Nitroxide spin labeled analogs of the non-ionic detergent triton X-100

Two nitroxide spin labeled analogs of the non-ionic detergent, Triton X-100, have been synthesized. Electron spin resonance spectra of these compounds in solution, in egg lecithin multilayers, and in aqueous dispersion of dimyristoylphosphatidylcholine vesicles are described.     

Studies on the cloud point and micellar phases of triton X-100 using 220 MHz proton magnetic resonance techniques

High-resolution proton magnetic resonance techniques at 220 MHz were employed to follow the transformation of Triton X-100 between its micellar and cloud point phases as a function of temperature. The results obtained suggest that while a phase separation occurs rather sharply above the cloud point, the increase in temperature below the cloud point is accompanied by the gradual formation of very large structures suspended in the aqueous phase. The proton magnetic resonance studies show that the separation of phases, which occurs above the cloud point, appears to be accompanied by a fractionation of the polydisperse detergent. In addition, a lowering of the cloud point of Triton X-100 by dipalmitoyl phosphatidylcholine was observed by visual means and the results are reported here.     

Inhibition of tumor cell growth by triton X-100 through specific effects on cell-cycle-regulatory components

A cross-linked form of the detergent Triton X-100, called Triton WR-1339, has been shown to reduce the spread of tumor cells in laboratory animals. However, some of these effects were controversial, probably due to the use of different tumor cell lines and varying sites of injection. In order to better understand these processes, we have used Triton X-100 and performed a molecular analysis of its growth-inhibitory function. Using the T24 bladder carcinoma cell line, we have shown that treatment of cells with this detergent caused a potent antiproliferative effect resulting from the downregulation of the key cell cycle regulators, the cyclin-dependent kinases (CDKs). CDK activity was lost due to a twofold effect, the increased expression of the CDK inhibitors p21^Cip1 and p27^Kip1 in combination with the reduced expression of cyclin A, a regulatory CDK subunit that is essential for CDK function. Taken together, our results provide a molecular basis for the antiproliferative effects of the Triton detergent, namely its differential effects on various parts of the cell cycle machinery.     

Spectroscopic properties of protochlorophyll forms in Triton X-100 detergent micelles

Protochlorophyll (PChl) forms were performed in Triton X-100 detergent micelles. The concentration of Triton X-100 was 7·10^?4 M (above the critical micellar concentration); the concentration of PChl varied between 1.6·10^?5 and 1.8·10^?4 M. Absorption, fluorescence and circular dichroism (CD) spectra were registered. The absorption spectra were resolved into Gaussian components by computer analysis. PChl forms with absorption bands at 632–634, 638, 652–654, 663–664, 668 and 676 nm and with fluorescence emission bands at 634–636, 640–644, 652–655, 677–678, 686 and 694–696 nm were observed in micellar solutions of different PChl concentrations. The CD spectra showed a strong dependence on the concentration of PChl: positive CD signals or positive Cotton effects were observed in the vicinity of 650 nm. The intensity of these signals increased in parallel with increasing concentration of PChl. No CD signals were found in the region of the longer wavelength absorption bands. These data show that the PChl exists in many different forms in this system, and the spectroscopic properties of these forms are determined by different molecular interactions viz., interactions of PChl with Triton X-100 or water molecules and/or by the aggregation of PChl.     

Triton X—100加KI对燕麦根细胞质膜ATP酶的溶解

在酸性条件下,1％ Triton X—100加 0.25mol/L KI能有效地溶解燕麦根细胞质膜ATP酶。溶解的ATP酶水解ATP的最适pH在6.5左右,酶活性受到Na_3VO_4和DES的强烈抑制,而不受Na_2MoO_4和NaN_3的抑制。溶解的酶液经透析后,K~ —ATP酶活性占Mg~(2 ),KCl—ATP酶活性的85％。     

Membrane-surfactant interactions The effect of triton X-100 on sarcoplasmic reticulum vesicles

