Amounts Of Neutral Lipids Research Articles

Cheese whey as a renewable substrate for microbial lipid and biomass production by Zygomycetes

AbstractThree Zygomycetes, Mortierella isabellina, Thamnidium elegans and Mucor sp., were tested for their ability of producing biomass and lipid‐containing γ‐linolenic acid (GLA) during their cultivation on cheese whey. M. isabellina consumed all of the available lactose and a significant amount of the available protein. On the contrary, the two other fungi seemed incapable of consuming lactose after protein exhaustion. In the second series of experiments, for M. isabellina a supplementary quantity of lactose was added into the medium in order to increase the C/N ratio and hence to increase the production of fat. In the case of T. elegans and Mucor sp., a supplementary quantity of ammonium sulfate was added in order to favor the consumption of lactose and the production of biomass. Indeed, enhancement of lipid production was observed for M. isabellina and biomass production for T. elegans and Mucor sp.. Fatty acid analysis of the microbial lipid showed a composition that presented non‐negligible changes in relation with the age of the culture and the C/N molar ratio of the medium. Further analysis of the fat showed that the quantity of neutral lipids was the more abundant. The fatty acid composition of neutral lipids resembled to that of total lipids. Phospholipids were the more unsaturated fraction for Mucor sp. and M. isabellina. GLA was synthesized in all trials but its concentration presented differences related with the utilized strains and the fermentation time. Growth of M. isabellina on lactose‐supplemented whey resulted in a maximum GLA production of 301 mg/L.

Extraction and analysis of neutral lipids from activated sludge with and without sub-critical water pre-treatment

Activated sludge with its high lipid content is a potential raw material for producing biodiesel. Sub-critical water (SCW) treatment has been known to be a cheap and effective method for reducing organic compounds, especially carcinogenic aromatic compounds such as polycyclic aromatic hydrocarbons or polychlorobiphenyls in activated sludge as well as increasing the amount of extractable lipids. In this work, the amount of neutral lipids and fatty acid profiles in neutral lipids extracted from activated sludge both with and without SCW pre-treatment were investigated. Results showed that the amount of neutral lipid extracted from SCW treated activated sludge is almost four times to those without SCW treatment. However, there is no observable difference between the fatty acid profiles of neutral lipids obtained from activated sludge with and without SCW treatment. At least 14 kinds of fatty acids were identified in the neutral lipids of sludge oil.

Neutral Lipid Characterization of Non‐Water‐Soluble Fractions of Carica Papaya Latex

AbstractThe non‐water‐soluble fraction of Carica papaya latex (CPL) constitutes a waste material from papain production; very little information exists regarding its chemical composition. The non‐water‐soluble fraction of CPL was fractionated by liquid chromatography into neutral lipids, glycolipids and phospholipids. The most abundant compounds were found to be the polar lipids, accounting for 79.2% (w/w) of the total extractible matter, while the total amount of neutral lipids was only around 20%. It was composed of free fatty acids, sterols and triterpenic alcohols, but no glycerides were detected. A high content of saturated fatty acids was measured; these saturated fatty acids were represented by very long chains with C24:0, C26:0 and C28:0 accounting for 6.3, 11.0 and 6.3%, respectively, in the total extractible matter and 7.3, 9.0 and 3.9% in the FFA fraction. The monounsaturated fatty acids were about 23–25% in both samples, with oleic acid (C18:1) being the most abundant. The polyunsaturated fatty acids that were 25.1% in the total matter and 21.6% in the FFA fraction were mainly represented by linoleic acid (C18:2n‐6). Finally, a very interesting characteristic of the FA composition of this latex concerns the presence of odd‐numbered fatty acids in significant amounts (around 22% in the total extract and 24.3% in the FFA fraction).

Fat‐Soluble Bioactives, Fatty Acid Profile and Radical Scavenging Activity of Semecarpus anacardium Seed Oil

AbstractSemecarpus anacardium (family Anacardiaceae) has many applications in the Ayurvedic and Siddha systems of medicine in India. Detailed knowledge on the composition of S. anacardium oil, in consideration of potential utilization, is of major importance. In this investigation, column chromatography, gas chromatography, thin layer chromatography and liquid chromatography techniques were performed to analyze lipid classes, fatty acids and fat‐soluble bioactives of S. anacardium crude seed oil. The amount of neutral lipids in the crude seed oil was the highest, followed by glycolipids and phospholipids, respectively. Linoleic followed by palmitic and oleic were the major fatty acids. The ratio of unsaturated fatty acids to saturated fatty acids was higher in neutral lipid classes than in the polar lipids. The main sterol compounds were β‐sitosterol, campesterol and stigmasterol. δ‐Tocopherol followed by β‐tocopherol were the main tocopherols. When S. anacardium seed oil and extra virgin olive oil were compared for their radical scavenging activity toward 1,1‐diphenyl‐2‐picrylhydrazyl radical and galvinoxyl radical (by electron spin resonance spectrometry), S. anacardium seed oil exhibited a stronger RSA.

