High-efficiency liquid chromatography–mass spectrometry separations with 50 mm, 250 mm,and 1 m long polymer-based monolithic capillary columns for the characterization of complex proteolytic digests

In this study, high-efficiency LC–MS/MS separations of complex proteolytic digests are demonstrated using 50 mm, 250 mm, and 1 m long poly(styrene-co-divinylbenzene) monolithic capillary columns. The chromatographic performance of the 50 and 250 mm monoliths was compared at the same gradient steepness for gradient durations between 5 and 150 min. The maximum peak capacity of 400 obtained with a 50 mm column, increased to 485 when using the 250 mm long column and scaling the gradient duration according column length. With a 5-fold increase in column length only a 20% increase in peak capacity was observed, which could be explained by the larger macropore size of the 250 mm long monolith. When taking into account the total analysis time, including the dwell time, gradient time and column equilibration time, the 50 mm long monolith yielded better peptide separations than the 250 mm long monolithic column for gradient times below 80 min (n_c = 370). For more demanding separation the 250 mm long monolith provided the highest peak production rate and consequently higher sequence coverage. For the analysis of a proteolytic digest of Escherichia coli proteins a monolithic capillary column of 1 m in length was used, yielding a peak capacity of 1038 when applying a 600 min gradient.     

Fast ion chromatography using short anion exchange columns

A fast ion chromatographic system is described which uses shorter column lengths and compares various eluent profiles in order to maximise the performance without sacrificing the chromatographic resolution. Both isocratic and gradient elution profiles were considered to find the most efficient mode of separation. The separation and determination of seven target anions (chloride, chlorate, nitrate, chromate, sulfate, thiocyanate and perchlorate) was achieved using a short (4 mm ID, 50 mm long) column packed with Dionex AS20 high-capacity anion exchange material. A hydroxide eluent was used at an initial concentration of 25 mM (at a flow-rate of 1.0 mL/min) and two performance maxima were found. The maximum efficiency occurred at a normalised gradient ramp rate of 5 mM/t₀, resulting in a peak capacity of 16, while the fastest separation (<3 min) occurred at a normalised ramp rate of 30 mM/t₀. The retention time, peak width and resolution using the different eluent profiles on varying column lengths is also compared. Further investigations in this study determined that the highest peak capacity separation under gradient conditions could be approximated using an isocratic separation. The advantage of using this novel approach to approximate the maximum efficiency separation removes the need for column re-equilibration that is required for gradient elution resulting in faster analyses and enhanced sample throughput, with benefits in particular for multidimensional chromatography.     

Fast gradient screening of pharmaceuticals with 5 cm long, narrow bore reversed-phase columns packed with sub-3 μm core-shell and sub-2 μm totally porous particles

The performance of 5 cm long narrow-bore columns packed with 2.6-2.7 μm core-shell particles and a column packed with 1.7 μm totally porous particles was compared in very fast gradient separations of polar neutral active pharmaceutical compounds. Peak capacities as a function of flow-rate and gradient time were measured. Peak capacities around 160-170 could be achieved within 25 min with these 5 cm long columns. The highest peak capacity was obtained with the Kinetex column however it was found that as the flow-rate increases, the peak capacity of the new Poroshell-120 column is getting closer to that obtained with the Kinetex column. Considering the column permeability, peak capacity per unit time and per unit pressure was also calculated. In this comparison the advantage of sub-3 μm core-shell particles is more significant compared to sub-2 μm totally porous particles. Moreover it was found that the very similar sized (d_p = 2.7 μm) and structured (ρ = 0.63) new Poroshell-120 and the earlier introduced Ascentis Express particles showed different efficiency. Results obtained showed that the 5 cm long narrow bore columns packed with sub-3 μm core-shell particles offer the chance of very fast and efficient gradient separations, thus these columns can be applied for fast screening measurements of routine pharmaceutical analysis such as cleaning validation.     

