Method and apparatus for multi-transition monitoring

Abstract

The present invention discloses a method for performing multi-transition monitoring using a liquid chromatography mass spectrometry device (111). The method comprises the following steps: a) determining at least one data set (112) from at least one database (114), the data set (112) comprising at least one reference measurement (116) of at least one transition of at least one analyte performed using the liquid chromatography mass spectrometry device (111); b) determining at least one reference peak information of the transition of the analyte using an initial setting of a measurement window (120), wherein the measurement window (120) is defined by a time range of a retention time; c) determining an actual setting of the measurement window (120) taking into account the reference peak information, wherein the determination comprises adjusting the time range; d) measuring the transition of the analyte using the liquid chromatography mass spectrometry device (111), and determining the measured peak information of the transition of the analyte using the actual setting of the measurement window (120).

Description

Technical Field

[0001] The present invention relates to a method and apparatus for multi-transition monitoring using mass spectrometry, in particular liquid chromatography and mass spectrometry. Background Art

[0002] Multiple transition monitoring (MRM) during an LC run reduces the retention time for each transition as the number of observers increases. Known methods such as scheduled MRM use only the time range in the LC run that is relevant to a specific transition. For example, analyte X produces peak Z at time x by MRM Y, and therefore only specific relevant times can be recorded, such as x-15s to x+15s. The specific time range is usually defined relative to a specific time around the peak maximum of the peak, usually called the retention time.

[0003] US 2017 / 0328874 A1 describes a chromatographic mass spectrometer, comprising: an MSn-1 analysis setter, which is used to set an analysis execution period for performing MSn-1 analysis, an execution time for analysis, and a loop time; an analysis period divider, which is used to divide the analysis period into multiple segments according to the number of MSn-1 analyses to be performed within the same time window or changes in analysis conditions; an MSn analysis setter, which is used to perform MSn-1 analysis to obtain mass spectrum data and to plan MSn analysis, and ions corresponding to peaks that meet the set conditions are designated as precursor ions; an MSn analysis execution time allocator, which is used to allocate a time period for performing MSn analysis in each segment, which time period is calculated by subtracting the event execution time from the loop time; and an analysis executor, which is used to repeatedly perform MSn-1 analysis and MSn analysis in each segment.

[0004] WO 2017 / 216934 A1 describes an analysis schedule that is pre-created so that multiple liquid chromatography streams can operate in parallel and a mass spectrometer can collect data at the timing of each component elution. A control unit controls so as to: divide the time required for analyzing each sample in multiple liquid chromatography systems into pre-collection time, collection period time, and post-collection time; search for and assign time positions where the collection period times in the liquid chromatography units do not overlap; determine the start time for the multiple liquid chromatography units, thereby creating an analysis schedule; and perform analysis thereafter. The control unit further: stores a parameter set for changing the component elution time, adjusts the analysis parameters so that the data collection timing is suitable for creating the analysis schedule, and changes the component elution time.

[0005] US 2015 / 0102219 A1 describes that before optimizing the measurement conditions for multiple reaction monitoring, the analysis operator prepares two lists on the product ion selection condition setting screen for each precursor ion of the target compound, namely, a list showing ions to be preferentially selected as product ions for which optimization needs to be performed and a list showing ions to be excluded from the optimization. When performing measurement, a product ion scanning measurement for the precursor ions of the target compound is performed and a mass spectrum is obtained. Among the ions extracted from this mass spectrum, any ions registered in the list of excluding ions are excluded, and any ions registered in the list of preferred ions are preferentially selected as product ions. For each combination of the m / z values ​​of the precursor ion and the product ion thus determined, the optimal conditions for MRM measurement are searched.

[0006] US 9,040,903 B2 describes mass spectrometry systems and methods utilizing dynamic data acquisition / instrument control methods. These systems and methods employ artificial intelligence algorithms to greatly improve the quantitative accuracy and / or identification accuracy during data acquisition. In one embodiment, the algorithm is capable of adapting the instrument method and system during data acquisition to direct data acquisition resources to improve the quantitative accuracy or identification accuracy of target analytes (such as proteins, peptides, and peptide fragments).

[0007] WO 2018 / 116039 A1 describes a system and method for identifying an actual XIC peak of a target compound from a sample. In one system, a standard sample is used to identify the actual XIC peak. The ratio of the amount of the target compound in any two different samples is known, so this ratio is compared with the intensity of the XIC peak calculated in the two samples to identify the actual XIC peak. In another system, information about other target compounds in multiple samples is used to identify the actual XIC peak. It is known that the XIC peak of the target compound in the same sample has similar retention time distributions in these samples, so the retention time distributions of the XIC peaks are compared to identify the actual XIC peak.

[0008] JP 05835086 B2 describes the creation of a mass calibration table. After the sample is injected into the sample vaporization chamber of the GC, mass analysis is not performed and data is not collected until the period when the sample solvent is completely eluted from the column. Therefore, it is prevented that mass calibration is performed by mistakenly using the analysis results of the solvent. After that, data collection starts by scanning measurement, and when the signal intensity of the TIC becomes equal to or higher than the threshold value, it is recognized that the component for mass calibration begins to be eluted. Then, scanning measurement is performed while changing the scanning speed, mass spectra of different scanning speeds for the component for mass calibration are collected, and a mass calibration table is created based on the mass spectrum.

