2D Peak Capacity Research Articles

Comprehensive two-dimensional gas chromatography using a miniaturized multiloop splitter-based non-cryogenic artificial trapping (M-SNAT) modulation device for analysis of cannabis samples

Multiloop splitter-based non-cryogenic artificial trapping (M-SNAT) modulation technique was developed, miniaturized and applied in comprehensive two-dimensional gas chromatography (GC×GC) for analysis of cannabis samples. The approach employed deactivated fuse silica (DFS) columns configured into multiple loop splitter system halving the perimeters of the progressively upstream loops. This splitter device was located between the first (1D) semi-nonpolar column outlet and a microfluidic Deans switch (DS). Each splitter loop splits a peak into two subpeaks having the same area with different void times. Three loops were then applied resulting in the number of the split subpeaks (nsplit) of 8 for each peak, and retention time differences between any two adjacent subpeaks (∆tR,split) were the same. By applying periodic heartcut event (H/C) within every artificial modulation period (PAM) of nsplit×∆tR,split, comprehensive split-and-trapped modulation profiles of analytes could be selectively transferred onto the second (2D) polar column (30 m) without cryogen consumption. This artificial modulation system was applied for analysis of cannabis samples with enhanced 2D peak capacity (2nc∼15). The established method was applied to analyse cannabis extracts using vegetable oils with or without frying process. This reveals 454 different peaks with 76, 92, 35 and 70 specific components specifically observed by using olive oil extraction (OE), fried OE, coconut oil extraction (CE) and fried CE, respectively.

Comprehensive two-dimensional gas chromatography under low-pressure conditions

The analysis of thermally labile and high-boiling point compounds by gas chromatography (GC) can be a challenge. One technique to overcome these challenges is low-pressure GC, which uses the vacuum produced from the mass spectrometer and wide-bore columns to elute compounds at significantly lower temperatures. While GC-MS is a powerful technique, comprehensive two-dimensional gas chromatography (GC × GC), allows for resolution of compounds that would typically coelute using GC. In this study, a pesticide standard mixture (8270 MegaMix Standard) was analyzed using a conventional GC × GC-TOFMS configuration (0.25 mm inner diameter (i.d.) to a 0.18 mm i.d. column) and low-pressure GC × GC-TOFMS configuration (0.53 mm i.d. to a 0.53 mm i.d. column). Elution temperatures, sensitivity, and peak capacity were investigated for both configurations. Compounds eluted an average of 30 °C less on the low-pressure GC × GC-TOFMS configuration compared to the conventional GC × GC-TOFMS configuration. Moreover, the compounds were separated in ∼13 min on the low-pressure GC × GC-TOFMS as opposed to 33 min for conventional GC × GC-TOFMS. However, due to the wide-bore columns and faster runtimes the low-pressure GC × GC-TOFMS had a lower, β corrected 2D peak capacity, nc,β,2D, of 1260 while the conventional GC × GC-TOFMS was 3588. Interestingly, both configurations yielded a similar peak capacity production of 93 peaks/min and 107 peaks/min for low-pressure and conventional GC × GC-TOFMS, respectively. A “real world” sample of diesel fuel was tested on the low-pressure and conventional GC × GC-TOFMS configurations and similar results were obtained compared to the pesticide standard mix except the peak capacity production of the low-pressure GC × GC-TOFMS configuration was higher than that of the conventional GC × GC-TOFMS method.

Development and Application of a Novel Multiloop Splitter-Based Non-cryogenic Artificial Trapping Modulation System in Comprehensive Two-Dimensional Gas Chromatography.

A multiloop splitter-based non-cryogenic artificial trapping (M-SNAT) modulation technique was established, which applied the first (1D) nonpolar and the second (2D) polar columns, deactivated fused silica (DFS) columns, a microfluidic Deans switch (DS), and splitters located between the 1D column outlet and the DS. The splitters were connected into multiple loops with a progressively doubled perimeter of the next loop. This enabled a duplex splitting mechanism within each loop consisting of splitting of analyte pulses, the pulse delay, and their combination which led to equally split peaks of the same analytes with the number of split peaks (nsplit) equal to 2m (m = number of loops). This system resulted in local profiles of artificially split-and-trapped analytes prior to their selective transfers onto the 2D column by means of periodic multiple heart-cuts (H/C). The developed SNAT approach can be successful, providing that the ratio of modulation period to sampling time (PM/tsamp) is equal to nsplit. The approach with nsplit = 16 was further developed into a single device platform and applied for the modulation of a wide range of compounds in waste tire pyrolysis samples with the RSD of ≤0.01 and <10% for the one-dimensional modulated peak times and peak areas, respectively (n = 50). The method enabled an artificial modulation mechanism without cryogen consumption and enhanced the 2D peak capacity (2nc) and 2D separation by use of a longer 2D column.