Separation of intact proteins on porous layer open tubular (PLOT) columns

Porous layer open tubular (PLOT) polystyrene divinylbenzene columns have been used for separating intact proteins with gradient elution. The 10 μm I.D. × 3 m columns were easily coupled to standard liquid chromatography–mass spectrometry (LC–MS) instrumentation with commercially available fittings. Standard proteins separated on PLOT columns appeared as narrow and symmetrical peaks with good resolution. Average peak width increased linearly with gradient time (t_G) from 0.14 to 0.33 min (t_G 20 and 120 min, respectively) using a 3 m column. With shorter columns, peak widths were larger and increased more steeply with gradient time. Theoretical peak capacity (n_c) increased with column length (tested up to 3 m). The n_c increased with t_G until a plateau was reached. The highest peak capacity achieved (n_c = 185) was obtained with a 3 m column, where a plateau was reached with t_G 90 min. The within- and between column retention time repeatabilities were below 0.6% and below 2.5% (relative standard deviation, RSD), respectively. The carry-over following injection of 0.5 ng per protein was less than 1.1%. The retention time dependence on column temperature was investigated in the range 20–50 °C. Proteins in a skimmed milk sample were separated using the method.     

Probing the kinetic performance limits for ion chromatography. I. Isocratic conditions for small ions

The first use of the kinetic plot method to characterise the performance of ion-exchange columns for separations of small inorganic anions is reported. The influence of analyte type (mono- and divalent), particle size (5 and 9 μm), temperature (30 and 60 °C) and maximum pressure drop upon theoretical extrapolations was investigated using data collected from anion-exchange polymeric particulate columns. The quality of extrapolations was found to depend upon the choice of analyte, but could be verified by coupling a series of columns to demonstrate some practical solutions for ion chromatography separations requiring relatively high efficiency. Separations of small anions yielding 25–40,000 theoretical plates using five serially connected columns (9 μm particles) were obtained and yielded deviations of <15% from the kinetic plot predictions. While this approach for achieving high efficiencies results in a very long analysis time (t₀ = 21 min), separations yielding approximately 10,000 theoretical plates using two serially connected columns (t₀ < 5 min) were shown to be more practically useful for isocratic separations when compared to use of a single column operated at optimum linear velocity (t₀ > 10 min).     

Some solutions to obtain very efficient separations in isocratic and gradient modes using small particles size and ultra-high pressure

The UHPLC strategy which combines sub-2 μm porous particles and ultra-high pressure (>1000 bar) was investigated considering very high resolution criteria in both isocratic and gradient modes, with mobile phase temperatures between 30 and 90 °C. In isocratic mode, experimental conditions to reach the maximal efficiency were determined using the kinetic plot representation for ΔP_max = 1000 bar. It has been first confirmed that the molecular weight of the compounds (MW) was a critical parameter which should be considered in the construction of such curves. With a MW around 1000 g mol⁻¹, efficiencies as high as 300,000 plates could be theoretically attained using UHPLC at 30 °C. By limiting the column length to 450 mm, the maximal plate count was around 100,000. In gradient mode, the longest column does not provide the maximal peak capacity for a given analysis time in UHPLC. This was attributed to the fact that peak capacity is not only related to the plate number but also to column dead time. Therefore, a compromise should be found and a 150 mm column should be preferentially selected for gradient lengths up to 60 min at 30 °C, while the columns coupled in series (3× 150 mm) were attractive only for t_grad > 250 min. Compared to 30 °C, peak capacities were increased by about 20–30% for a constant gradient length at 90 °C and gradient time decreased by 2-fold for an identical peak capacity.     

