Dwight Stoll

Gustavus Adolphus College, Chemistry, Faculty Member

Followers

Following

Public Views

Interests

Uploads

Papers by Dwight Stoll

Trapping‐Enrichment Multi‐dimensional Liquid Chromatography with On‐Line Deuterated Solvent Exchange for Streamlined Structure Elucidation at the Microgram Scale

Angewandte Chemie, 2022

At the forefront of chemistry and biology research, development timelines are fast-paced and larg... more At the forefront of chemistry and biology research, development timelines are fast-paced and large quantities of pure targets are rarely available. Herein, we introduce a new framework, which is built upon an automated, online trapping-enrichment multi-dimensional liquid chromatography platform (TE- Dt -mDLC) that enables: 1) highly efficient separation of complex mixtures in a first dimension ( 1 D -UV); 2) automated peak trapping-enrichment and buffer removal achieved through a sequence of H 2 O and D 2 O washes using an independent pump setup; and 3) a second dimension separation ( 2 D -UV-MS) with fully deuterated mobile phases and fraction collection to minimize protic residues for immediate NMR analysis while bypassing tedious drying processes and minimizing analyte degradation. Diverse examples of target isolation and characterization from organic synthesis and natural product chemistry laboratories are illustrated, demonstrating recoveries above 90% using as little as a few micrograms of material.

In Silico Multifactorial Modeling for Streamlined Development and Optimization of Two-Dimensional Liquid Chromatography

Analytical Chemistry, 2021

Continued adoption of two-dimensional liquid chromatography (2D-LC) in industrial laboratories wi... more Continued adoption of two-dimensional liquid chromatography (2D-LC) in industrial laboratories will depend on the development of approaches to make method development for 2D-LC more systematic, less tedious, and less reliant on user expertise. In this paper, we build on previous efforts in these directions by describing the use of multifactorial modeling software that can help streamline and simplify the method development process for 2D-LC. Specifically, we have focused on building retention models for second dimension (2D) separations involving variables including gradient time, temperature, organic modifier blending, and buffer concentration using LC simulator (ACD/Labs) software. Multifactorial retention modeling outcomes are illustrated as resolution map planes or cubes that enable straightforward location of 2D conditions that maximize resolution while minimizing analysis time. We also illustrate the practicality of this approach by identifying conditions that yield baseline separation of all compounds co-eluting from a first dimension (1D) separation using a single combination of 2D stationary phase and elution conditions. The multifactorial retention models were found to be very accurate for both the 1D and 2D separations, with differences between experimental and simulated retention times of less than 0.5%. Pharmaceutical applications of this approach for multiple heartcutting 2D-LC were demonstrated using IEC-IEC or achiral RPLC-chiral RPLC for 2D separations of multicomponent mixtures. The framework outlined here should help make 2D-LC method development more systematic and streamline development and optimization for a variety of 2D-LC applications in both industry and academia.

Fast methods for structurally similar compounds using carbon HPLC columns

Lc Gc North America, 2002

Method development can be challenging for structurally similar compounds in reversed-phase liquid... more Method development can be challenging for structurally similar compounds in reversed-phase liquid chromatography (RPLC). A new carbon-clad zirconia phase offers dramatically different chromatographic selectivity for thesetypes of compounds when compared with traditional silica-based bonded phases. This note shows baseline separation of six structurally similar sulfate-steroid conjugates using a zirconia-based ZirChrom®-CARB HPLC column.

Fast Separation of Nitrosamines

Lc Gc North America, 2002

Interest in the analysis of nitrosamine compounds is increasing as researchers become more aware ... more Interest in the analysis of nitrosamine compounds is increasing as researchers become more aware of their toxicity and presence in the environment, foods, and pharmaceuticals. The extraordinary chemical and thermal stability of zirconia-based stationary phases allows for the rapid separation of these polar, basic compounds with good peak shape and efficiency. This note shows the separation of nine low molecular weight nitrosamines using a DiamondBond-C18 column.