[0009] V. Lange et al., "Selected reaction monitoring for quantitativeproteomics: a tutorial", MOLEC-ULAR SYSTEMS BIOLOGY, Vol. 4, October 14, 2008, XP055033380, ISSN: 1744-4292, DOI: 10.1038 / msb.2008.61, describes selected reaction monitoring (SRM) for the reliable quantification of low-abundance analytes in complex mixtures. In an SRM experiment, a predefined precursor ion and one of its fragments are selected by two mass filters in a triple quadrupole instrument and monitored over time for precise quantification. A series of transitions combined with the retention time of the target peptide enables a definitive determination. Typically, a large number of peptides are quantified during a single LC-MS experiment. V. Lange et al. illustrate the application of SRM for quantitative proteomics, including the selection of proteotypic peptides and the optimization and validation of transitions.

[0010] Scheduled MRM works particularly well for very well-defined measurements and / or analyte mixtures with high concentrations. Despite the advantages of scheduled MRM, however, there is a growing interest in maximizing column lifetime, which can be closely associated with significant peak shifts, so that scheduled MRM with a fixed time range for peak recording can lead to unreliable and incorrect results.

[0011] Issues to be resolved

[0012] Therefore, the object of the present invention is to provide a method and a device for multiple transition monitoring which avoid the above-mentioned disadvantages of the known methods and devices. In particular, the method and the device will allow reliable and correct multiple transition monitoring even in the case of retention time shifts due to column aging, capillary replacement, inaccurate solvent composition and other factors. In addition, the method and the device will allow a reduction in the limit of quantification, in particular for key analytes. Summary of the invention

[0013] This problem is solved by a method and a device for multi-transition monitoring having the features of the independent claims. Preferred embodiments of the invention are disclosed in the dependent claims, which can be realized individually or in any arbitrary combination.

[0014] As used hereinafter, the terms "having", "including" or "comprising" or any of their arbitrary grammatical variations are used in a non-exclusive manner. Thus, these terms may refer both to situations in which, in addition to the features introduced by these terms, no further features are present in the entity described in this context, and to situations in which one or more further features are present. As an example, the expressions "A has B", "A includes B" and "A contains B" may refer both to situations in which, in addition to B, no other elements are present in A (i.e., situations in which A consists solely and exclusively of B), and to situations in which, in addition to B, one or more further elements (such as element C, element C and element D or even further elements) are present in entity A.

[0015] In addition, as used below, the terms "preferably", "more preferably", "particularly", "more particularly", "specifically", "more specifically" or similar terms are used in conjunction with optional features without limiting the possibilities of alternatives. Therefore, the features introduced by these terms are optional features and are not intended to limit the scope of the claims in any way. As will be appreciated by those skilled in the art, the present invention can be implemented by using alternative features. Similarly, the features introduced by "in one embodiment of the present invention" or similar expressions are intended to be optional features without any limitation on alternative embodiments of the present invention, without any limitation on the scope of the present invention, and without any limitation on the possibility of combining the features introduced in this manner with other optional or non-optional features of the present invention.

[0016] In a first aspect of the present invention, a method for multi-transition monitoring using a liquid chromatography-mass spectrometry apparatus is disclosed.

[0017] As used herein, the term "multiple transition monitoring", also denoted as multiple reaction monitoring (MRM), is a broad term and is given its ordinary and customary meaning to one of ordinary skill in the art, and is not limited to a special or customary meaning. The term may specifically refer to, but is not limited to, methods used in mass spectrometry, particularly in tandem mass spectrometry, in which multiple product ions from one or more precursor ions are monitored. As used herein, the term "monitoring" is a broad term and is given its ordinary and customary meaning to one of ordinary skill in the art, and is not limited to a special or customary meaning. The term may specifically refer to, but is not limited to, the determination and / or detection of multiple product ions.

[0018] As used herein, the term "liquid chromatography mass spectrometry device" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art and is not limited to a special or customized meaning. The term may specifically refer to, but is not limited to, a combination of liquid chromatography and mass spectrometry. The liquid chromatography mass spectrometry device may be or may include at least one high performance liquid chromatography (HPLC) device or at least one micro liquid chromatography (µLC) device. The liquid chromatography mass spectrometry device may include a liquid chromatography (LC) device and a mass spectrometry (MS) device, wherein the LC device and the MS are coupled via at least one interface.

[0019] As used herein, the term "liquid chromatography (LC) device" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, an analytical module configured to separate one or more target analytes of a sample from other components of the sample for use in detecting the one or more analytes using a mass spectrometry device. The LC device may include at least one LC column. For example, the LC device may be a single-column LC device or a multi-column LC device having a plurality of LC columns. The LC column may have a stationary phase through which a mobile phase is pumped to separate and / or elute and / or transfer the target analyte. The LC column may be replaceable, such as after a predefined or predetermined time and / or number of runs and / or other suitable counters. For example, the LC column may be replaceable if one or more thresholds of one or more of the following are reached: solvent volume, number of valve switching events, number of runs, number of injections, number of samples, number of samples of a certain type, LC pressure / curve. As used herein, the term "mass spectrometer" is a broad term and is given its ordinary and customary meaning to those of ordinary skill in the art and is not limited to a special or customized meaning. The term specifically may refer to, but is not limited to, a mass analyzer configured to detect at least one analyte based on a mass-to-charge ratio. A mass spectrometer may be or may include at least one quadrupole mass spectrometer. An interface coupling an LC device and a MS may include at least one ionization source configured to generate molecular ions and to transfer the molecular ions into a gas phase.