Axial temperature gradient and mobile phase gradient in microcolumn high-performance liquid chromatography

The effect of axial temperature gradient (ATG) along a microcolumn on the separation performance at both isocratic and gradient elution mode was investigated. A thermostat system was designed to form an ATG along the packed column. Polycyclic aromatic hydrocarbons (PAHs) were separated on a 0.53 mm  × 150 mm i.d. 5 μm C₁₈ microcolumn, with water and acetonitrile as mobile phase. The separation results obtained at mobile phase gradient (MPG) and ATG in microcolumn HPLC were compared with the results performed at ambient conditions. Extrapolated curves of peak width at half height (wh)versus lnk showed that wh is narrower at the same retention time when ATG was applied in addition to MPG. The column efficiency was enhanced 20-30% and the resolution was slightly reduced because of reduction of selectivity at elevated temperature at ATG condition. The RSD of retention time in ATG mode was less than 2.5%.     

Compact type-I coil planet centrifuge for counter-current chromatography

A compact type-I coil planet centrifuge has been developed for performing counter-current chromatography. It has a revolution radius of 10 cm and a column holder height of 5 cm compared with 37 and 50 cm in the original prototype, respectively. The reduction in the revolution radius and column length permits application of higher revolution speed and more stable balancing of the rotor which leads us to learn more about its performance and the future potential of type-I coil planet centrifuge. The chromatographic performance of this apparatus was evaluated in terms of retention of the stationary phase (S_f), peak resolution (R_s), theoretical plate (N) and peak retention time (t_R). The results of the experiment indicated that increasing the revolution speed slightly improved both the retention of the stationary phase and the peak resolution while the separation time is remarkably shortened to yield an excellent peak resolution at a revolution speed of 800 rpm. With a 12 ml capacity coiled column, DNP-DL-glu, DNP-β-ala and DNP-l-ala were resolved at R_s of 2.75 and 2.16 within 90 min at a flow rate of 0.4 ml/min. We believe that the compact type-I coil planet centrifuge has a high analytical potential.     

Optimizing the peak capacity per unit time in one-dimensional and off-line two-dimensional liquid chromatography for the separation of complex peptide samples

To obtain the best compromise between peak capacity and analysis time in one-dimensional and two-dimensional (2D) liquid chromatography (LC), column technology and operating conditions were optimized. The effects of gradient time, flow rate, column temperature, and column length were investigated in one-dimensional reversed-phase (RP) gradient nano-LC, with the aim of maximizing the peak per unit time for peptide separations. An off-line two-dimensional LC approach was developed using a micro-fractionation option of the autosampler, which allowed automatic fractionation of peptides after a first-dimension ion-exchange separation and re-injection of the fractions onto a second-dimension RP nano-LC column. Under the applied conditions, which included a preconcentration/desalting time of 5 min, and a column equilibration time of 12.5 min, the highest peak capacity per unit time in the 2D-LC mode was obtained when applying a short (10 min) first-dimension gradient and second-dimension RP gradients of 20 min duration. For separations requiring a maximum peak capacity of 375, one-dimensional LC was found to be superior to the off-line strong cation-exchange/×/RPLC approach in terms of analysis time. Although a peak capacity of 450 could be obtained in one-dimensional LC when applying 120-min gradients on 500-mm long columns packed with 3-μm particles, for separations requiring a peak capacity higher than 375 2D-LC experiments provide a higher peak capacity per unit time. Finally, the potential of off-line 2D-LC coupled to tandem mass spectrometry detection is demonstrated with the analysis of a tryptic digest of a mixture of nine proteins and an Escherichia coli digest.     

Development of fast enantioselective gas-chromatographic analysis using gas-chromatographic method-translation software in routine essential oil analysis (lavender essential oil)

