An official website of the United States government
Abstract This review is in support of the development of selective, precise, fast, and validated capillary electrophoresis (CE) methods. It follows up a similar article from 1998, Wätzig H, Degenhardt M, Kunkel A. “Strategies for capillary electrophoresis: method development and validation for pharmaceutical and biological applications,” pointing out which fundamentals are still valid and at the same time showing the enormous achievements in the last 25 years. The structures of both reviews are widely similar, in order to facilitate their simultaneous use. Focusing on pharmaceutical and biological applications, the successful use of CE is now demonstrated by more than 600 carefully selected references. Many of those are recent reviews; therefore, a significant overview about the field is provided. There are extra sections about sample pretreatment related to CE and microchip CE, and a completely revised section about method development for protein analytes and biomolecules in general. The general strategies for method development are summed up with regard to selectivity, efficiency, precision, analysis time, limit of detection, sample pretreatment requirements, and validation.
more »
« less
Taylor dispersion analysis in fused silica capillaries: a tutorial review
Biological and pharmaceutical analytes like liposomes, therapeutic proteins, nanoparticles, and drug-delivery systems are utilized in applications, such as pharmaceutical formulations or biomimetic models, in which controlling their size is often critical. Many of the common techniques for sizing these analytes require method development, significant sample preparation, large sample quantities, and lengthy analysis times. In other cases, such as DLS, sizing can be biased towards the largest constituents in a mixture. Therefore, there is a need for more rapid, sensitive, accurate, and straightforward analytical methods for sizing macromolecules, especially those of biological origin which may be sample-limited. Taylor dispersion analysis (TDA) is a sizing technique that requires no calibration and consumes only nL to pL sample volumes. In TDA, average diffusion coefficients are determined via the Taylor–Aris equation by characterizing band broadening of an analyte plug under well-controlled laminar flow conditions. Diffusion coefficient can then be interpreted as hydrodynamic radius ( R H ) via the Stokes–Einstein equation. Here, we offer a tutorial review of TDA, intended to make the method better understood and more widely accessible to a community of analytical chemists and separations scientists who may benefit from the unique advantages of this versatile sizing method. We first provide a tutorial on the fundamental principles that allow TDA to achieve calibration-free sizing of analytes across a wide range of R H , with an emphasis on the reduced sample consumption and analysis times that result from utilizing fused silica capillaries. We continue by highlighting relationships between operating parameters and critically important flow conditions. Our discussion continues by looking at methods for applying TDA to sample mixtures via algorithmic approaches and integration of capillary electrophoresis and TDA. Finally, we present a selection of reports that demonstrate TDA applied to complex challenges in bioanalysis and materials science.
more »
« less
- Award ID(s):
- 2054748
- PAR ID:
- 10313797
- Date Published:
- Journal Name:
- Analytical Methods
- Volume:
- 13
- Issue:
- 21
- ISSN:
- 1759-9660
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
Mass spectrometry is a powerful analytical technique used at every stage of the pharmaceutical research process. A specialized subset of this technique, mass spectrometry imaging (MSI) has emerged as an important technique for determining the spatial distribution of drugs in biological samples. Despite the importance of MSI, its quantitative capabilities are still limited due to the complexity of biological samples and the lack of separation prior to analysis. This makes the simultaneous quantification and visualization of analytes challenging. Several techniques have been developed to address this challenge and enable quantitative MSI. One of these techniques is the mimetic tissue model, which involves the incorporation of an analyte of interest into tissue homogenates at several concentrations. A calibration curve that accounts for signal suppression by the complex biological matrix is then created by measuring the signal of the analyte in the series of tissue homogenates. Herein, we use the mimetic tissue model on a triple quadrupole mass spectrometer (QqQ) in multiple reaction monitoring (MRM) mode to demonstrate the quantitative abilities of nanospray desorption electrospray ionization (nano-DESI) and compare these results with those obtained using atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI). For the tested compounds, our findings indicate that nano-DESI achieves lower standard deviations than AP-MALDI which contributes to nano-DESI also achieving lower limits of detection (LOD) for the analytes studied. Additionally, we discuss the limitations of the mimetic tissue model in the quantification of certain analytes and the challenges involved with the implementation of the model.more » « less