The effect of Triton X-100 on purified sarcoplasmic reticulum vesicles has been studied by means of chemical, ultrastructural and enzymic techniques. At low detergent/membrane ratios (about 1 Triton X-100 per 60 phospholipid molecules) the only effect observed is an increase in vesicle permeability. Higher surfactant concentrations, up to a 1:1 detergent/phospholipid ratio, produce a large enhancement of ATPase activity. Membrane solubilization occurs as a critical phenomenon when the surfactant/phospholipid molar ratio reaches a value around 1.5:1, corresponding to 2 μmol Triton X-100/mg protein. At this point, the suspension turbidity drops, virtually all the protein and phospholipid is solubilized and every organized structure disappears. Simultaneously, a dramatic increase in the specific activity of the solubilized ATPase is observed. The sudden solubilization of almost all the bilayer components at a given detergent concentration is attributed to the relative simplicity of this membrane system. Solubilization takes place at the same surfactant/membrane ratio, at least between 0.5 and 4 mg membrane protein/ml. The non-solubilized residue seems to consist mainly of delipidized aggregated forms of ATPase.     

Hydrogenation of Triton X-100 eliminates its fluorescence and ultraviolet light absorption while preserving its detergent properties

The ultraviolet-light absorption and fluorescence of Triton X-100 were virtually eliminated by hydrogenation to its reduced cyclohexyl analog, RTX-100. The critical micelle concentration of RTX-100 was 12% higher than that of Triton X-100. RTX-100 and Triton X-100 were quite similar in their abilities to extract proteins from human erythrocyte membranes.     

Cytoskeletal structures inEuglena gracilis after triton X-100 extraction and dry cleaving

Summary A three-dimensional network of structural filaments was visible with common electron microscopes in the cytoplasm ofEuglena gracilis green cells extracted with buffers containing the nonionic detergent Triton X-100. A similar filamentous web was detected at the periphery of critical point dried cells cleaved on grids by means of an adhesive tape. SDS-polyacrylamide gel electrophoresis of the detergent-resistent cytoskeleton showed that actin or actin-like proteins of molecular weight in the range of 43–45 K are not among the components having a structural role inEuglena. The significance of these findings was discussed in relation to the capability of the alga to change the cell shape.The study was supported by grants from Consiglio Nazionale delle Ricerche (CNR) and Ministero della Pubblica Istruzione of Italy.     

The effect of triton X-100 on bovine brain synaptic membranes

Bovine brain synaptic membranes which were frozen and then extensively washed showed low affinity [³H]muscimol binding. These membranes contained GABA and calmodulin, apparently tightly bound within the membrane fraction. Membranes which were additionally treated with the detergent Triton X-100 showed high affinity [³H]muscimol binding. These membranes did not appear to contain GABA or calmodulin. Transmission electron microscopy studies demonstrated that the washed membrane fraction contained many synaptosomal and vesicular structures. Triton treatment led to the extensive rupture of these structures. These studies explain the well-reported findings of tightly bound GABA and calmodulin in brain membrane fractions, as being due to the entrapment of these compounds inside sealed membrane-bound structures which are still present after a freezethaw and extensive wash treatment, their complete removal requiring Triton-treatment to rupture the vesicles.     

The phospholipid fatty acids of Porphyridium purpureum cultured in the presence of triton x-100 and sodium desoxycholate

The phospholipid fatty acid composition of Porphyridium purpureum grown on a solid medium was studied in the presence of Triton X-100 (TX) and sodium desoxycholate (SDC). The most common fatty acids in PC and PE were palmitic (16:0), stearic (18:0), linoleic (18:2ω6), arachidonic (20:4ω6) and eicosapentaenoic (20:5ω3) acids, 20:4ω6 being very abundant. In PG the most common acids were 16:0, trans-hexaenoic acid (tr16:1ω3), oleic acid (18:1) and 20:4ω6. Both detergents caused an increase in the saturation of PC and, to a lesser extent, of PE. The relative amounts of short chain fatty acids increased. Both detergents increased the amounts of 16:0 and, correspondingly, decreased the amounts of 20:4ω6. In PG the amounts of both 16:0 and tr 16:1ω3 increased and the amounts of 18:0, 18:2ω6 and 20:4ω6 decreased in the presence of detergents. The changes were always greatest at the concentrations of 5–10 ppm TX or SDC. At 20 ppm the fatty acid compositions, especially with SDC, were very similar to the controls, which suggests a change in the detergent effect between 10–20 ppm. The normal PC/PE ratio was 5.6 and the (PC+ PE)/PG ratio 39.0. Both detergents caused a marked decrease in these ratios. Because the detergent effects are not linear, it seems that even very low detergent concentrations have an important influence on algae in polluted waters.     