The study aimed to find the best trade-off between separation of the most critical peak pair and analysis time, in enantioselective GC–FID and GC–MS analysis of lavender essential oil, using the GC method-translation approach. Analysis conditions were first optimized for conventional 25 m × 0.25 mm inner diameter (d_c) column coated with 6^I–VII-O-tert-butyldimethylsilyl-2^I–VII-3^I–VII-O-ethyl-β-cyclodextrin (CD) as chiral stationary phase (CSP) diluted at 30% in PS086 (polymethylphenylpolysiloxane, 15% phenyl), starting from routine analysis. The optimal multi-rate temperature program for a pre-set column pressure was determined and then used to find the pressures producing the efficiency-optimized flow (EOF) and speed-optimized flow (SOF). This method was transferred to a shorter narrow-bore (NB) column (11 m × 0.10 mm) using method-translation software, keeping peak elution order and separation. Optimization of the enantioselective GC method with the translation approach markedly reduced the analysis time of the lavender essential oil, from about 87 min with the routine method to 40 min with an optimal multi-rate temperature program and initial flow with a conventional inner diameter column, and to 15 min with FID as detector or 13.5 min with MS with a corresponding narrow-bore column, while keeping enantiomer separation and efficiency.     

Effect of an open tube in series with a packed capillary column on liquid chromatographic performance: The influence of particle diameter,temperature, and system pressure

A postcolumn reactor or a simple open tube connecting a capillary column to, for example, a mass spectrometer affects the performance of a capillary liquid chromatography system in two ways: stealing pressure from the column and adding band-spreading. This effect is especially intolerable in fast separations. Our calculations show that in the presence of a 25 μm radius postcolumn reactor, column (50 μm radius) efficiency (number of theoretical plates) is severely reduced by more than 75% with a t₀ of 10 s and a particle diameter from 1 to 5 μm for unretained solutes at room temperature. Therefore, it is necessary to minimize the reactor's effect and to improve the column efficiency by optimizing postcolumn conditions. We derived an equation that defines the observed number of theoretical plates (N_obs) taking into account the two effects stated above, which is a function of the maximum pressure P_m, the particle diameter d_p, the reactor radius a_r, the column radius a_c, the desired dead time t₀, the column temperature T and zone capacity factor k″. Poppe plots were obtained by calculations using this equation. The results show that for a t₀ shorter than 18 s, a P_m of 4000 psi, and a d_p of 1.7 μm, a 5 μm radius reactor has to be used. Such a small reactor is difficult to fabricate. Fortunately, high temperature helps to minimize the reactor effect so that reactors with manageable radius (larger than 12.5 μm) can be used in many practical conditions. Furthermore, solute retention diminishes the influence of a postcolumn reactor. Thus, a 12.5 μm reactor supersedes a 5 μm reactor for retained solutes even at a t₀ of 5 s (k″ > 3.8, or k′ > 2.0).     

Characterization of new types of stationary phases for fast and ultra-fast liquid chromatography by signal processing based on AutoCovariance Function: A case study of application to Passiflora incarnata L. extract separations

In this study, a comparative investigation was performed of HPLC Ascentis^® (2.7 μm particles) columns based on fused-core particle technology and Acquity^® (1.7 μm particles) columns requiring UPLC instruments, in comparison with Chromolith™ RP-18e columns. The study was carried out on mother and vegetal tinctures of Passiflora incarnata L. on one single or two coupled columns. The fundamental attributions of the chromatographic profiles are evaluated using a chemometric procedure, based on the AutoCovariance Function (ACVF). Different chromatographic systems are compared in terms of their separation parameters, i.e., number of total chemical components (m_tot), separation efficiency (σ), peak capacity (n_c), overlap degree of peaks and peak purity. The obtained results show the improvements achieved by HPLC columns with narrow size particles in terms of total analysis time and chromatographic efficiency: comparable performance are achieved by Ascentis^® (2.7 μm particle) column and Acquity^® (1.7 μm particle) column requiring UPLC instruments. The ACVF plot is proposed as a simplified tool describing the chromatographic fingerprint to be used for evaluating and comparing chemical composition of plant extracts by using the parameters D% – relative abundance of the deterministic component – and c_EACF – similarity index computed on ACVF.     