Second-Dimension Temperature Programming System for Comprehensive Two-Dimensional Gas Chromatography. Part 1: Precise Temperature Control Based on Column Electrical Resistance.

A second-dimension temperature programming system (2DTPS) for comprehensive two-dimensional gas chromatography (GC × GC) is introduced, and its performance is characterized. In the system, a commercial stainless-steel capillary column was used for the separation, as a heating element, and as a temperature sensor. The second dimension (2D) column was resistively heated and controlled using an Arduino Uno R3 microcontroller. Temperature measurement was accomplished by measuring the overall 2D column's electrical resistance. A diesel sample was used to compare the 2D peak capacity (2nc) and resolution (2Rs), while a perfume sample was used to compare the reproducibility of the system for within-day (n = 5) and day-to-day (n = 5) results. The 2nc improved by 52% with the 2DTPS compared to the secondary oven. The GC × GC system utilizing the 2DTPS had an average within-day and day-to-day relative standard deviation (RSD) of 0.02 and 0.12% for the 1D retention time (1tR), 0.56 and 0.58% for the 2D retention time (2tR), and 1.18 and 1.53% for the peak area, respectively.

Second dimension temperature programming system for comprehensive two-dimensional gas chromatography with true temperature control

A proof-of-concept second dimension (²D) temperature programming system for comprehensive two-dimensional gas chromatography (GC × GC) with true temperature control is introduced and its performance characterized. The ²D temperature programming system utilized a commercial stainless-steel capillary column that was resistively heated and controlled using an Arduino UNO microcontroller. Temperature measurement was accomplished using a differential thermocouple system attached to the ²D column. Diesel was injected in triplicate to compare the reproducibility of the GC × GC system without additional ²D heating, with ²D temperature programming, and with a constant offset, all accomplished using the ²D temperature programming system. Relative standard deviations for the ²D temperature programming were 0.02%, 0.49% and 1.2% for the first dimension (¹D) retention time, ²D retention time, and peak area, respectively. The estimated ²D peak capacity improved by 48% with ²D temperature programming compared to the separation without additional ²D heating.

Towards a miniaturized on-site nano-high performance liquid chromatography electrospray ionization ion mobility spectrometer with online enrichment

Safeguarding water quality is resource-intensive and costly. Especially in cases of accidents or disasters, compact devices that allow quick assessment of the current situation are lacking. The objective of this work is the development of a portable measuring device for on-site detection of pollutants in water based on nano-high performance liquid chromatography (nano-HPLC) and electrospray ionization (ESI) ion mobility spectrometry (IMS). Integrated online enrichment by means of solid phase extraction (SPE) further improves sensitivity. In this work, an SPE cartridge exchange unit is presented, which was developed by additive manufacturing in a cost-effective and resource-efficient way. Prerequisites are an easy adaptation to commercially available SPE cartridges and pressure stability of up to 50 bar. In addition, a compact ESI-IMS with 75 mm drift tube length and a resolving power of R = 100 is presented that enables ionization, separation based on ion mobility and detection of analytes at a flow rate of 0.6 – 1.8 µL/min from the liquid phase. This approach also allows miniaturization of the overall system leading to a reduction in energy requirements and the amount of solvents used. For future environmentally benign systems, complete elimination of toxic solvents would be ideal. Therefore, acetonitrile and the nontoxic ethanol are compared as organic mobile phase in terms of elution and ionization efficiency. For characterization, a test mixture containing relevant target analytes, such as chlortoluron, isoproturon, pyrimethanil, mepanipyrim, cyprodinil and carbamazepine, is analyzed. In addition, the analytical greenness metric approach tool is used to evaluate the overall system in terms of its green credentials. It was shown that ethanol can be well used as an organic solvent for the mobile phase and even exhibits better ionization than acetonitrile for the selected model analytes. Furthermore, a relatively high orthogonality of O=0.53 and an effective 2D peak capacity of 2Dneff=174 are reached. Due to the overall miniaturization and splitless coupling, the aims of green chemistry can be met, and ideally a value of 0.92 can be achieved using the AGREE tool for the presented system.

