Triton X-100 (TX-100) is a widely used detergent to prevent viral contamination of manufactured biologicals and biopharmaceuticals, and acts by disrupting membrane-enveloped virus particles. However, environmental concerns about ecotoxic byproducts are leading to TX-100 phase out and there is an outstanding need to identify functionally equivalent detergents that can potentially replace TX-100. To date, a few detergent candidates have been identified based on viral inactivation studies, while direct mechanistic comparison of TX-100 and potential replacements from a biophysical interaction perspective is warranted. Herein, we employed a supported lipid bilayer (SLB) platform to comparatively evaluate the membrane-disruptive properties of TX-100 and a potential replacement, Simulsol SL 11W (SL-11W), and identified key mechanistic differences in terms of how the two detergents interact with phospholipid membranes. Quartz crystal microbalance-dissipation (QCM-D) measurements revealed that TX-100 was more potent and induced rapid, irreversible, and complete membrane solubilization, whereas SL-11W caused more gradual, reversible membrane budding and did not induce extensive membrane solubilization. The results further demonstrated that TX-100 and SL-11W both exhibit concentration-dependent interaction behaviors and were only active at or above their respective critical micelle concentration (CMC) values. Collectively, our findings demonstrate that TX-100 and SL-11W have distinct membrane-disruptive effects in terms of potency, mechanism of action, and interaction kinetics, and the SLB platform approach can support the development of biophysical assays to efficiently test potential TX-100 replacements.
In light of regulatory considerations, there are ongoing efforts to identify Triton X-100 (TX-100) detergent alternatives for use in the biological manufacturing industry to mitigate membrane-enveloped pathogen contamination. Until now, the efficacy of antimicrobial detergent candidates to replace TX-100 has been tested regarding pathogen inhibition in endpoint biological assays or probing lipid membrane disruption in real-time biophysical testing platforms. The latter approach has proven especially useful to test compound potency and mechanism of action, however, existing analytical approaches have been limited to studying indirect effects of lipid membrane disruption such as membrane morphological changes. A direct readout of lipid membrane disruption by TX-100 detergent alternatives would be more practical to obtain biologically relevant information to guide compound discovery and optimization. Herein, we report the use of electrochemical impedance spectroscopy (EIS) to investigate how TX-100 and selected replacement candidates—Simulsol SL 11W (Simulsol) and cetyltrimethyl ammonium bromide (CTAB)—affect the ionic permeability of tethered bilayer lipid membrane (tBLM) platforms. The EIS results revealed that all three detergents exhibited dose-dependent effects mainly above their respective critical micelle concentration (CMC) values while displaying distinct membrane-disruptive behaviors. TX-100 caused irreversible membrane disruption leading to complete solubilization, whereas Simulsol caused reversible membrane disruption and CTAB induced irreversible, partial membrane defect formation. These findings establish that the EIS technique is useful for screening the membrane-disruptive behaviors of TX-100 detergent alternatives with multiplex formatting possibilities, rapid response, and quantitative readouts relevant to antimicrobial functions.
Membrane-disrupting lactylates are an important class of surfactant molecules that are esterified adducts of fatty acid and lactic acid and possess industrially attractive properties, such as high antimicrobial potency and hydrophilicity. Compared with antimicrobial lipids such as free fatty acids and monoglycerides, the membrane-disruptive properties of lactylates have been scarcely investigated from a biophysical perspective, and addressing this gap is important to build a molecular-level understanding of how lactylates work. Herein, using the quartz crystal microbalance–dissipation (QCM-D) and electrochemical impedance spectroscopy (EIS) techniques, we investigated the real-time, membrane-disruptive interactions between sodium lauroyl lactylate (SLL)—a promising lactylate with a 12-carbon-long, saturated hydrocarbon chain—and supported lipid bilayer (SLB) and tethered bilayer lipid membrane (tBLM) platforms. For comparison, hydrolytic products of SLL that may be generated in biological environments, i.e., lauric acid (LA) and lactic acid (LacA), were also tested individually and as a mixture, along with a structurally related surfactant (sodium dodecyl sulfate, SDS). While SLL, LA, and SDS all had equivalent chain properties and critical micelle concentration (CMC) values, our findings reveal that SLL exhibits distinct membrane-disruptive properties that lie in between the rapid, complete solubilizing activity of SDS and the more modest disruptive properties of LA. Interestingly, the hydrolytic products of SLL, i.e., the LA + LacA mixture, induced a greater degree of transient, reversible membrane morphological changes but ultimately less permanent membrane disruption than SLL. These molecular-level insights support that careful tuning of antimicrobial lipid headgroup properties can modulate the spectrum of membrane-disruptive interactions, offering a pathway to design surfactants with tailored biodegradation profiles and reinforcing that SLL has attractive biophysical merits as a membrane-disrupting antimicrobial drug candidate.
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