[0020] As used herein, the term "sample" is a broad term and is given its common and customary meaning to those of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, any sample, such as a biological sample (also referred to as a test sample), a quality control sample, an internal standard sample. The sample may include one or more target analytes. For example, the test sample may be selected from a group consisting of the following items: physiological fluids, including blood, serum, plasma, saliva, lens fluid, cerebrospinal fluid, sweat, urine, emulsion, ascites, mucus, synovial fluid, peritoneal fluid, amniotic fluid, tissue, cells, etc. The sample may be used directly when obtained from the corresponding source, or may be pretreated and / or sample preparation workflow. The sample may be pretreated by adding an internal standard and / or by diluting with another solution and / or by mixing with a reagent, etc. For example, in general, the target analyte may be vitamin D, drugs of abuse, therapeutic drugs, hormones, and metabolites. The quality control sample may be a sample of one or more quality control substances that simulates the test sample and contains known values. The quality control material can be the same as the target analyte, or can be an analyte generated by reaction or derivatization of the analyte identical to the target analyte, and / or can be an analyte whose concentration is known, and / or can be a simulated target analyte or a material that can otherwise be associated with a certain target analyte. An internal standard sample can be a sample comprising at least one internal standard substance with a known concentration. For more details about the sample, reference is made to, for example, EP 3 425 369 A1, the entire disclosure of which is included herein by reference. Other target analytes are also possible.

[0021] The method comprises the following steps, which can be performed in a given order as an example. However, it should be noted that different orders are also possible. In addition, one or more method steps can also be performed once or repeatedly. In addition, two or more method steps can be performed simultaneously or in a timely overlapping manner. The method may include other method steps not listed.

[0022] The method comprises the following steps:

[0023] a) determining at least one data set from at least one database, the data set comprising at least one reference measurement of at least one transition of at least one analyte performed with the liquid chromatography-mass spectrometry device;

[0024] b) determining at least one reference peak information of the transition of the analyte using an initial setting of a measurement window, wherein the measurement window is defined by a time range of retention time;

[0025] c) determining an actual setting of the measurement window taking into account the reference peak information, wherein the determination comprises adjusting the time range;

[0026] d) measuring the transition of the analyte with the liquid chromatography mass spectrometry device, and determining measured peak information of the transition of the analyte using the actual setting of the measurement window.

[0027] As used herein, the term "data set" is a broad term and is given its ordinary and customary meaning to those of ordinary skill in the art and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, information stored and / or stored about at least one prior MRM measurement (e.g., from a prior run). Information about a prior MRM measurement may include at least one chromatogram and / or at least one information evaluated from the chromatogram, such as peak maximum, retention time, peak start time, peak end time, peak width, in particular full width at half maximum, peak shape, tailing factor, and / or any type of peak fitting and filtering. The tailing factor T can be expressed by T=W 0.05 / (2d) Determine, where W 0.05 is the peak width at 0.05 of the peak height, and d is the distance between the vertical line through the peak maximum and the peak front edge at 0.05 of the peak height. As used herein, the term "database" is a broad term and is given its ordinary and customary meaning to those of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, a data set including the at least one data set. The database may include at least one table and / or at least one lookup table in which the at least one data set is stored. The database may include at least one storage unit configured to store a data set. As used herein, the term "determining at least one data set from at least one database" is a broad term and is given its ordinary and customary meaning to those of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, accessing a database and retrieving a data set from a database.

[0028] As used herein, the term "reference measurement" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, at least one prior MRM measurement, such as an MRM measurement of a prior run and / or at least one prediction about an MRM measurement. The reference measurement may include or may be at least one known MRM transition. The reference measurement may be: at least one measurement of at least one quality control sample collected during a prior quality control run, and / or at least one measurement of at least one internal standard sample collected during a prior internal standard sample run, and / or at least one measurement of a test sample collected during a prior run. The reference measurement may be at least one measurement collected and / or determined and / or measured in the same manner as the measurement of the actual test sample and under the same or at least similar and / or comparable conditions as the measurement of the actual test sample. The reference measurement and the measurement of the transition of the analyte may be performed under substantially the same conditions. For example, the reference measurement and the measurement of the transition of the analyte can be performed under constant chromatographic conditions (in particular using the same LC column and eluent). The method can be applicable to known compounds and well-known conditions. However, the method can also include predicting at least one reference measurement, in particular in the case of gradient changes. The prediction can include taking into account aging of the LC column, capillary changes, solvent composition inaccuracies and other factors.