A systematic investigation of comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry with dynamic pressure gradient modulation for high peak capacity separations

Dynamic pressure gradient modulation (DPGM) in full modulation mode is optimized for comprehensive two-dimensional (2D) gas chromatography (GC × GC) with time-of-fight mass spectrometry (TOFMS) detection to obtain high peak capacity separations and demonstrate broad applicability for complex samples. A pulse valve introduces an auxiliary carrier gas flow at a T-union connecting the first dimension (1D) column to the second dimension (2D) column. At a sufficiently high auxiliary pressure (Paux) the 1D flow is temporarily stopped. Then, during each modulation period (PM) the valve is turned off briefly, a period termed the pulse width (pw), allowing the 1D effluent to essentially be reinjected onto the 2D column for the modulated separations. Modifications to the modulator assembly are provided to improve performance. Method optimization is demonstrated for a 116-component test mixture by tuning the Paux and the pw. For a PM = 2 s and 1F of 0.10 ml/min, the optimal pw and initial Paux selected were 200 ms and 330.9 kPa (33 psig), respectively. The 30 min separation of the test mixture provided a 1D peak capacity of 1nc = 330 and a 2D peak capacity of 2nc = 15, hence an ideal 2D peak capacity nc,2D = 1nc × 2nc = 4950. Likewise, the 2D peak capacity corrected for undersampling of the 1D separation was 4500 and corrected for both undersampling and sampling variation via statistical overlap theory was 4090. These results provide a 2-fold improvement in peak capacity relative to the previous DPGM study in full modulation mode for GC × GC-TOFMS. The optimized conditions were applied for a variety of applications: diesel fuel, derivatized cow serum, solid phase microextraction (SPME) of coffee headspace, and SPME of river water headspace. Additionally, the fraction of 2D separation space utilized (fcoverage), as defined by the minimum convex hull method, ranged from 0.60 to 0.85. We observed that any fcoverage correction to 2D peak capacity is highly sample dependent, since all samples, except for the diesel sample, were run with the same separation conditions, and yet the fcoverage ranged from 0.60 to 0.80.

Dynamic pressure gradient modulation for comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry detection

Dynamic pressure gradient modulation (DPGM) is investigated for comprehensive two-dimensional gas chromatography (GC × GC) with time-of-flight mass spectrometry (TOFMS) detection. With DPGM, a commercial pneumatic “pulse” valve is opened to introduce a suitably high auxiliary gas pressure at a T-junction connecting the first dimension (1D) and second dimension (2D) columns during the modulation period (PM), temporarily stopping the 1D flow. The valve is then closed for the duration of a pulse width (pw) to “re-inject” temporally focused 1D eluate onto the 2D column for separation. This flow modulation technique is observed to be compatible with TOFMS detection using a 2D flow rate of 4 ml/min for the separation of a 90-component test mixture. A 25 min separation window using a PM = 1 s and pw = 200 ms for full modulation (and 100% duty cycle) provided an average 1Wb = 4.5 s and 2Wb = 130 ms for a 2D peak capacity of nc,2D = 2700 (100 peaks per min). The detector response enhancement factor (DREF) serves as a metric for the enhanced sensitivity of the modulated relative to the unmodulated 1D peaks, with DREFs ranging between 10 and 20 and about a 5-fold improvement in signal-to-noise ratio (S/N). The bilinear “quality” of the GC × GC data is studied using the chemometric method parallel factor analysis (PARAFAC). Since PARAFAC requires sufficiently trilinear data, the reproducibility of the 2D peak shape for a given analyte is confirmed using lack-of-fit (LOF) and percent variation (R2) metrics. The limit-of-detection (LOD) for the representative analyte hexadecane is determined using PARAFAC, providing an LOD of 0.7 ppb (±0.03 ppb) for three replicates. Seven heavily overlapped analytes are also fully resolved by PARAFAC down to the part-per-million (ppm) concentration level, producing reproducible spectra with a majority of spectral match values (MV) over 800 (RSD ≤ 7.1%). This study provides promising results for DPGM as a flow modulation technique compatible with GC × GC-TOFMS, providing high sensitivity data suitable for chemometric analysis.