Fingerprint analysis of Dioscorea nipponica by high-performance liquid chromatography with evaporative light scattering detection

High-performance liquid chromatographic method (HPLC) with evaporative light scattering detection (ELSD) coupled with microwave-assisted extraction (MAE) as an efficient sample preparation technique has been developed for fingerprint analysis of Dioscorea nipponica. The samples were separated with an Agilent C8 column using water (A) and acetonitrile (B) under gradient conditions (0-10 min, linear gradient 20-40% B; 10-12 min, linear gradient 40-42% B; 12-25 min, isocratic 42% B) as the mobile phase at a flow rate of 1 mL min⁻¹ within 22 min. The ELSD conditions were optimized at nebulizer-gas flow rate 2.7 L min⁻¹ and drift tube temperature 90 °C. Precision experiments showed relative standard deviation (R.S.D.) of peak area and retention time were better than 2.5%; inter-day and intra-day variabilities showed that R.S.D. was ranged from 0.78% to 4.74%. Limit of detection was less than 50 μg mL⁻¹ and limit of quantification was less than 80 μg mL⁻¹. Accuracy validation showed that average recovery was between 97.39% and 104.07%. The method was validated to achieve the satisfactory precision and recovery. Relative retention time and relative peak area were used to identify the common peaks for fingerprint analysis. There are nine common peaks in the fingerprint. The quality of seven batches of D. nipponica samples was evaluated to be qualified or unqualified by the parameters “difference” and “total difference” of common peaks. Furthermore, the contents of important medicinal compounds (dioscin, prodioscin and gracillin) in different batches of D. nipponica samples were determined simultaneously using the developed HPLC-ELSD method. The results indicated variation of the herb quality which might be related to different producing area, growing condition, climate, harvest time, drug processing and so on. The developed analytical procedure was proved to be a reliable and rapid method for the quality control of D. nipponica.     

Protein separation using toroidal columns by type-J synchronous counter-current chromatography towards preparative separation

Separation of large bioactive molecules such as proteins, DNAs and RNAs using aqueous two-phase systems (ATPSs) and liquid–liquid partition-based counter-current chromatography (CCC) can avoid risks of sample loss and denaturation, and greatly reduce processing time. We have constructed toroidal columns (length 26–140 m, column volume 51–280 ml, bore size 1.6 mm) suitable for mounting onto a commercially available preparative CCC apparatus. With the use of an ATPS containing 12.5% (w/w) PEG1000 and 12.5% (w/w) K₂HPO₄ and at a rotational speed of 800 rpm for the rotor of the CCC device, the lower phase (i.e. the phosphate-enriched phase) has been used as the mobile phase and a pair of proteins, myoglobin and lysozyme, as model proteins for demonstrating the separation capability of the CCC system. For a toroidal column with a length of 53.5m and a column volume of 107.5 ml, and operated for the Coriolis force parallel flow mode at 0.62 ml/min, protein sample loading (containing 2.2 mg/ml myoglobin and lysozyme, respectively) at 1.7% and 7.4% to the column volume led to peak resolution (with theoretical plate number TP and stationary phase retention S_f shown in the parenthesis) of R_s = 1.5 (N = 211 and N = 113 TP for myoglobin and lysozyme, respectively, and S_f = 45.0%), and R_s = 1.4 (218 and 152 TP, and S_f = 34.0%). However, further increase of the loading to 13% failed to separate the two proteins. Although proteins eluted at positions predictable from the distribution coefficients, they showed broader peaks when compared with small dipeptides under identical CCC operating conditions. This confirms that the molecular weight of the partitioned species is an important factor causing peak broadening on CCC chromatograms. These results paved the way for further scaling-up toroidal CCC columns for processing larger quantities of samples containing large biomolecules.     

