and fragmentation patterns of corresponding trimethylsilyl derivatives. The used derivatizing agents N‐methyl‐N‐(trimethylsilyl)trifluoroacetamide (MSTFA) and N,O‐bis(trimethylsilyl)trifluoroacetamide (BSTFA) are equally effective for phytosterol content determination. As expected, the predominant phytosterols were Δ7‐sterols that ranged from 91.0 to 94.2% of the total phytosterol content given as sum of free sterols and steryl esters. Comparison of extraction methods revealed that, although the oil yields obtained by extraction with hexane, petroleum ether and supercritical CO2 were 43.37, 44.65, and 36.17%, respectively, the total phytosterol content in supercritical CO2 extract (294 mg/100 g oil, for both MSTFA and BSTFA) was about 30% higher than in hexane and about 20% higher than in petroleum ether extract.Practical applications: Extraction with the application of fluids in supercritical state opens a new approach to the production of edible vegetable oils. Many economical, health, and environmental aspects are interlaced in this modern approach. By applying the supercritical extraction, the selectivity of the extraction can be significantly improved compared to traditional methods of extraction. Thus, depending on the operating conditions of supercritical CO2 extraction higher total phytosterol content can be achieved resulting in improved nutritional value of the pumpkin seed oil.

Lipid droplet formation on opposing sides of the endoplasmic reticulum

In animal cells, the primary repositories of esterified fatty acids and alcohols (neutral lipids) are lipid droplets that form on the lumenal and/or cytoplasmic side of the endoplasmic reticulum (ER) membrane. A monolayer of amphipathic lipids, intermeshed with key proteins, serves to solubilize neutral lipids as they are synthesized and desorbed. In specialized cells, mobilization of the lipid cargo for delivery to other tissues occurs by secretion of lipoproteins into the plasma compartment. Serum lipoprotein assembly requires an obligate structural protein anchor (apolipoprotein B) and a dedicated chaperone, microsomal triglyceride transfer protein. By contrast, lipid droplets that form on the cytoplasmic face of the ER lack an obligate protein scaffold or any required chaperone/lipid transfer protein. Mobilization of neutral lipids from the cytosol requires regulated hydrolysis followed by transfer of the products to different organelles or export from cells. Several proteins play a key role in controlling droplet number, stability, and catabolism; however, it is our premise that their formation initiates spontaneously, solely as a consequence of neutral lipid synthesis. This default pathway directs droplets into the cytoplasm where they accumulate in many lipid disorders.