High resolution two-dimensional liquid chromatography coupled with mass spectrometry for robust and sensitive characterization of therapeutic antibodies at the peptide level

Separations of complex peptide mixtures have been a common target application for two-dimensional liquid chromatography over the last few decades. These separations have most frequently been carried out at the capillary scale, with columns on the order of 75 µm i.d. and flow rates on the order of 500 nL/min. Recently, however, several groups have worked to optimize comprehensive 2D-LC (LC × LC) separations of peptides at the analytical scale (i.e., 2 mm i.d. columns, and ca. 1 mL/min flow rates) and demonstrated peak capacities on the order of 5000 in analysis times of a few hours, using reversed-phase separations in both dimensions. In this paper we aim to advance the performance of such separations in two primary ways. First, we demonstrate that active solvent modulation (ASM) can be used to improve the 2D peak capacity by both enabling use of long and highly efficient first dimension (1D) columns, and by mitigating the deleterious effects of injecting large fractions of 1D effluent into the small columns that are required for fast and highly sensitive second dimension (2D) separations. Taken together these two benefits enable the realization of a peak capacity of 10,000 in an analysis time of four hours. This comes at the cost of increased instrument complexity compared to 1D-LC separations, but the 2D-LC approach is unquestionably the most efficient way to improve upon the resolving power of existing 1D-LC. Second, we have systematically studied the compromise between the peak capacity of each 2D separation and the operating pressure required to achieve that peak capacity. Understanding this compromise will be important to the development of LC × LC methods that both produce high peak capacities, and are sufficiently robust to operate for days at a time without significant losses in separation performance. Based on the results of this study we chose conditions for subsequent separations that required less than 400 bar operating pressure in the second dimension, but yielded a 2D peak capacity of about 3500 in 2 h. After 160 h of continuous operation of the LC × LC separation under these conditions (and about 20,000 injections into the 2D column) the 2D column had only lost about 18% of its initial isocratic efficiency. These results should motivate further development and implementation of such high performing and robust separations for the identification and quantification of peptides in a variety of application areas, including digests of therapeutic proteins such as monoclonal antibodies.

Dynamic pressure gradient modulation for comprehensive two-dimensional gas chromatography

We report the discovery, preliminary investigation, and demonstration of a novel form of differential flow modulation for comprehensive two-dimensional (2D) gas chromatography (GC×GC). Commercially available components are used to apply a flow of carrier gas with a suitable applied auxiliary gas pressure (Paux) to a T-junction joining the first (1D) and second (2D) dimension columns. The 1D eluate is confined at the T-junction, and introduced for 2D separation with a cyclic rhythm, dependent upon the relationship of the modulation period (PM) to the pulse width (pw), where pw is defined as the time interval when the auxiliary gas flow at the T-junction is off. We refer to this flow modulation technique as “dynamic pressure gradient modulation” (DPGM) since a pressure gradient oscillates with the PM along the 1D and 2D column ensemble providing temporary stop-flow conditions and fast 2D flow rates, resulting in 100% duty cycle and full modulation. A 90-component test mixture was used to evaluate the technique with a pw of 60 ms and a PM of 750 ms. The resulting peaks were narrow, with 2Wb ranging from about 20–180 ms. With an average 1Wb of 3 s and a 2nc of 10, a 2D peak capacity, nc,2D, for the 25 min separation was 5000. The detector response enhancement factor (DREF) is reported, defined as the peak height of the highest modulated 2D peak divided by the unmodulated 1D peak height (DREF = 2h/1h). The DREF ranged from about 7–87, depending on the 1Wb and 2Wb for a given analyte. A diesel sample was analyzed to demonstrate performance with a complex sample. Based upon the average 1Wb of 5 s and an average 2Wb of 168 ms, a nc,2D of 8640 was obtained for the 60 min diesel separation. Finally, the modulation principle was investigated as a function of PM, pw, and the volumetric flow rates, 1F and 2F. The measured 2Wb correlate well with the theoretical 2D injected width, given by 2Winj = (1F/2F) ·PM. However, the relevant 1F appears to be dictated by the 1D flow rate when no pressure is applied (during the pw interval), instead of 1F being the average flow rate on 1D (defined by the 1D dead time). The findings provide strong evidence for a differential flow modulation mechanism.

Development of Ultrafast Separations Using Negative Pulse Partial Modulation To Enable New Directions in Gas Chromatography.