Identification and characterization of a Triton X-100 replacement for virus inactivation

Triton X-100 detergent treatment is a robust enveloped virus inactivation unit operation included in biopharmaceutical manufacturing processes. However, the European Commission officially placed Triton X-100 on the Annex XIV authorization list in 2017 because a degradation product of Triton X-100, 4-(1,1,3,3-tetramethylbutyl) phenol (also known as 4-tert-octylphenol), is considered to have harmful endocrine disrupting activities. As a result, the use of Triton X-100 in the European Economic Area (EEA) would not be allowed unless an ECHA issued authorization was granted after the sunset date of January 4, 2021. This has prompted biopharmaceutical manufacturers to search for novel, environment-friendly alternative detergents for enveloped virus inactivation. In this study, we report the identification of such a novel detergent, Simulsol SL 11W. Simulsol SL 11W is an undecyl glycoside surfactant produced from glucose and C11 fatty alcohol. We report here that Simulsol SL 11W was able to effectively inactive enveloped viruses, such as xenotropic murine leukemia virus (XMuLV) and pseudorabies virus (PRV). By using XMuLV as a representative enveloped virus, the influence of various parameters on the effectiveness of virus inactivation was evaluated. Virus inactivation by Simulsol SL 11W was effective across different clarified bioreactor harvests at broad concentrations, pH, and temperature ranges. Simulsol SL 11W concentration, temperature of inactivation, and treatment time were identified as critical process parameters for virus inactivation. Removal of Simulsol SL 11W was readily achieved by Protein A chromatography and product quality was not affected by detergent treatment. Taken together, these results have shown the potential of Simulsol SL 11W as a desirable alternative to Triton X-100 for enveloped virus inactivation that could be readily implemented into biopharmaceutical manufacturing processes.     

Release of remnant plasma membrane from milk fat globules by Triton X-100

The nonionic detergent, Triton X-100, was investigated as an agent for releasing plasma membrane from milk fat globules. The sedimentable material ($\text{50 000 × g}$, 1 h) derived by treating washed goat globules with the detergent (0.2%) was compared to membrane made by the classical globule churning procedure. Characterization included lipid and protein analyses, gel electrophoresis of peptide components, determination of enzymatic activities, and examination with the electron microscope. The results established that the detergent-released material is membrane with similarities to the product by churning. Evaluation of variables revealed that a detergent concentration of 0.1 to 0.2% and reaction temperature of 20–22°C appear optimum with respect to membrane yield when a reaction time of 2 min is employed. At higher detergent concentrations or temperatures removal of phospholipid from the membrane was maximized. Triton X-100 was observed to release membrane from milk fat globules of the goat, human and cow, the latter with a minor procedural modification. The detergent based method is a convenient procedure for obtaining plasma membrane material in good yield for biochemical studies. It also should aid investigations of milk fat globule structure.     