Lipid, fatty acid and tocol distribution of coriander fruit's different parts

High amounts of neutral lipids (NL) were found (95.65% of total lipids) in whole fruit oil, while glycolipids (GLs) occurred at high levels in pericarp oil (73.21% of total lipids). Triacylglycerol was found to be the principal lipid fraction of NL which formed 93% in seed and whole fruit. However, monoacylglycerol constituted the major fraction of NL in pericarp (34.5%). At least two GLs and five phospholipids (PL) were identified. The GL composition of the pericarp is characterized by monogalactosylacylglycerol as the major fraction with a percentage of 53.39% of total lipids. The major individual PL subclasses were phosphatidylcholine followed by phosphatidylethanolamine in seed and whole fruit. However, the PL were not detected in pericarp oil. The principal fatty acids identified in most lipid classes were petroselinic acid (C18:1 n − 12), linoleic acid (C18:2 n − 6), palmitic (C16:0) and oleic acid (C18:1 n − 9). The total tocopherol and tocotrienol (tocol) contents were 27.78 mg/100 g oil in whole fruit, 26.42 mg/100 g oil in seed and 5.36 mg/100 g oil in pericarp. Fruit and seed oils were characterized by a high amount of γ-tocotrienol with 19.56 mg/100 g oil. However, α-tocopherol (1.82 mg/100 g oil) was found to be the tocol marker in pericarp oils. The results are important as an indication of the potentially economical utility of Coriandrum sativum L. seed oil as a new source of PL.

Characteristics of Oil from Hulless Barley (Hordeum vulgare L.) Bran from Tibet

AbstractThe bran of hulless barley (Hordeum vulgare L.) from Tibet was investigated. This paper reports on the physicochemical characteristics, lipid classes and fatty acids of the oil from the bran. The petroleum (60–90 °C) extract of hulless barley bran was found to be 8.1%. The investigated physiochemical parameters included density at 40 °C (0.96 g/cm3), refractive index at 40 °C (1.41), melting point (30.12 °C), acid value (11.6 mg KOH/g), peroxide value (19.41 μg/g), saponification value (337.62 mg KOH/g), iodine value (113.51 mg iodine/g) and unsaponifiable matter (4.5% of total lipids).The amount of neutral lipids in the crude oil was the highest (94.55% of total lipids), followed by glycolipids (4.20% of the total lipid) and phospholipids (1.25% of the total lipid). Linoleic acid (75.08% of total fatty acids) followed by palmitic acid (20.58% of total fatty acids), were the two major fatty acids in the oil. The results show that the oil from the hulless barley bran could be a good source of valuable essential fatty acids.

Fatty acids, bioactive lipids and radical scavenging activity of Celastrus paniculatus Willd. seed oil

In the present contribution lipid composition and characteristics of Indian Celastrus paniculatus seed oil were determined. C. paniculatus seeds are a rich source of oil (46%). The amounts of neutral lipids in the oil were the highest, followed by glycolipids and phospholipids. Oleic followed by palmitic and linoleic were the major fatty acids in C. paniculatus seed oil and its lipid classes. C. paniculatus seed oil being characterized by a relatively high amount of phytosterols, wherein the sterol marker was β-sitosterol followed by campesterol and stigmasterol. γ-Tocopherol was the major tocopherol isomer while the rest being α-tocopherol. When C. paniculatus oil and extra virgin olive oil were compared upon their radical scavenging activity (RSA) toward the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical C. paniculatus oil exhibited stronger RSA. In consideration of potential utilization, detailed knowledge on the composition of C. paniculatus oil is of major importance. The diversity of applications to which C. paniculatus oil can be put gives this plant great industrial importance.