Metabolism profile of scutellarin in urine following oral administration to rats by ultra performance liquid chromatography coupled to time-of-flight mass spectrometry

Scutellarin, a flavone glucuronide of 5,6,4′-trihydroxyflavone-7-O-glucoronide, is the main active component of the traditional Chinese botanic drug Erigeron breviscapus (Vant.) Hand.-Mazz. In this study, a method based on ultra performance liquid chromatography coupled with a time-of-flight mass spectrometer (UPLC/TOF MS) was established and validated to profile the metabolites of scutellarin in Sprague-Dawley rat urine following oral administration of single dose of scutellarin at 80.8 mg/kg. The column utilized was an Acquity BEH C18 (150 mm × 2.1 mm, 1.7 μm). The mobile phase was 0.2% formic acid and acetonitrile with gradient condition. Two standard curves of scutellarin were obtained for the concentration range of 1.065-10.65 μg/mL and 10.65-63.92 μg/mL, respectively. By automating the data processing of the software Masslynx developed by Waters Ltd., 17 metabolites of scutellarin were found and determined in rat urine, with the corresponding reactions in vivo such as isomerism, reduction, methylation, glucuronide conjugation, hydroxylation, hydroxylation and methylation, etc., most of which were discovered for the first time. For most metabolites, the time (T_p) of peak excretion was 8-12 h. Calculated as scutellarin, the cumulative urine excretion rate of the metabolites was 1.93%.     

Development of a new analytical method for online simultaneous qualitative determination of aluminium (free aluminium ion, aluminium-fluoride complexes) by HPLC-FAAS

The paper presents a novel method for simultaneous online examination of inorganic forms of aluminium: AlF₂⁺, AlF^2+, and Al³⁺ by means of the high performance liquid chromatography hyphenated with a detection by the atomic absorption spectrometry with flame atomization (HPLC-FAAS) without post-column reaction. The application of optimization procedure conditions of chromatographic separation of inorganic forms of aluminium was achieved by the analytical column IonPac CS5A (Dionex) with guard column IonPac CG5A (Dionex) and an aqueous ammonium chloride mobile phase, at pH about 3 with gradient elution. The separation of Al forms with nominal charge of 1+, 2+, 3+ required a run time of less than 8 min during a single analysis. The proposed method has been successfully used for the examination of aluminium forms formation AlF_n⁽³⁻ⁿ⁾⁺ in environmental samples.     

Investigation of the validity of the kinetic plot method to predict the performance of coupled column systems operated at very high pressures under different thermal conditions

The present study investigates how strong the kinetic plot method is influenced by the changes in plate height, retention factor and apparent column permeability that arise under conditions of very high pressure. More precisely, the study investigates how well a set of performance measurements conducted on a single short column can be used to predict the performance of a long sequence of coupled columns. This has been investigated for the two practically most relevant thermal conditions, i.e., that of a forced-air oven and that of a still-air oven. Measuring column performance data for acetophenone and benzene on a series of coupled 3.5 μm columns that could be operated up to 1000 bar, it was found that the kinetic plot method provides accurate predictions of time versus efficiency for the still-air oven systems, over the entire range of investigated pressures and column lengths (up to 60 cm), provided k′ and K_v0 are evaluated at the maximal pressure. For the forced-air oven which leads to worse performances than the still-air oven, the kinetic plot prediction is less accurate, partly because the thermal conditions (near-isothermal) tend to vary if the number of coupled columns increases. The fact that the thermal conditions of the column wall might vary with the column length is an additional complexity making very-high pressure separations less predictable and harder to interpret and model.     

Robustness study of a reversed-phase liquid chromatographic method for the analysis of carboxylic acids in industrial reaction mixtures