[0029] As used herein, the term "reference peak information" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, at least one information of a peak (i.e., a local maximum) of a chromatogram corresponding to and / or associated with a target analyte of a reference measurement, which is suitable for limiting the relevant time range of the measurement for the target analyte. The reference peak information may include one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, in particular half-height full width, peak shape, tailing factor, and / or any type of peak fitting and filtering. The determination of the reference peak information may include evaluating the reference measurement. The evaluation may include performing at least one data analysis, the at least one data analysis including performing at least one peak search algorithm and / or performing at least one peak fitting algorithm. The evaluation may include one or more of the following: checking raw data, preprocessing, smoothing, background reduction or removal, peak detection, peak integration.

[0030] As used herein, the term "measurement window" is a broad term and is given its common and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, the time range in which the measurement of the target analyte is performed. The measurement window is defined by the time range of the retention time. It is well known that the measurement of the target analyte is limited to a certain predefined time range. As used herein, the term "setting" of the measurement window is a broad term and is given its common and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, the value of one or two limits of the measurement window. Specifically, the setting of the measurement window may include one or both of the following: the value of the lower limit of the measurement window, i.e., the retention time at the start of the measurement; and the value of the upper limit of the measurement window, i.e., the retention time at the stop of the measurement.

[0031] The term "initial setting" is a broad term and is given its common and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, the setting of a measurement window for determining reference peak information. The initial setting may be predetermined and / or predefined. For example, in the case of the first measurement after changing the column of an LC device, the initial setting may be a default setting that may be stored in a database. The method may include at least one initial calibration step, wherein the reference measurement and / or the initial setting may be determined in the initial calibration step. The initial calibration step may be performed during and / or after at least one quality control run of the liquid chromatography mass spectrometry device, and / or during and / or after at least one internal standard sample run. The start of the initial calibration step may be triggered by a change in the column of the liquid chromatography mass spectrometry device and / or after a predefined or predetermined time and / or after a predefined or predetermined number of runs or other suitable counters. As used herein, the term "trigger" may refer to an automatic process that is automatically initiated and executed, or a warning that is generated for a user and prompts the user to manually set the setting to the initial setting. In case of repeated execution of the method, the initial settings may be settings of the measurement window determined from measurements of at least one previous run or multiple previous runs, for example an average value of the limits of the time range.

[0032] The term "actual setting" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art, and is not limited to a special or customized meaning. The term may specifically refer to, but is not limited to, the setting of the measurement window determined by considering the reference peak information determined in step b). In addition, in order to consider only the reference peak information of the previous measurement, the actual setting may be determined by considering the previous measurement or multiple previous measurements. The determination of the actual setting may include evaluating the reference peak information and thereby determining the lower limit and / or upper limit of the measurement window. Specifically, at least one automatic analysis of the retention time may be performed. In addition, the measurement window may be automatically reallocated. The actual setting may be determined and / or calculated based on the expected retention time and tailing factor. The actual setting may be determined by comparing the peak value (particularly the signal intensity) and the background. For example, the signal intensity and the background may be compared by defining and / or using at least one threshold at the beginning of the peak and / or at least one threshold at the end of the peak. The actual setting may be determined by predicting based on multiple data sets.

[0033] The initial setting may include a retention time with a wider time range than the actual setting, and / or the actual setting may be shifted in the retention time compared to the initial setting. In the latter case, the width of the measurement window may be maintained. Specifically, the initial setting of the measurement window may be selected to be so wide that it is ensured that the peak corresponding to the target analyte is within the time range. Subsequently, the initial setting of the measurement window may be optimized in view of the measurement results, which allows the width of the measurement window to be reduced and / or the measurement window to be positioned, particularly in view of changes in the LC column (e.g., due to aging) or other changes. The reference measurement may include or may be a known MRM transition, wherein the method includes optimizing their measurement. The term "determining the actual setting" is a broad term and is given its ordinary and customary meaning to a person of ordinary skill in the art, and is not limited to a special or custom meaning. The term may specifically refer to, but is not limited to, adapting and / or changing the initial setting of the measurement window according to at least one subsequent measurement. The timing of the measurement may be fixed. However, due to changes in the peak position caused by the ongoing analysis, particularly in the case of repeated execution of method steps a) to d), the retention time may be shifted and may be adapted based on a pre-measurement. The method may include adjusting the timing of the MRM schedule based on the time varying parameter. Thus, the optimized retention time may be used to optimize the scheduled MRM measurements, thereby making time for more MRM transitions.

[0034] The method comprises measuring in step d) a transition of an analyte in the sample using a liquid chromatography mass spectrometry device. The measurement may be triggered by a user, for example, by applying at least one input to at least one human-machine interface of the liquid chromatography mass spectrometry device.

[0035] The actual setting of the measurement window is used to determine the measured peak information of the transition of the analyte. As used herein, the term "measured peak information" is a broad term and is given its common and customary meaning to a person of ordinary skill in the art, and is not limited to a special or customized meaning. The term specifically may refer to, but is not limited to, at least one information of a peak of a chromatogram corresponding to and / or related to the target analyte measured using the actual setting of the measurement window in step d). The measured peak information includes one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, in particular half-height full width, peak shape, tailing factor. The determination of the measured peak information may include evaluating the actual measurement. The evaluation may include performing at least one data analysis, and the at least one data analysis includes performing at least one peak search algorithm and / or performing at least one peak fitting algorithm.