Simulation of Self-Assembly of Cationic Lipids and DNA into Structured Complexes

We present a comprehensive set of simulations that elucidates several features of experimentally observed self-assembled structures in solutions of DNA and mixtures of neutral and cationic lipids. Our simulations are based on the Noguchi-Takasu implicit-solvent coarse-grained model of lipids as rigid trimer molecules [Noguchi, H.; Takasu, M. Phys. Rev. E 2001, 64, 041913.]. This model is extended in our work so that a certain fraction, varphi(c), of the lipids carries +1e charge and DNA molecules are introduced as uniformly charged rods. The simplified coarse-grained modeling approach provides a feasible way to study the long time-scale dynamics associated with the evolution of mesoscopically large complexes from initially disordered systems. Our simulations show that, depending on the rigidity parameter kappa(s) which governs the stiffness of the membranes, both lamellar and inverted hexagonal complexes are formed at intermediate charged lipid densities. Disordered structures are formed both when large amounts of neutral lipids (small varphi(c)) are introduced which leads to the "erosion" of the spatial order, as well as for large charge densities that result in membrane rupture. A novel phase, where DNA rods and cylindrical micelles form a two-dimensional square lattice analogous to the three-dimensional cubic NaCl-type structure, is observed in the large varphi(c) regime for very soft membrane material (kappa(s) = 0).

Extranodal Rosai-Dorfman disease of the kidney

Extranodal Rosai-Dorfman disease of the kidney

Structural and Biochemical Properties of Lipid Particles from the Yeast Saccharomyces cerevisiae

The two most prominent neutral lipids of the yeast Saccharomyces cerevisiae, triacylglycerols (TAG) and steryl esters (SE), are synthesized by the two TAG synthases Dga1p and Lro1p and the two SE synthases Are1p and Are2p. In this study, we made use of a set of triple mutants with only one of these acyltransferases active to elucidate the contribution of each single enzyme to lipid particle (LP)/droplet formation. Depending on the remaining acyltransferases, LP from triple mutants contained only TAG or SE, respectively, with specific patterns of fatty acids and sterols. Biophysical investigations, however, revealed that individual neutral lipids strongly affected the internal structure of LP. SE form several ordered shells below the surface phospholipid monolayer of LP, whereas TAG are more or less randomly packed in the center of the LP. We propose that this structural arrangement of neutral lipids in LP may be important for their physiological role especially with respect to mobilization of TAG and SE reserves.

Lipoprotein lipase (LPL) is highly expressed and active in the ovary of European sea bass ( Dicentrarchus labrax L.), during gonadal development

The oocytes of many fish species accumulate high amounts of neutral lipids as a caloric reserve for embryonic and larval development. We propose that lipoprotein lipase (LPL, EC 3.1.1.34) plays an important role in supplying the oocytes with fatty acids and we have cloned its cDNA from the ovary of sea bass, and determined the patterns of LPL activity and LPL mRNA expression in the ovary. The cDNA obtained was 3051 bp long with an open reading frame encoding 518 amino acids. The amino acid sequence has a high similarity and shows similar structural features to LPL of other species. Northern blot analysis revealed LPL expression in adipose tissue and gonads only. LPL activity and LPL mRNA expression in the ovary was very high in fish with a gonadosomatic index (GSI) above 5, coinciding with the appearance of a high number of lipid droplets in the ooplasm. The LPL mRNA expression was localised to the follicle cells surrounding the oocyte. Our results suggest that LPL is likely to play an important role in the incorporation of neutral lipids into the oocytes, and that follicle cells, in addition to participating in steroidogenesis, also may be important in building up oocyte lipid reserves.

Cytokine response to lipoprotein lipid loading in human monocyte-derived macrophages

BackgroundMacrophage foam cell formation is a prominent feature of human atherosclerotic plaques, usually considered to be correlated to uptake of and inflammatory response to oxidized low density lipoproteins (OxLDL). However, there are alternative pathways for formation of macrophage foam cells and the effect of such lipid loading on macrophage function remains to be fully characterized. In the present study we investigated basal and inducible cytokine expression in primary human macrophages either loaded with triglycerides through incubation with very low density lipoproteins (VLDL) or with cholesterol through incubation with aggregated LDL (AgLDL). We then analyzed how foam cell lipid content affected secretion of three pro-inflammatory cytokines: interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), and of one chemokine: interleukin-8 (IL-8), all of which are considered pro-inflammatory, pro-atherosclerotic, and are expressed by cells in atherosclerotic tissue.ResultsFormation of triglyceride-loaded foam cells resulted in a four-fold increase in basal IL-1β secretion, whereas cholesterol loading lacked significant effect on IL-1β secretion. In contrast, secretion of TNF-α and IL-6 decreased significantly following both cholesterol and triglyceride loading, with a similar trend for secretion of IL-8. Lipid loading did not affect cell viability or expression of caspase-3, and did not significantly affect macrophage ability to respond to stimulation with exogenous TNF-α.ConclusionLipid loading of primary human macrophages resulted in altered cytokine secretion from cells, where effects were similar regardless of neutral lipid composition of cells. The exception was IL-1β, where triglyceride, but not cholesterol, lipid loading resulted in a stimulation of basal secretion of the cytokine. It is apparent that macrophage cytokine secretion is affected by lipid loading by lipoproteins other than OxLDL. As both VLDL and AgLDL have been found in the vessel wall, macrophage cytokine response to uptake of these lipoproteins may have a direct effect on atherosclerotic development in vivo. However, macrophage neutral lipid amount and composition did not affect cellular activation by exogenous TNF-α, making it likely that lipoprotein lipid loading can affect foam cell cytokine secretion during basal conditions but that the effects can be overruled by TNF-α during acute inflammation.