Download

Implications of dispersion in connecting capillaries for separation systems involving post-column flow splitting

Journal of Chromatography A, 2021

Implications of dispersion in connecting capillaries for separation systems involving post-column... more

format_quoteStrong correlation between estimated and measured peak standard deviations validates conservation of mass at split points.format_quote

Download

Mapping the Separation Landscape in Two-Dimensional Liquid Chromatography: Blueprints for Efficient Analysis and Purification of Pharmaceuticals Enabled by Computer-Assisted Modeling

Analytical Chemistry, 2020

Recent developments in two-dimensional liquid chromatography (2D-LC) now make separation and anal... more Recent developments in two-dimensional liquid chromatography (2D-LC) now make separation and analysis of very complex mixtures achievable. Despite being such a powerful chromatographic tool, current 2D-LC technology requires a series of arduous method development activities poorly suited for a fastpaced industrial environment. Recent introductions of new technologies including active solvent modulation and a support for multicolumn 2D-LC are helping to overcome this stigma. However, many chromatography practitioners believe that the lack of a systematic way to effectively optimize 2D-LC separations is a missing link in securing the viability of 2D-LC as a mainstay for industrial applications. In this work, a computer-assisted modeling approach that dramatically simplifies both offline and online 2D-LC method developments is introduced. Our methodology is based on mapping the separation landscape of pharmaceutically relevant mixtures across both first (1 D) and second (2 D) dimensions using LC Simulator (ACD/Labs) software. Retention models for 1 D and 2 D conditions were built using a minimal number of multifactorial modeling experiments (2 × 2 or 3 × 3 parameters: gradient slope, column temperature, and different column and mobile phase combinations). The approach was first applied to online 2D-LC analysis involving achiral and chiral separations of complex mixtures of enantiomeric species. In these experiments, the retention models proved to be quite accurate for both the 1 D and 2 D separations, with retention time differences between experiments and simulations of less than 3.5%. This software-based concept was also demonstrated for offline 2D-LC purification of drug substances.

format_quoteRetention models for achiral and chiral separations showed less than 3.5% differences between experimental and simulated retention times.format_quote

Download

Orthogonal Middle-up Approaches for Characterization of the Glycan Heterogeneity of Etanercept by Hydrophilic Interaction Chromatography Coupled to High-Resolution Mass Spectrometry

Analytical Chemistry, 2018

Etanercept is a recombinant Fc-fusion protein widely used to treat rheumatic diseases. This prote... more Etanercept is a recombinant Fc-fusion protein widely used to treat rheumatic diseases. This protein is highly glycosylated and contains numerous O-and N-glycosylation sites. Since glycosylation is recognized as an important critical quality attribute (CQA) that can affect immunogenicity, solubility, and stability of Fc-fusion proteins, it should be thoroughly characterized. In this work, hydrophilic interaction chromatography (HILIC) was combined with high resolution mass spectrometry (HRMS) by using a QTOF/MS instrument to assess glycosylation of etanercept at the middle-up level of analysis (fragments of circa 25-30 kDa). In addition, a combination of different enzymatic digestion procedures (i.e. glycosidase, sialidase and protease) was systematically employed to facilitate spectra deconvolution. With the developed procedure, the main post-translational modifications (PTMs) of etanercept were assessed, and a global overview of the subunit-specific distribution of the glycosylation pattern was obtained at a middle-up level of analysis.

Download

Robust methods with high efficiency, bonded-carbon HPLC columns

Lc Gc North America, 2002

Loss of stationary phase, retention drift, and short column life are common problems using bonded... more Loss of stationary phase, retention drift, and short column life are common problems using bonded-phase silicas, especially with aggressive mobile phases. A new bonding technology overcomes these problems by attaching C18 groups to a carbon surface with ultra-stable carbon-carbon bonds. This note shows that robust methods with high efficiency are now possible using ultra-stable bonded-carbon columns.