Partial modulation via a pulse flow valve operated in the negative pulse mode is developed for high-speed one-dimensional gas chromatography (1D-GC), comprehensive two-dimensional (2D) gas chromatography (GC × GC), and comprehensive three-dimensional gas chromatography (GC3). The pulse flow valve readily provides very short modulation periods, PM, demonstrated herein at 100, 200, and 300 ms, and holds significant promise to increase the scope and applicability of GC instrumentation. The negative pulse mode creates an extremely narrow, local analyte concentration pulse. The reproducibility of the negative pulse mode is validated in a 1D-GC mode, where a pseudosteady-state analyte stream is modulated, and 8 analytes are baseline resolved (resolution, Rs ≥ 1.5) in a 200 ms window, providing a peak capacity, nc, of 14 at unit resolution ( Rs = 1.0). Additionally, the pulse width, pw, of the pulse flow valve "injection" relationship to peak width-at-base, wb, resolution between peaks and detection sensitivity are studied. To demonstrate the applicability to GC × GC, a high-speed separation of a 20-component test mixture of similar, volatile analytes is shown. Analytes were separated on the second-dimension column, 2D, with 2 wb ranging from 7 to 12 ms, providing an exceptional 2D peak capacity, 2 nc, of ∼12 using a modulation period ( PM) of 100 ms. Next, a 12 min separation of a diesel sample using a PM of 300 ms is presented. The 1 wb is ∼4 s, resulting in a 1 nc of ∼180, and 2 wb is ∼18 ms, resulting in a 2 nc of ∼17, thus achieving a nc,2D of ∼3000 in this rapid GC × GC diesel separation. Finally, GC3 with time-of-flight mass spectrometry (TOFMS) detection using a PM of 100 ms applied between the 2D and 3D columns is reported. Narrow third dimension, 3D, peaks with 3 wb of ∼15 ms were obtained, resulting in a GC3 peak capacity, nc,3D, of ∼35 000 in a 45 min separation.

Likelihood of total resolution in selective comprehensive two-dimensional liquid chromatography with parallel processing: Simulation and theory

The probability Pr(sLC×LC) that all peaks are separated by a resolution of 1.5 or more in selective comprehensive two-dimensional liquid chromatography (sLC × LC) is computed for simple model systems of 5 to 60 peaks and first-dimension (1D) gradient times of 100 to 2000 s. The computations include mimics of a commercial instrument, whose fixed second-dimension (2D) gradient time and use of one cycle time for initialization reduces Pr(sLC×LC) relative to an earlier report. For serial sLC × LC, in which a single device collects and transfers 1D multiplets to the second dimension, Pr(sLC×LC) under practical conditions is predicted to be only slightly larger than the probability of total resolution in LC × LC for separations of the same duration in each case. To increase Pr(sLC×LC), two model systems are proposed based on parallel processing, in which one device collects multiplets from the first separation while a second device simultaneously transfers fractions from previously collected multiplets to the second dimension for further separation. A sum of probabilities guideline is proposed by which optimal fixed 2D gradient times, ranging from 9.5 to 12 s, are found for both serial and parallel models. The increases of Pr(sLC×LC) based on parallel processing are modest; the largest is only 0.062 for one system and 0.106 for the other, relative to the serial model. A theory is derived that rationalizes the modesty of the increase, which was unexpected. It shows that Pr(sLC×LC) equals the probability of total resolution in the first dimension, plus the product of the probability that all 1D multiplets are transferred to the second dimension and the probability that all multiplets are separated in the second dimension. The theory shows that, although parallel processing is better than serial processing for multiplet transfer, the ability to leverage this gain is offset by the limited probability that all multiplets are then actually separated in the second dimension, which is only about 0.55 for conditions where the change from serial to parallel processing is most beneficial. With these findings in hand, two scenarios are examined for future consideration: one in which the 2D peak capacity is doubled, and another in which multiplets are always transferred to the second dimension. The latter shows considerable promise for increasing Pr(sLC×LC) substantially beyond its counterpart in LC × LC. For example, a 50% probability of separating all peaks in a 15-component mixture can be reached in 1150 s using LC × LC. The same probability can be reached in the same time for a sample with nearly twice as many components (27) in the case of sLC × LC, assuming transfer of all multiplets to the second dimension. These findings will be useful to those considering systematic approaches to developing 2D-LC methods for moderately complex mixtures, and to those interested in instrument development for 2D-LC.