Loss of Glial Lazarillo, a Homolog of Apolipoprotein D, Reduces Lifespan and Stress Resistance in Drosophila

The vertebrate Apolipoprotein D (ApoD) is a lipocalin secreted from subsets of neurons and glia during neural development and aging . A strong correlation exists between ApoD overexpression and numerous nervous system pathologies as well as obesity, diabetes, and many forms of cancer . However, the exact relationship between the function of ApoD and the pathophysiology of these diseases is still unknown. We have generated loss-of-function Drosophila mutants for the Glial Lazarillo (GLaz) gene , a homolog of ApoD in the fruit fly, mainly expressed in subsets of adult glial cells. The absence of GLaz reduces the organism's resistance to oxidative stress and starvation and shortens male lifespan. The mutant flies exhibit a smaller body mass due to a lower amount of neutral lipids stored in the fat body. Apoptotic neural cell death increases in aged flies or upon paraquat treatment, which also impairs neural function as assessed by behavioral tests. The higher sensitivity to oxidative stress and starvation and the reduced fat storage revert to control levels when a GFP-GLaz fusion protein is expressed under the control of the GLaz natural promoter. Finally, GLaz mutants have a higher concentration of lipid peroxidation products, pointing to a lipid peroxidation protection or scavenging as the mechanism of action for this lipocalin. In agreement with Walker et al. (, in this issue of Current Biology), who analyze the effects of overexpressing GLaz, we conclude that GLaz has a protective role in stress situations and that its absence reduces lifespan and accelerates neurodegeneration.

Profile and levels of fatty acids and bioactive constituents in mahua butter from fruit-seeds of buttercup tree [Madhuca longifolia (Koenig)

Madhuca longifolia Syn. M. indica (Sapotaceae) is an important economic tree growing throughout the subtropical region of the Indo-Pak subcontinent. Information regading the exact composition of mahua butter (also known as mowrah butter) from fruit-seeds of buttercup or mahua tree (Madhuca longifolia) is scare. In the present contribution, lipid composition and characteristics of mahua butter from the Indian buttercup tree were determined. A combination of different chromatographic techniques were performed to analyze lipid classes, fatty acids, and fat-soluble bioactives of mahua fat. n-Hexane extract of finely-ground buttercup fruit-seed meal was found to be about 58%. This observation confirmed the fact that the buttercup fruit-seeds is a rich source of fat. The extracted fat was yellow in color and solid at room temperature. The amount of neutral lipids in the crude mahua fat was the highest (ca. 95.4% of total lipids), followed by glycolipids and phospholipids, respectively. Oleic, stearic, palmitic, and linoleic were the major fatty acids in mahua fat and its’ lipid classes. The ratio of unsaturates to saturates was higher in the neutral lipid subclasses than in the polar lipid fractions. Mahua butter being characterized by a relatively high amount of phytosterols, wherein the sterol markers were Δ5-avenasterol and β-sitosterol. γ-Tocopherol was the major tocopherol isomer while the rest being β and α isomers. When mahua butter and extra-virgin olive oil were compared upon their radical scavenging activity (RSA) toward the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, mahua butter exhibited stronger RSA. In consideration of potential utilization, detailed knowledge on the composition of mahua butter is of major importance. The diversity of applications to which mahua butter can be put gives this substance great industrial importance.