-
Field-flow fractionation (FFF) is a family of techniques that was created especially for separating and characterizing macromolecules, nanoparticles, and micrometer-sized analytes. It is coming of age as new nanomaterials, polymers, composites, and biohybrids with remarkable properties are introduced and new analytical challenges arise due to synthesis heterogeneities and the motivation to correlate analyte properties with observed performance. Appreciation of the complexity of biological, pharmaceutical, and food systems and the need to monitor multiple components across many size scales have also contributed to FFF's growth. This review highlights recent advances in FFF capabilities, instrumentation, and applications that feature the unique characteristics of different FFF techniques in determining a variety of information, such as averages and distributions in size, composition, shape, architecture, and microstructure and in investigating transformations and function.more » « less
-
null (Ed.)Sucrose is among the main products of photosynthesis that are deemed necessary for plant growth and survival. It is produced in the mesophyll cells of leaves and translocated to different parts of the plant through the phloem. Progress in understanding this transport process remains fraught with experimental difficulties, thereby prompting interest in theoretical approaches and laboratory studies. The Münch pressure and mass flow model is one of the accepted hypotheses describing the physics of sucrose transport in the phloem. It is based on osmosis creating an energy potential difference between the source and the sink. The flow responding to this energy potential is assumed laminar and described by the Hagen–Poiseuille equation. This study revisits such osmotically driven flows in tubes with membrane walls by including the effects of Taylor dispersion on mass transport. This effect has been overlooked in phloem flow studies. Taylor dispersion can increase the effective transport of solutes by increasing the apparent diffusion coefficient. It is shown that, in addition to the conventional diffusive correction derived for impermeable tube walls, a new adjustment to the mean advective terms arises because of osmotic effects. Because the molecular Schmidt number is very large for sucrose in water, the sucrose front speed and travel times have a direct dependence on the Péclet number for different ranges of the Münch number. This study establishes upper limits on expected Taylor dispersion enhancement of sucrose transport.more » « less
-
RationaleSulfur isotope analysis of organic sulfur‐containing molecules has previously been hindered by challenging preparatory chemistry and analytical requirements for large sample sizes. The natural‐abundance sulfur isotopic compositions of the sulfur‐containing amino acids, cysteine and methionine, have therefore not yet been investigated despite potential utility in biomedicine, ecology, oceanography, biogeochemistry, and other fields. MethodsCysteine and methionine were subjected to hot acid hydrolysis followed by quantitative oxidation in performic acid to yield cysteic acid and methionine sulfone. These stable, oxidized products were then separated by reversed‐phase high‐performance liquid chromatography (HPLC) and verified via offline liquid chromatography/mass spectrometry (LC/MS). The sulfur isotope ratios (δ34S values) of purified analytes were then measured via combustion elemental analyzer coupled to isotope ratio mass spectrometry (EA/IRMS). The EA was equipped with a temperature‐ramped chromatographic column and programmable helium carrier flow rates. ResultsOn‐column focusing of SO2in the EA/IRMS system, combined with reduced He carrier flow during elution, greatly improved sensitivity, allowing precise (0.1–0.3‰ 1 s.d.) δ34S measurements of 1 to 10 μg sulfur. We validated that our method for purification of cysteine and methionine was negligibly fractionating using amino acid and protein standards. Proof‐of‐concept measurements of fish muscle tissue and bacteria demonstrated differences up to 4‰ between the δ34S values of cysteine and methionine that can be connected to biosynthetic pathways. ConclusionsWe have developed a sensitive, precise method for measuring the natural‐abundance sulfur isotopic compositions of cysteine and methionine isolated from biological samples. This capability opens up diverse applications of sulfur isotopes in amino acids and proteins, from use as a tracer in organisms and the environment, to fundamental aspects of metabolism and biosynthesis.more » « less
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1039/d1ay00588j