Comprehensive two-dimensional gas chromatography using partial modulation via a pulsed flow valve with a short modulation period

Partial modulation via a pulsed flow valve for comprehensive two-dimensional (2D) gas chromatography (GC × GC) is demonstrated, producing narrow peak widths, 2Wb, on the secondary separation dimension, 2D, coupled with short modulation periods, PM, thus producing a high peak capacity on the 2D dimension, 2nc. The GC × GC modulator is a pulse flow valve that injects a pulse of carrier gas at the specified PM, at the connection between the primary, 1D, column and the 2D column. Using a commercially available pulse flow valve, this injection technique performs a combination of vacancy chromatography and frontal analysis, whereby each pulse disturbance in the analyte concentration profile as it exits the 1D column results in data that is readily converted into a 2D separation. A three-step process converts the raw data into a format analogous to a GC × GC separation, incorporating signal differentiation, baseline correction and conversion to a GC × GC chromatogram representation. A 115-component test mixture with a wide range of boiling points (36–372°C) of nine compound classes is demonstrated using modulation periods of PM = 50, 100, 250, and 500ms, respectively. For the test mixture with a PM of 250ms, peak shapes on 2D are symmetric with apparent 2Wb ranging from 12 to 45ms producing a 2nc of ~ 10. Based on the average peak width of 0.93s on the 1D separation for a time window of 400s, the 1D peak capacity is 1nc ∼ 430. Thus, the ideal 2D peak capacity nc,2D is 4300 or a peak capacity production of 650 peaks/min using the PM of 250ms. Additionally, for a PM of 50, 100 and 500ms, the 2nc are 4, 7, and 12, respectively. Retention times on 2D, 2tR, are reproducible having standard deviations less than 1ms. Finally, the processed data is shown to be quantitative, with an average RSD of 4.7% for test analytes.

In Silico Screening of Two-Dimensional Separation Selectivity for Ion Chromatography × Capillary Electrophoresis Separation of Low-Molecular-Mass Organic Acids.

A prerequisite for ordered two-dimensional (2D) separations and full utilization of the enhanced 2D peak capacity is selective exploitation of the sample attributes, described as sample dimensionality. In order to take sample dimensionality into account prior to optimization of a 2D separation, a new concept based on construction of 2D separation selectivity maps is proposed and demonstrated for ion chromatography × capillary electrophoresis (IC×CE) separation of low-molecular-mass organic acids as test analytes. For this purpose, 1D separation selectivity maps were constructed based on calculation of pairwise separation factors and identification of critical pairs for four IC stationary phases and 28 levels of background electrolyte pH in CE. The derived IC and CE maps were then superimposed and the effectiveness of the respective 2D separations assessed using an in silico approach, followed by testing examples of one successful and one unsuccessful 2D combination experimentally. The results confirmed the efficacy of the predictions, which require a minimal number of experiments compared to the traditional one-at-a-time approach. Following the same principles, the proposed framework can also be adapted for optimization of separation selectivity in various 2D combinations and for other applications.

Implications of phase ratio for maximizing peak capacity in comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry

The relationship between the phase ratio, β, of the primary (1D) and secondary (2D) separation dimensions of comprehensive two-dimensional (2D) gas chromatography (GC×GC) separations, and the implications of β on realization of maximal 2D peak capacity, nc,2D, are examined. A GC×GC chromatographic system with time-of-flight mass spectrometry, TOFMS, was otherwise held constant for the separation of a multi-component test mixture spanning a range of chemical functionalities, while only the β of the two analytical columns were changed, 1β for 1D and 2β for 2D. Six column sets were studied using common, commercially available β values. The β ratio, βR=1β/2β, is defined as a quantitative metric to facilitate this study. It is demonstrated that βR plays a key role in maximizing nc,2D. Overall, βR substantially affected nc,2D by influencing retention factors on the 2D column, 2k, and thereby changing the modulation period, PM, necessary for proper 2D column separations. The necessary changes to PM modify the modulation ratio, MR, which affects the 1D column peak widths and 1nc due to the impact of undersampling. Through changes to 1β, the range of 2k can be controlled, with subsequent effects to both 2nc and 1nc. These effects were opposite in direction, such that improvements to 2nc may result in declines in 1nc. It is observed that due to the pseudo-isothermal nature of the 2D separation, there are diminishing returns to extending the 2nc at the cost of 1nc. In this particular study, column set 3 (1D: 20m length, 250μm i.d., 0.25μm film; 2D: 2m, 180μm i.d., 0.2μm film; βR=1.11) with a PM of 3s provided the highest theoretical nc,2D of ∼8200, though this was at a relatively low MR of ∼1.8. Column set 2 (1D: 20m length, 250μm i.d., 0.5μm film; 2D: 2m, 180μm i.d., 0.2μm film; βR=0.56) with a PM of 1.5s provided a high theoretical nc,2D of ∼5800, at a much higher MR of ∼3.7. Though column set 2 had a lesser total peak capacity than column set 3, its higher MR suggests that by improving the 1D column efficiency (i.e., narrowing the 1D column peak widths) to improve 1nc, can result in an increased theoretical nc,2D.