Characterisation of fatty acids and bioactive compounds of kachnar ( Bauhinia purpurea L.) seed oil

Information concerning the exact composition of kachnar ( Bauhinia purpurea) seed oil is scare. In the present contribution, a combination of CC, GC, TLC and normal-phase HPLC were performed to analyse lipid classes, fatty acids and fat-soluble bioactives of kachnar seed oil. n-Hexane extract of kachnar oilseeds was found to be 17.5%. The amount of neutral lipids in the crude seed oil was the highest (ca. 99% of total lipids), followed by glycolipids and phospholipids, respectively. Linoleic, followed by palmitic, oleic and stearic, were the major fatty acids in the crude seed oil and its lipid classes. The ratio of unsaturated fatty acids to saturated fatty acid, was higher in neutral lipid classes than in the polar lipid fractions. The oil was characterised by a relatively high amount of phytosterols, wherein the sterol markers were β-sitosterol and stigmasterol. β-Tocopherol was the major tocopherol isomer with the rest being δ-tocopherol. In consideration of potential utilisation, detailed knowledge of the composition of kachnar ( B. purpurea) seed oil is of major importance.

Respiratory and metabolic responses during exercise and skeletal muscle morphology in obesity

Obese subjects need additional energy to move larger bodies and a greater amount of O2 to accomplish an equal external work-load than lean subjects. Moreover, obesity is characterized by a marked insulin resistance, which may modify metabolic responses to physical exercise. We measured the respiratory and metabolic responses to a progressive ergometric test in a group of 10 men obese and in 10 lean controls. Muscle biopsies of the left quadriceps were performed to study correlations between metabolic responses and muscle morphology. Oxygen consumption, carbon dioxide production and the respiratory exchange ratio (RER) were assessed, as were plasma concentrations of K+ and catecholamines in the basal state (rest) and during the last 30 s of each work load at exercise. Lactic acid was also measured. Obese subjects achieved a lower maximal power output (129 vs. 140 W) and a significantly lower anaerobic threshold (82 vs. 114 W, p<0.05). RER was significantly lower in the obese group, and catecholamines rose earlier and with a lower peak in correspondence of maximal power outputs. K+ and lactic acid levels increased in both groups, although significantly less in obese subjects. The main feature of muscle specimens from obese subjects was hypertrophy of the fibers, present in 9 of 10 cases, and increased amounts of neutral lipids (Oil Red O staining), present in 7 of 10 individuals. Low RER in obese subjects indicated a condition of insulin resistance, which persisted during physical activity. We conclude that fats are the fuel preferred by obese individuals during strenuous exercise.

Differential temperature regulation of three sunflower microsomal oleate desaturase (FAD2) isoforms overexpressed in Saccharomyces cerevisiae

AbstractThe oxygen‐independent temperature regulation of three sunflower microsomal oleate desaturase (FAD2) isoforms has been investigated using Saccharomyces cerevisiae cells expressing each FAD2 gene. Yeast cells transformed with the FAD2‐1 gene showed the highest percentage of dienoic acids when they were grown at 10–15 °C. In contrast, the maximal level of dienoic acids for S. cerevisiae cells expressing the FAD2‐2 and FAD2‐3 genes were obtained at 30 and 35 °C, respectively. Temperature shifts in the phase of exponential growth, from 30 to 15 °C or from 15 to 30 °C, produced changes in the final percentage of dienoic acids, mostly in yeast cells transformed with the FAD2‐1 gene, to reach the content corresponding to the new temperature. Low temperature (15 °C) increased the amount of neutral lipids in all transformed yeast cells, mainly because it favored triacylglycerol accumulation. In addition, the FAD2‐expressing yeast cells showed a higher polar lipid content than those transformed with the empty vector. Dienoic acids were present in all lipids, although high temperature (30 °C) favored their accumulation in neutral lipids. As the main conclusion, the low thermal stability observed for the major and seed specific isoform (FAD2‐1) is the key factor controlling the direct temperature regulation in sunflower seeds.