[0036] The method may further comprise updating the data set by adding the measurement of the transition of the analyte to the data set. In particular, in case of repeated execution of method steps a) to d), the data set may be updated after execution of step d) so that the subsequent step a) is executed using the updated initial setting. Thus, the updating may be performed permanently.

[0037] Method steps a) to d) may be performed repeatedly. In step a), the most recent measurement may be used as a reference measurement. In step b), the reference peak information of the transition of the analyte may be determined using the most recently determined actual setting as the initial setting. After multiple repetitions of method steps a) to d), in step a), the average of multiple previous measurements may be used as a reference measurement. The average may be determined using at least five previous measurements. Additionally or alternatively, particularly in the case of large variations between runs, a moving average of multiple previous measurements may be used as a reference measurement. Additionally or alternatively, the maximum amount of previous measurements used to calculate the average may be limited. Using more than one measurement ensures that the reference data is corrected for outliers or sample effects.

[0038] The width of the measurement window may be narrowed with the number of repetitions. Thus, the measurement window becomes even better and / or more beneficial to clear more MS detection windows. The detection window may be the time range in which the mass spectrometer must perform a measurement of a sample. Specifically, the detection window may be the maximum time possible between two consecutive sample inputs in the LC column.

[0039] The method may further include at least one In situ Adjust the steps. In situ The adjustment step may be performed during step d). In situIn the adjustment step, the intensity of this transition of the analyte can be monitored during the measurement and compared with at least one predetermined or predefined threshold level. The liquid chromatography mass spectrometry device may include at least one other database, and the at least one other database is configured to store at least one definition of threshold level and / or threshold value. For example, the at least one threshold level and / or the at least one threshold value can be defined by the percentage change of signal intensity relative to background. Additionally or alternatively, at least one absolute value can be used as a threshold value for determining the number of excesses or undershoots. The other database may be configured to receive input information from a database, such as the value of the threshold level. Therefore, the threshold level may be data-driven. If the intensity drops to a level lower than the predefined threshold level, the collection of the transition can be stopped. The predetermined or predefined threshold level can be defined by multiplying the signal-to-noise ratio by a factor X, and the signal-to-noise ratio is also known from the beginning. In situ The adjustment step can be implemented as a feedback loop with automatic real-time adjustment of the measurement parameters. For example, a measurement (i.e., a specific MRM) can be started at a time known based on a previous measurement. The running time of the measurement can be determined based on the actual measured intensity of this MRM. When the intensity drops below a predetermined or predefined threshold level, the acquisition of the MRM can be stopped, and residence time can be allowed for other MRMs running simultaneously. This method may be reliable for clear peaks. However, problems may arise when the signal-to-noise ratio is low, for example, small peaks that vary greatly in separate signals. It may be advantageous for these situations that a predetermined or predefined threshold level can be given by the end of a fitting curve (e.g., a Gaussian curve). The fitting curve allows the definition of peak regions, such as the starting point of a peak and the end point of a peak. In particular, a certain peak height at the end point of a peak (represented as the end of the fitting curve) can be used as a threshold level. In principle, it is problematic to use a fitting curve during measurement because there is no complete signal at this stage. However, it is possible to use a fitted curve such as a Gaussian curve, because the fitting parameters of the Gaussian curve or other fitted curve can be determined from the one or more previous measurements. This allows all parameters of the fitted curve to be determined with only a single iteration. Additionally or alternatively, the fitted curve can be determined from the measurement of an internal standard. This allows the fitting process to be accelerated. Additionally or alternatively, a combination of fitting results for different peaks of the same analyte can be used to enhance the robustness of the fitting result. The fitted curve, in particular the Gaussian curve, can be independent of the background or less dependent on the background, and is therefore advantageous even under high noise.

[0040] The method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) can be performed by at least one computer. In particular, the method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) can be performed fully automatically. In particular, the method can be fully or partially computer-implemented, in particular implemented on a computer (such as a processor) of a device for multi-transition monitoring.

[0041] The method may include compensating for column aging and / or other effects such as capillary replacement, solvent composition inaccuracies, etc.

[0042] In a further aspect, a computer program is disclosed, comprising computer executable instructions for performing a method according to any one of the embodiments described herein (particularly method steps a) to c) and determination of measured peak information in step d) when the program is executed on a computer or a computer network (particularly a processor of an apparatus for multi-transition monitoring).

[0043] Thus, in general, a computer program is disclosed and proposed herein, which includes computer executable instructions for executing the method according to the present invention in one or more embodiments attached herein when the program is executed on a computer or a computer network. In particular, the computer program can be stored on a computer-readable data carrier. Thus, in particular, one, more than one or even all method steps as indicated above can be performed by using a computer or a computer network, preferably by using a computer program. In particular, the computer can be fully or partially integrated into the device for multi-transition monitoring, and the computer program can be specifically embodied as software. However, alternatively, at least a part of the computer can also be located outside the device for multi-transition monitoring.

[0044] This document further discloses and proposes a computer program product with program code means, so that when the program is executed on a computer or a computer network, the method according to the present invention, such as one or more method steps mentioned above, is performed in one or more embodiments attached hereto. Specifically, the program code means can be stored on a computer readable data carrier.