Download

Direct identification of rituximab main isoforms and subunit analysis by online selective comprehensive two-dimensional liquid chromatography - mass spectrometry

Analytical chemistry, Jan 6, 2015

In this proof-of-concept study, rituximab, a reference therapeutic monoclonal antibody (mAb), was... more In this proof-of-concept study, rituximab, a reference therapeutic monoclonal antibody (mAb), was characterized through the implementation of online, selective comprehensive two-dimensional liquid chromatography (sLCxLC) coupled with mass spectrometry (MS), and using a middle-up approach. In this setup, cation exchange chromatography (CEX) and reversed phase liquid chromatography (RPLC) were used as the first and second separation dimensions, respectively. As illustrated in this work, the combination of these two chromatographic modes allows a direct assignment of the identities of CEX peaks using data from the TOF/MS detector, because RPLC is directly compatible with MS detection, whereas CEX is not. In addition, the resolving power of CEX is often considered to be limited and therefore, this 2D approach provides an improvement in peak capacity and resolution when high performance second dimension separations are used, instead of simply using the second dimension separation as a de...

format_quoteRituximab characterized through sLCxLC coupled with MS, employing a novel middle-up analytical approach.format_quote

Download

Effect of pressure pulses at the interface valve on the stability of second dimension columns in online comprehensive two-dimensional liquid chromatography

Journal of chromatography. A, Jan 23, 2015

Users of online comprehensive two-dimensional liquid chromatography (LCxLC) frequently acknowledg... more Users of online comprehensive two-dimensional liquid chromatography (LCxLC) frequently acknowledge that the mechanical instability of HPLC columns installed in these systems, particularly in the second dimension, is a significant impediment to its use. Such instability is not surprising given the strenuous operating environment to which these columns are subjected, including the large number (thousands per day) of fast and large pressure pulses resulting from interface valve switches (on the timescale of tens of milliseconds) associated with very fast second dimension separations. There appear to be no published reports of systematic studies of the relationship between second dimension column lifetime and any of these variables. In this study we focused on the relationship between the lifetimes of commercially available columns and the pressure pulses observed at the inlet of the second dimension column that occur during the switching of the valve that interfaces the two dimensions ...

Evaluation of detection sensitivity in comprehensive two-dimensional liquid chromatography separations of an active pharmaceutical ingredient and its degradants

Analytical and bioanalytical chemistry, 2015

In this paper, we describe the findings of a study aimed at assessing the detection sensitivity o... more In this paper, we describe the findings of a study aimed at assessing the detection sensitivity of comprehensive two-dimensional high-performance liquid chromatography (LCxLC) separation of a degraded active pharmaceutical ingredient (API) with UV absorption as the detection technique. Specifically, we have examined the impact of the volume and solvent composition (referred to as "interface conditions") of fractions of first-dimension column effluent transferred to the second dimension for further separation on the ability to resolve and detect low-abundance compounds. Historically, LCxLC has been perceived as being inferior to 1D-LC from the point of view of detection sensitivity. In this work, we demonstrate that LCxLC is sufficiently sensitive to be useful in the pharmaceutical context where in general impurities present at 0.05 % (relative to the API concentration) should be quantified. Moreover, we find that this level of sensitivity is only attained under certain con...

Two dimensional assisted liquid chromatography – a chemometric approach to improve accuracy and precision of quantitation in liquid chromatography using 2D separation, dual detectors, and multivariate curve resolution

Analytica Chimica Acta, 2015

A novel method for quantitation in LC Â LC is proposed based on use of dual detectors. Curve reso... more A novel method for quantitation in LC Â LC is proposed based on use of dual detectors. Curve resolution was carried out on data from both detectors. The combined use of data from both detectors lead to improved quantitative results. In some cases, use of only first dimension data results in improved quantification.