Understanding the trophic role of the Antarctic ctenophore, Callianira antarctica, using lipid biomarkers

To better understand the trophic role of ctenophores in Antarctica during austral fall and winter, a major species of cydippid ctenophore, Callianira ant- arctica, was collected during April/May (fall) and August/September (winter) 2002 in the vicinity of Marguerite Bay. Lipid content, lipid classes, fatty acids, fatty alcohols and sterols were analyzed in animals, to- gether with lipid biomarkers in krill and copepod species representing potential ctenophore prey. Lipid content in ctenophores collected in winter was slightly higher than from animals in fall (4.8 and 3.5% of dry weight, respectively). Polar lipids were the dominant lipid class in ctenophores, accounting for over half of the lipid content, with significant amounts of free fatty alcohols (more than 10% of total lipid content) detected. Lipid- class composition, however, differed significantly be- tween seasons, with significant amounts of neutral lipid (wax esters and triacylglycerols) only detected in animals from fall. Although the dominant lipid classes in cte- nophores varied between fall and winter, individual lipids (i.e., fatty acids, alcohols and sterols) showed only minor changes between seasons. Specifically, long-chain polyunsaturated fatty acids (20:5(n-3) and 22:6(n-3)) found in high abundance in larval krill were also ele- vated in ctenophores collected in winter. Very high amounts of monounsaturated fatty alcohols, particu- larly 20:1(n-9) and 22:1(n-11), known to be important components of wax esters in calanoid copepods, were also observed. Multivariate analysis using the suite of lipids found indicated that copepods are an important diet item for ctenophores in the study area. Results further suggest that C. antarctica feed actively year- round, with larval krill providing a food resource during austral winter.

The Saccharomyces cerevisiae alcohol acetyl transferase Atf1p is localized in lipid particles.

The yeast alcohol acetyl transferase I, Atf1p, is responsible for the major part of volatile acetate ester production in fermenting Saccharomyces cerevisiae cells. Some of these esters, such as ethyl acetate and isoamyl acetate, are important for the fruity flavours of wine, beer and other fermented beverages. In order to reveal the subcellular localization of Atf1p and further unravel the possible physiological role of this protein, ATF1::GFP fusion constructs were overexpressed in brewer's yeast. The transformant strain showed a significant increase in acetate ester formation, similar to that of an ATF1 overexpression strain, indicating that the Atf1p-GFP fusion protein was active. UV fluorescence microscopy revealed that the fusion protein was localized in small, sphere-like organelles. These organelles could be selectively stained by the fluorescent dye Nile red, indicating that they contained high amounts of neutral lipids and/or sterols, a specific characteristic of yeast lipid particles. Purification of lipid particles from wild type and ATF1 deletion cells confirmed that the Atf1p-GFP fusion protein was located in these organelles. Furthermore, a clear alcohol acetyl transferase activity could be measured in the purified lipid particles of both wild type and transformed cells. The localization of Atf1p in lipid particles may indicate that Atf1p has a specific role in the lipid and/or sterol metabolism that takes place in these particles.

Cationic lipid-DNA complexes for gene therapy: understanding the relationship between complex structure and gene delivery pathways at the molecular level.

Cationic liposomes (CLs) are used as gene vectors (carriers) in worldwide human clinical trials of non-viral gene therapy. These lipid-gene complexes have the potential of transferring large pieces of DNA of up to 1 million base-pairs into cells. As our understanding of the mechanisms of action of CL-DNA complexes remains poor, transfection efficiencies are still low when compared to gene delivery with viral vectors. We describe recent studies with a combination of techniques (synchrotron x-ray diffraction for structure determination, laser-scanning confocal microscopy to probe the interactions of CL-DNA particles with cells, and luciferase reporter-gene expression assays to measure transfection efficiencies in mammalian cells), which collectively are beginning to unravel the relationship between the distinctly structured CL-DNA complexes and their transfection efficiency. The work described here is applicable to transfection optimization in ex vivo cell transfection, where cells are removed and returned to patients after transfection. CL-DNA complexes primarily form a multilayered sandwich structure with DNA layered between the cationic lipids (labeled L(alpha)(C)). On rare occasions, an inverted hexagonal structure with DNA encapsulated in lipid tubules (labeled H(II)(C)) is observed. A major recent insight is that for L(alpha)(C) complexes the membrane charge density sigma(M) of the CL-vector, rather than the charge of the cationic lipid alone, is a key universal parameter that governs the transfection efficiency of L(alpha)(C) complexes in cells. The parameter sigma(M) is a measure of the average charge per unit area of the membrane, thus taking into account the amount of neutral lipids. In contrast to L(alpha)(C) complexes, H(II)(C) complexes containing the lipid 1,2-dioleoyl-sn-glycerophosphatidylethanolamine (DOPE) exhibit no dependence on sigma(M). The current limiting factor to transfection by cationic lipid vectors appears to be the tight association of a fraction of the delivered exogenous DNA with cationic cellular molecules, which may prevent optimal transcriptional activity. Future directions are outlined, which make use of surface-functionalized CL-DNA complexes suitable for transfection in vivo.