[0045] The present invention further discloses and proposes a data carrier having a data structure stored thereon, which, after being loaded into a computer or a computer network, such as into a working memory or a main memory of the computer or the computer network, can execute a method according to one or more embodiments disclosed herein, in particular one or more method steps mentioned above.

[0046] This article further discloses and proposes a computer program product having program code means stored on a machine-readable carrier, so that when the program is executed on a computer or a computer network, a method according to one or more embodiments disclosed herein, specifically one or more method steps mentioned above, is performed. As used herein, a computer program product refers to a program that is a tradable product. The product can generally exist in any format (such as in paper format) or on a computer-readable data carrier. Specifically, the computer program product can be distributed on a data network.

[0047] Finally, this document discloses and proposes a modulated data signal, which contains instructions readable by a computer system or a computer network for executing a method according to one or more embodiments disclosed herein, specifically one or more method steps mentioned above.

[0048] Specifically, this article further discloses:

[0049] - a computer or a computer network comprising at least one processor, wherein the processor is adapted to perform the method according to one of the embodiments described in this description,

[0050] - a computer loadable data structure, which is suitable for performing a method according to one of the embodiments described in this description when the data structure is executed on a computer,

[0051] - a computer program, wherein the computer program is suitable for executing the method according to one of the embodiments described in this description when the program is executed on a computer,

[0052] - a computer program comprising program means for carrying out the method according to one of the embodiments described in the present description when the computer program is executed on a computer or on a computer network,

[0053] - a computer program comprising program means according to the preceding embodiments, wherein these program means are stored on a computer-readable storage medium,

[0054] - a storage medium, on which a data structure is stored and wherein the data structure is suitable, after being loaded into a main memory and / or a working memory of a computer or a computer network, for carrying out a method according to one of the embodiments described in the present description, and

[0055] - A computer program product having program code means, wherein the program code means can be stored or are stored on a storage medium for carrying out the method according to one of the embodiments described in the present description when the program code means are executed on a computer or a computer network.

[0056] In a further aspect of the present invention, a device for performing multi-transition monitoring of at least one analyte in a sample is disclosed. The device comprises

[0057] - at least one liquid chromatography mass spectrometry device configured for multi-transition monitoring;

[0058] - at least one database configured to store at least one data set comprising at least one reference measurement of at least one transition of at least one analyte;

[0059] - at least one evaluation device, wherein the evaluation device is configured to determine at least one reference peak information of the transition of the analyte using an initial setting of a measurement window, wherein the measurement window is defined by a time range of retention times, wherein the evaluation device is configured to determine an actual setting of the measurement window taking into account the reference peak information, wherein the determination comprises adjusting the time range, wherein the evaluation device is configured to determine measured peak information of the transition of the analyte in subsequent measurements of the transition of the analyte using the actual setting of the measurement window with the liquid chromatography-mass spectrometry device.

[0060] The device may be configured to perform a method according to any one of the preceding embodiments.For most of the terms and possible definitions used herein, reference may be made to the description of the above method.

[0061] As further used herein, the term "evaluation device" generally refers to any device suitable for performing the method steps as described above, preferably by using at least one data processing device, and more preferably by using at least one processor and / or at least one application specific integrated circuit. Thus, as an example, the at least one evaluation device may include at least one data processing device having software code stored thereon, the software code comprising a plurality of computer commands. The evaluation device may provide one or more hardware elements for performing one or more specified operations, and / or may provide one or more processors with software for running thereon for performing one or more method steps.

[0062] Compared with known methods and devices for multi-transition monitoring, the method and device according to the present invention can provide a large number of advantages. Therefore, in particular, dynamic MRM based on previous runs can release valuable channel time, thereby increasing the sampling rate or time for analytes. This is particularly beneficial for analytes that should be measured with high sensitivity. Influences that may be caused by changes in components during their service life (e.g., column aging) and / or other influences (such as capillary replacement, solvent composition inaccuracies and other factors) and the resulting retention time shifts or changes and effects on spray stability can all be compensated within the boundaries of the available time window. Deviations caused by column batch-to-batch changes can also be compensated. The method can be implemented as a fast and powerful in-situ algorithm without changing the hardware.

[0063] To summarize and without excluding further possible embodiments, the following embodiments may be envisaged:

[0064] Embodiment 1: A method for multi-transition monitoring using a liquid chromatography-mass spectrometry device, the method comprising the following steps:

[0065] a) determining at least one data set from at least one database, the data set comprising at least one reference measurement of at least one transition of at least one analyte performed with the liquid chromatography-mass spectrometry device;

[0066] b) determining at least one reference peak information of the transition of the analyte using an initial setting of a measurement window, wherein the measurement window is defined by a time range of retention time;

[0067] c) determining an actual setting of the measurement window taking into account the reference peak information, wherein the determination comprises adjusting the time range;

[0068] d) measuring the transition of the analyte with the liquid chromatography mass spectrometry device, and determining measured peak information of the transition of the analyte using the actual setting of the measurement window.

[0069] Embodiment 2: The method according to the preceding embodiment, wherein the method further comprises: updating the data set by adding the measurement of the transition of the analyte to the data set.

[0070] Embodiment 3: The method according to the previous embodiment, wherein method steps a) to d) are repeatedly performed, wherein in step a) the most recent measurement is used as a reference measurement, and wherein in step b) the reference peak information of the transition of the analyte is determined using the most recently determined actual setting as the initial setting.