The effect of cholesterol on the solubilization of phosphatidylcholine bilayers by the non-ionic surfactant Triton X-100

Most of the studies on the solubilization of model membranes by Triton X-100 (TR) involve one lipid. The aim of the present study was to evaluate the effect of the addition of cholesterol on the solubilization of bilayers made of palmitoyloleoylphosphatidylcholine (POPC) or dipalmitoylphosphatidylcholine (DPPC). Detailed investigation of the kinetics of solubilization of the cholesterol-containing bilayers by TR at different temperatures reveals that: (i) At 4 degrees C, solubilization of both systems is relatively slow. Hence, in order to prevent misleading conclusions from turbidity measurements it is important to monitor the solubilization after steady-state values of optical density (OD) are reached. (ii) Studies of the temperature-induced changes of the aggregates present in mixtures of TR, POPC and cholesterol indicate that the state of aggregation at all temperatures (including 4 degrees C) represents equilibrium. By contrast, for DPPC/cholesterol/TR mixtures "kinetic traps" may occur not only at 4 degrees C but at higher temperatures as well (e.g. 37 degrees C). (iii) The presence of cholesterol in POPC bilayers makes the bilayers more resistant to solubilization at low temperatures, especially at 4 degrees C. As a consequence, the temperature dependence of the TR concentration required for complete solubilization (Dt(sol)) is no longer a monotonically increasing function (as for POPC bilayers) but a bell-shaped function, with a minimum at about 25 degrees C. Inclusion of cholesterol in DPPC bilayers makes the bilayers more resistant to solubilization at all temperatures except 4 degrees C. In this system, we observe a bell-shaped dependence of Dt(sol) on temperature, with a minimum at 37 degrees C. (iv) Both the rate of vesicle size growth and the rate of the solubilization of POPC vesicles are not affected by the inclusion of cholesterol in the bilayers. Similarly, cholesterol did not affect significantly the rate of size growth of DPPC bilayers at all temperatures, but reduced the rate of solubilization at 4 degrees C.     

Effect of triton X-100 on compactin production from<Emphasis Type="Italic">Penicillium citrinum</Emphasis>

Glucose alone was found to be the most effective carbon source for producing compactin. An initial glucose concentration of 40 g/L gave the highest compactin concentration of 250 mg/L. Among the various nitrogen sources, when 5 g/L of pharmamedia and soybean meal as the sole nitrogen source were used, respectively, the compactin concentration was higher than 250 mg/L. Especially, in the case of the mixture of 6 g/L of pharmamedia and 8 g/L of soybean meal, the compactin concentration was 400 mg/L. To select the best surfactant for effective compactin production, various surfactants were investigated. When Triton X-100 was used, the maximum compactin concentration was 445 mg/L. With the initial concentration ranging from 1.5 to 2.0 g/L, the compactin concentration was the highest at 465–450 g/L. The cell concentration was similar to that of the control without the addition of Triton X-100. On the other hand, when the above 4.0 g/L of Triton X-100 were used, the cell concentration decreased. Using the based results the continuous fed-batch cultures by adding the Triton X-100 were carried out for 10 days in an air-lift bioreactor. When 1.5 g/L of Triton X-100 was added to the culture broth at 0, 4, and 8 days of culture, respectively, the compactin production was increased with the increase of culture time. The maximum compactin concentration after 10 days of culture was 1,200 mg/L, which was about 2.0-fold higher than that of the control without the addition of Triton X-100.     

Triton X-100-induced lipoteichoic acid release is correlated with the methicillin resistance in Staphylococcus aureus

We previously reported that Triton X-100 (TRX) reduced methicillin resistance in Staphylococcus aureus, although the degree of reduction varied among strains. One of the biological effects of TRX on S. aureus cells was enhancement of lipoteichoic acid (LTA) release. We investigated the correlation between the amount of LTA released and the degree of reduction in methicillin resistance induced by TRX. The strains showing the greatest reduction of methicillin resistance released the largest amount of LTA, compared to those showing slight or moderate reduction. A mutant whose resistance was not affected by TRX did not increase its release of LTA. These findings suggest that LTA release is associated with a reduction in methicillin resistance in the presence of TRX.     

Solubilization of human erythrocyte membrane glycoproteins by triton X-100.

1. The enzymic removal of sialic acid residues from the glycoproteins of the human erythrocyte decreases the solubilization of membrane glycoprotein by Triton X-100. 2. The solubilization of asialoglycoprotein by Triton X-100 may be restored by the addition of borate. 3. Use of this non-ionic detergent in the presence of borate, as a general procedure for the mild solubilization of membrane glycoproteins deficient in sialic acid residues, is discussed.     