The robustness study of the reversed-phase liquid chromatographic method developed for the quantitative analysis of carboxylic acids is a real asset to prepare method transfer because it provides an indication of its reliability during routine use. Indeed, it was possible to predict the consequences of small variations in operating conditions on the responses. The design of experiments approach was applied to model the effects and interactions of a high number of factors varying simultaneously with a limited number of runs. First we identified the factors which potentially affect the chromatographic responses used for carboxylic acids quantitation: detection wavelength (λ), column temperature (T), acetonitrile ratio in mobile phase (Me), duration of the plateau before the gradient (L) and gradient slope (S). Then we estimated the order of magnitude of realistic variations to assign factor levels. Finally a central composite design was carried out around the nominal conditions defined during method optimization. The statistical treatment of responses (retention factors, and concentrations) showed that the column temperature, the acetonitrile ratio in the mobile phase, the duration of the plateau before the gradient and the gradient slope were the most influent factors. The building of the robust domain from response-surfaces allowed us to give tolerance limits for the factors (216 nm < λ < 222 nm, 49.3 °C < T < 51.4 °C, 4.90% < Me < 5.18%, v/v, 4.5 min < L < 5.4 min, 9% < S < 11%) for which the performances of the method were maintained.     

Probability of failure of the watershed algorithm for peak detection in comprehensive two-dimensional chromatography

The watershed algorithm is the most common method used for peak detection and integration in two-dimensional chromatography. However, the retention time variability in the second dimension may render the algorithm to fail. A study calculating the probabilities of failure of the watershed algorithm was performed. The main objective was to calculate the maximum second-dimension retention time variability, Δ²t_R,crit, above which the algorithm fails. Several models to calculate Δ²t_R,crit were developed and evaluated: (a) exact model; (b) simplified model and (c) simple-modified model. Model (c) gave the best performance and allowed to deduce an analytical expression for the probability of failure of the watershed algorithm as a function of experimental Δ²t_R, modulation time and peak width in the first and second dimensions. It could be demonstrated that the probability of failure of the watershed algorithm under normal conditions in GC × GC is around 15–20%. Small changes of Δ²t_R, modulation time and/or peak width in the first and second dimension could induce subtle changes in the probability of failure of the watershed algorithm. Theoretical equations were verified with experimental results from a diesel sample injected in GC × GC and were found to be in good agreement with the experiments.     

Mobile phase selection for the combined use of liquid chromatography–inductively coupled plasma mass spectrometry and electrospray ionisation mass spectrometry

Four different organic solvents: dimethylformamide, 1,4-dioxane, n-propanol and ethanol were evaluated as alternative organic modifiers to acetonitrile for liquid chromatography (LC) separations. The aim was to establish common sets of chromatographic conditions that could be applied for LC hyphenation to inductively coupled plasma mass spectrometry (ICPMS) as well as to electrospray ionization MS (ESIMS). The approach was to evaluate candidate solvents that, compared to acetonitrile, potentially could give improved analytical performance (low solvent vapor loading, maximized analyte sensitivity and minimized carbon depositions on instrumental parts) in ICPMS analysis while retaining chromatographic and ESIMS performances. The study showed that dimethylformamide, 1,4-dioxane, n-propanol and ethanol all can be advantageous chromatographic modifiers for LC–ICPMS analysis, giving superior performance compared to acetonitrile. For the combined use of LC–ICPMS and LC–ESIMS with a common set of chromatographic conditions, n-propanol gave the best overall performance. The ¹⁹⁵Pt⁺ signal in ICPMS was continuously monitored during a 0–60% organic solvent gradient and at 25% of organic modifier, 100% of the signal obtained at the gradient start was preserved for n-propanol compared to only 35% of the signal when using acetonitrile. Platinum detection limits were 5–8 times lower using n-propanol compared with acetonitrile. Signal-to-noise ratio in continuous ESIMS signal measurements was 100, 90 and 110 for a 100 μg/ml solution of leucine–enkephaline using acetonitrile, ethanol and n-propanol, respectively. Chromatographic efficiency in reversed phase separations was preserved for n-propanol compared to acetonitrile for the analysis of the whole protein cytochrome C and the peptide bacitracin on a column with particle and pore sizes of 5 μm and 300 Å, but slightly deteriorated for the separation of the peptides leucine–enkephaline and bacitracin on a 3 μm and 90 Å column as the peak width at half height for both peptides increased by a factor of two. The performance on the smaller dimensioned column could however be improved by running the separations at 40 °C.