Fully Automated Portable Comprehensive 2-Dimensional Gas Chromatography Device.

We developed a fully automated portable 2-dimensional (2-D) gas chromatography (GC x GC) device, which had a dimension of 60 cm × 50 cm × 10 cm and weight less than 5 kg. The device incorporated a micropreconcentrator/injector, commercial columns, micro-Deans switches, microthermal injectors, microphotoionization detectors, data acquisition cards, and power supplies, as well as computer control and user interface. It employed multiple channels (4 channels) in the second dimension (2D) to increase the 2D separation time (up to 32 s) and hence 2D peak capacity. In addition, a nondestructive flow-through vapor detector was installed at the end of the 1D column to monitor the eluent from 1D and assist in reconstructing 1D elution peaks. With the information obtained jointly from the 1D and 2D detectors, 1D elution peaks could be reconstructed with significantly improved 1D resolution. In this Article, we first discuss the details of the system operating principle and the algorithm to reconstruct 1D elution peaks, followed by the description and characterization of each component. Finally, 2-D separation of 50 analytes, including alkane (C6-C12), alkene, alcohol, aldehyde, ketone, cycloalkane, and aromatic hydrocarbon, in 14 min is demonstrated, showing the peak capacity of 430-530 and the peak capacity production of 40-80/min.

High temperature diaphragm valve-based comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry

A high temperature diaphragm valve-based two-dimensional (2D) gas chromatography (GC×GC) with time-of-flight mass spectrometry (TOFMS) instrument, with the valve mounted directly in the GC oven, is demonstrated with separations up to 325°C. Use of the diaphragm valve allowed for the use of uncoupled carrier gas flows for 1D (first column dimension) and 2D (second column dimension), with a 1D flow rate of 1ml/min, and a 2D flow rate 3ml/min. The 3ml/min flow rate on 2D was selected to ensure compatibility with most TOFMS detectors. For valve-based modulation, a 5µl sample loop coupled with a 60ms pulse width was selected, providing sufficient sensitivity concurrent with an acceptable 2D peak capacity. A 44-component mixture of alkanes, alcohols, and polyaromatic hydrocarbons (including n-alkanes of heptane to triacontane) whose boiling points range from 98°C to 450°C was used to initially study instrument performance. For a 120min separation and a modulation period PM of 2s, average peak widths-at-base of 10s and 94ms were achieved for the alkanes on the 1D and 2D dimensions, respectively. Hence, the 1D peak capacity is 1nc~700, and the 2D peak capacity can in principle be 2nc~20. Thus the ideal 2D peak capacity could in principle approach nc,2D~14,000 using the 120min 1D run time. The limit of detection (LOD) for docosane, a representative analyte, was determined to be 1.5ppm injected concentration when 2µl of liquid sample was injected with a 20:1 split. A separation of diesel fuel demonstrated the practical utility of the instrument with this complex sample using a relatively fast run time of 20min and a short modulation period PM of 1s. Average peak widths-at-base of 5.4s and 166ms were achieved on the 1D and 2D dimensions, respectively. This yielded a 1nc~220 and a 2nc~ 6. Therefore, the 2D peak capacity is nc,2D~1320 with the 20min diesel separation.