[0071] Embodiment 4: The method according to the preceding embodiment, wherein in step a) an average value of a plurality of previous measurements is used as reference measurement.

[0072] Embodiment 5: The method according to the preceding embodiment, wherein the average value is determined by using at least five previous measurements.

[0073] Embodiment 6: The method according to any one of the two preceding embodiments, wherein the maximum number of previous measurements used to calculate the average value is limited.

[0074] Embodiment 7: The method according to embodiment 2 above, wherein in step a), a moving average of a plurality of previous measurements is used as a reference measurement.

[0075] Embodiment 8: The method according to any one of the preceding embodiments, wherein the method further comprises at least one In situ The adjustment step, wherein the In situ The adjustment step is performed during step d), wherein In situ In the adjusting step, the intensity of said transitions of said analyte is monitored during said measuring and compared to at least one predetermined or predefined threshold level, wherein if said intensity drops below said predefined threshold level, acquisition of said transitions is stopped.

[0076] Embodiment 9: The method according to the preceding embodiments, wherein the at least one threshold level and / or the at least one threshold is defined by a percentage change in signal intensity relative to background and / or at least one absolute value is used as a threshold for determining an excess or undershoot.

[0077] Embodiment 10: A method according to the preceding embodiments, wherein the predetermined or predefined threshold level is given by the end of a fitting curve, such as a Gaussian curve, wherein the parameters of the fitting curve are determined from one or more previous measurements, and / or wherein the fitting curve is determined from the measurement of an internal standard, and / or wherein a combination of fitting results of different peaks of the same analyte is used to enhance the robustness of the fitting result.

[0078] Embodiment 11: A method according to any one of the preceding embodiments, wherein the reference peak information and / or the measured peak information includes one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, especially half-height full width, peak shape, tailing factor and / or any type of peak fitting and filtering.

[0079] Embodiment 12: The method according to any of the preceding embodiments, wherein the method comprises at least one initial calibration step, wherein the reference measurement and / or the initial setting is determined in the initial calibration step.

[0080] Embodiment 13: The method according to the preceding embodiments, wherein the initial calibration step is performed during and / or after at least one quality control run of the liquid chromatography-mass spectrometry device, and / or during and / or after at least one internal standard sample run.

[0081] Embodiment 14: A method according to any one of the two preceding embodiments, wherein the start of the initial calibration step is triggered by a change of a column of the liquid chromatography-mass spectrometry device and / or after a predefined or predetermined time and / or after a predefined or predetermined number of runs or other suitable counters.

[0082] Embodiment 15: The method according to any one of the preceding embodiments, wherein the initial setting comprises a wide time range of retention time compared to the actual setting.

[0083] Embodiment 16: The method according to any one of the preceding embodiments, wherein the method comprises repeating method steps a) to d), wherein the width of the measurement window narrows with the number of repetitions.

[0084] Embodiment 17: The method according to any one of the preceding embodiments, wherein the reference measurement and the measurement of the transition of the analyte are performed under substantially the same conditions.

[0085] Embodiment 18: The method according to any one of the preceding embodiments, wherein the method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) are performed by at least one computer.

[0086] Embodiment 19: The method according to any one of the preceding embodiments, wherein the method comprises compensating for column aging and / or additional effects, such as capillary replacement, solvent composition inaccuracies, etc.

[0087] Embodiment 20: A method according to any one of the preceding embodiments, wherein the actual setting is determined and / or calculated based on the expected retention time and tailing factor, and / or wherein the actual setting is determined by comparing the peak and the background, and / or wherein the actual setting is determined by prediction based on multiple data sets.

[0088] Embodiment 21: A device for multi-transition monitoring of at least one analyte in a sample, the device comprising:

[0089] - at least one liquid chromatography mass spectrometry device configured for multi-transition monitoring;

[0090] - at least one database configured to store at least one data set comprising at least one reference measurement of at least one transition of at least one analyte;

[0091] - at least one evaluation device, wherein the evaluation device is configured to determine at least one reference peak information of the transition of the analyte using an initial setting of a measurement window, wherein the measurement window is defined by a time range of retention times, wherein the evaluation device is configured to determine an actual setting of the measurement window taking into account the reference peak information, wherein the determination comprises adjusting the time range, wherein the evaluation device is configured to determine measured peak information of the transition of the analyte in subsequent measurements of the transition of the analyte using the actual setting of the measurement window with the liquid chromatography-mass spectrometry device.

[0092] Embodiment 22: The apparatus according to the preceding embodiment, wherein the apparatus is configured to perform a method according to any one of the preceding embodiments relating to methods. BRIEF DESCRIPTION OF THE DRAWINGS

[0093] Preferably in conjunction with the dependent claims, additional optional features and embodiments will be disclosed in more detail in the subsequent description of the embodiments. Wherein, as will be appreciated by those skilled in the art, each optional feature can be implemented in a separate manner and in any arbitrarily feasible combination. The scope of the present invention is not limited by the preferred embodiments. Embodiments are schematically depicted in the accompanying drawings. Wherein, the same reference numerals in these drawings refer to the same or functionally equivalent elements.