Unusual enthalpy changes which accompany the titration of dimyristoylphosphatidylcholine vesicles with Triton X-100

The enthalpy changes which accompany the titration of 0.1% and 0.25% small unilamellar and multiameller vesicle samples of dimyristoylphosphatidylcholine with 2% Triton X-100 in 0.067 M phosphate buffer (pH 7.4) containing 0.15 M NaCl have been determined by titration calorimetry at 21 degrees C and 28 degrees C, the enthalpy change for both type of vesicles was zero within the limits of experimental error. At 21 degrees C, the multilamellar vesicle samples exhibited an enthalpy change of 1.35 +/- 0.48 and 2.47 +/- 0.98 kcal/mol dimyristoylphosphatidylcholine which was complete at a molar ratio of dimyristoylphosphatidylcholine to Triton of 3.21 +/- 0.84 and 5.77 +/- 1.05 for 0.1% and 0.25% dimyristoylphosphatidylcholine solutions, respectively. An exothermic transition of -2.39 +/- 0.30 and -2.05 +/- 0.69 kcal/mol phospholipid followed by an endothermic transition of 1.37 +/- 0.12 and 1.94 +/- 0.20 kcal/mol dimyristoylphosphatidylcholine was observed at 21 degrees C for 0.1% and 0.25% small unilamellar vesicle samples, respectively. In addition the nearly athermal association of the small unilemellar vesicle samples at 21 degrees C was observed, which may be an appropriate model for biological membrane fusion.     

Temperature-dependence of the solubilization of dipalmitoylphosphatidylcholine (DPPC) by the non-ionic surfactant Triton X-100, kinetic and structural aspects

Most of the studies on the solubilization of model membranes conducted thus far involved model membranes made of liquid-crystalline phospholipids. Relatively little is known on the influence of temperature and of the phase of the lipid bilayers on their solubilization by detergents. The aim of the present study was to gain knowledge about the temperature and phase dependence of the solubilization of phospholipid bilayers by the non-ionic detergent Triton X-100 (TR). Detailed investigation of the kinetics of the solubilization of dipalmitoylphosphatidylcholine (DPPC), as well as of palmitoyloleoylphosphatidylcholine (POPC) by TR at different temperatures reveals that: (i) solubilization of DPPC is a relatively slow process, especially below Tm. This means that in order to prevent misleading conclusions it is important to monitor the solubilization after a steady state is established. (ii) Both the steady state structure and size of DPPC/TR aggregates and the kinetics of solubilization depend on temperature. (iii) The TR concentration required for solubilization of POPC bilayers is an increasing function of temperature, although no phase change of bilayers occurs in the studied temperature range. (iv) Detailed studies of the temperature-induced changes of the aggregates present in DPPC/TR or POPC/TR mixtures suggest that the state of aggregation at any temperature above 23 degrees C represents equilibrium. By contrast, for DPPC/TR mixtures at 4 degrees C all the processes are very slow, which complicates the interpretation of results obtained through the common practice of studying "rafts" by investigating detergent-resistant membranes.     

Triton X-100 alters the resistance level of methicillin-resistant Staphylococcus aureus to oxacillin

Abstract We used population analysis to examine the effects of Triton X-100 on the level of resistance to oxacillin of 18 methicillin-resistant Staphylococcus aureus . In the presence of 0.02% Triton X-100, 17 formerly methicillin-resistiant strains exhibited enhanced sensitivity to oxacillin. One homogeneous isolate, KSAF1 was barely affected by the Triton X-100. Sensitivities of lysostaphin, 51 kDa N -acetylglucosaminidase and 62 kDa N -acetylmuramoyl-l-alanine amidase to heat-inactivated cells were not affected when the bacteria were grown in 0.02% Triton X-100. Our data, together with those of a previous study, suggested that Triton X-100 alters the resistance level of methicillin-resistant S. aureus by influencing a factor(s) other than PBPs, bacteriolytic enzymes, or femAB products.