High temperature diaphragm valve-based comprehensive two-dimensional gas chromatography

A high-temperature diaphragm valve-based comprehensive two-dimensional gas chromatography (GC×GC) instrument is demonstrated which readily allows separations up to 325°C. Previously, diaphragm valve-based GC×GC was limited to 175°C if the valve was mounted in the oven, or limited to 265°C if the valve was faced mounted on the outside of the oven. A new diaphragm valve has been commercially developed, in which the temperature sensitive O-rings that previously limited the separation temperatures have been replaced with Kalrez O-rings, a perfluoroelastomer, allowing for significantly higher temperatures permitting a greater range of volatile and semi-volatile compounds to be readily separated. In the current investigation, a separation temperature up to 325°C is demonstrated with the valve mounted directly in the oven. Since the temperature limit for most commonly used GC columns is at or below 325°C, the scope of diaphragm valve-based GC×GC is now dramatically broadened to encompass a majority of all column stationary phase chemistries. A 44-component mixture of alkanes, alcohols, and polyaromatic hydrocarbons is used to study this new configuration whose boiling points range from 98°C (n-heptane) to 450°C (n-triacontane). For the test mixture using a modulation period PM of 1.0s, peak shapes on second dimension separations, 2D, are symmetric with average widths at base of 79.4ms, producing a 2D peak capacity of 2nc∼12. Based on the average peak width of 2.4s for the first dimension separation with a run time of 32.5min, the 1D peak capacity is 1nc∼800. Thus, the ideal two-dimensional peak capacity nc,2D is 9600. Little variation in within-analyte 2D peak width was observed with an average %RSD of less than 3.0%. Furthermore, retention time on 2D was very reproducible with an average %RSD less than 0.5%. Measured peak areas (sum of all 2D peaks for given analyte) had an average %RSD of 4.4%. The transfer fraction from 1D to 2D was experimentally determined to be ∼30%, while the detection sensitivity for valve-based GC×GC was ∼8 times higher than one dimensional GC due to zone compression. After a year of use with temperatures consistently up to 325°C, there has been no deterioration of the valve or its performance for GC×GC. Separations of vacuum pump oil and orange oil are also reported to demonstrate practical utility.

Improving peak capacity in fast online comprehensive two-dimensional liquid chromatography with post-first-dimension flow splitting.

The use of flow splitters between the two dimensions in online comprehensive two-dimensional (2D) liquid chromatography (LC × LC) has not received very much attention, in comparison with their use in 2D gas chromatography (GC × GC), where they are quite common. In principle, splitting the flow after the first dimension column and performing online LC × LC on this constant fraction of the first dimension effluent should allow the two dimensions to be optimized almost independently. When there is no flow splitting, any change in the first-dimension flow rate has an immediate impact on the second dimension. With a flow splitter, one could, for example, double the flow rate into the first dimension column and perform a 1:1 flow split without changing the sample loop size or the sampler's collection time. Of course, the sensitivity would be diminished, but this can be partially compensated through the use of a larger injection; this will likely only amount to a small price to pay for this increased resolving power and system flexibility. Among other benefits, we found a 2-fold increase in the corrected 2D peak capacity and the number of observed peaks for a 15-min analysis time, using a post-first-dimension flow splitter. At a fixed analysis time, this improvement results primarily from an increase in the gradient time, resulting from the reduced system re-equilibration time, and, to a smaller extent, it is due to the increased peak capacity achieved by full optimization of the first dimension.

Generation and Limitations of Peak Capacity in Online Two-Dimensional Liquid Chromatography

The different operating conditions of an online two-dimensional liquid chromatographic separation (2D-LC), such as the length of the column, the linear velocity and the composition of the mobile phase used in the second dimension, its initial organic content if this separation is carried out in gradient elution, the number of fractions of the first column eluent collected, and the analysis time of the first dimension all affect the achievable separation power of 2D-LC online systems. The influences of these factors on the separation performance were investigated, and an equation was derived for the calculation of the achievable peak capacity in online 2D-LC assuming (1) that the option of undersampling the first-dimension separation is acceptable, (2) that the solutes follow linear-solvent-strength behavior, and (3) that all the separations are made in gradient elution. This theoretical discussion shows that (1) highly efficient separations made with online 2D-LC require the second-dimension peaks to be very narrow, (2) the separation power of 2D-LC systems is maximum for an optimum number of fractions collected in the first dimension, (3) higher peak capacities can be achieved by using shorter second-dimension columns and collecting a relatively large number of fractions, (4) the achievable 2D peak capacity is maximum for a certain eluent flow rate and column length of the second-dimension column, and (5) the maximum achievable peak capacity increases with decreasing velocity and initial organic content of the second-dimension eluent. As a consequence, due to the time restriction of the second-dimension gradient time, online 2D-LC schemes cannot realistically afford peak capacities exceeding 10,000, even if they are implemented with exceptionally efficient columns and if long analysis times are accepted.