[0094] In the attached picture:

[0095] 1A and B show a flow chart of a method according to the present invention;

[0096] 2A and B show a comparison of a conventional method ( FIG. 2A ) and a method according to the present invention ( FIG. 2B );

[0097] 3A and B show a further comparison of the measurement windows of the conventional method (FIG. 3A) and the method according to the present invention (FIG. 3B); and

[0098] FIG. 4 shows a device for multi-transition monitoring according to the present invention. DETAILED DESCRIPTION

[0099] FIG. 1A shows a flow chart of a method for performing multi-transition monitoring using a liquid chromatography-mass spectrometry device 111 according to the present invention, an embodiment of which is shown in FIG. 4 .

[0100] The method comprises determining at least one data set 112 from at least one database 114 in step a). The data set 112 comprises at least one reference measurement 116 of at least one transition of at least one analyte with a liquid chromatography mass spectrometry device 111. The data set 112 may be and / or may comprise information stored and / or stored about at least one previous MRM measurement (e.g., from a previous run). The information about the previous MRM measurement may comprise at least one chromatogram and / or at least one information evaluated from the chromatogram, such as peak maximum, retention time, peak start time, peak end time, peak width, in particular full width at half maximum, peak shape, tailing factor and / or any type of peak fitting and filtering. The database 114 may comprise at least one table and / or at least one lookup table in which the at least one data set 112 is stored. The database may comprise at least one storage unit configured to store the data set.

[0101] The reference measurement may include or may be at least one known MRM transition. The reference measurement may be: at least one measurement of at least one quality control sample collected during a prior quality control run, and / or at least one measurement of at least one internal standard sample collected during a prior internal standard sample run, and / or at least one measurement of a test sample collected during a prior run. The reference measurement may be at least one measurement collected and / or determined and / or measured in the same manner as the measurement of the actual test sample and under the same or at least similar and / or comparable conditions as the measurement of the actual test sample. The reference measurement and the measurement of the transition of the analyte may be performed under substantially the same conditions. For example, the reference measurement and the measurement of the transition of the analyte may be performed under constant chromatographic conditions (particularly using the same LC column and eluent). The method may be applicable to known compounds and well-known conditions. The method may include predicting at least one reference measurement, particularly in the case of a gradient change. The prediction may include taking into account aging of the LC column, capillary replacement, solvent composition inaccuracies and other factors.

[0102] The method comprises determining at least one reference peak information of the transition of the analyte using an initial setting of a measurement window 120 in step b) (indicated by reference numeral 118), wherein the measurement window 120 is defined by a time range of retention times. An example of a measurement window 120 is shown in FIG. 2B . The reference peak information may be at least one information of a peak (i.e., a local maximum) of a chromatogram corresponding to and / or related to the target analyte of the reference measurement, which is suitable for limiting the relevant time range for the measurement of the target analyte. The reference peak information may include one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, in particular half-height full width, peak shape, tailing factor and / or any type of peak fitting and filtering. The determination of the reference peak information may include evaluating 122 the reference measurement. The evaluation 122 may include performing at least one data analysis, the at least one data analysis including performing at least one peak finding algorithm and / or performing at least one peak fitting algorithm. The evaluation may include one or more of the following: checking raw data, preprocessing, smoothing, background reduction or removal, peak detection, peak integration.

[0103] The measurement window 120 is defined by a time range of retention times. It is well known to limit the measurement of a target analyte to a certain predefined time range. The setting of the measurement window 120 may include values ​​for one or two limits of the measurement window 120. Specifically, the setting of the measurement window 120 may include one or both of the following: a value for a lower limit of the measurement window, i.e., the retention time at which the measurement starts; and a value for an upper limit of the measurement window, i.e., the retention time at which the measurement stops.

[0104] The initial setting may be a setting of a measurement window 120 for determining reference peak information. The initial setting may be predetermined and / or predefined. For example, in the case of the first measurement after changing the column of the LC device, the initial setting may be a default setting that can be stored in a database. The method may include at least one initial calibration step (not shown here), in which the reference measurement and / or the initial setting may be determined in the initial calibration step. The initial calibration step may be performed during and / or after at least one quality control run of the liquid chromatography mass spectrometry device, and / or during and / or after at least one internal standard sample run. The start of the initial calibration step may be triggered by a change in the column of the liquid chromatography mass spectrometry device 111 and / or after a predefined or predetermined time and / or after a predefined or predetermined number of runs or other suitable counters. In the case of repeatedly performing the method, the initial setting may be a setting of a measurement window 120 determined from at least one pre-run or multiple pre-run measurements, such as an average value of the limits of a time range.

[0105] The method further comprises determining the actual setting of the measurement window 120 taking into account the reference peak information in step c), wherein the determination comprises adjusting the time range. The actual setting may be the setting of the measurement window 120 determined by taking into account the reference peak information determined in step b). Furthermore, in order to take into account only the reference peak information of a previous measurement, the actual setting may be determined taking into account a previous measurement or a plurality of previous measurements. The determination of the actual setting may comprise evaluating the reference peak information and thereby determining the lower limit and / or the upper limit of the measurement window. Specifically, at least one automatic analysis of the retention time may be performed, indicated by reference numeral 124. Furthermore, the measurement window may be automatically reallocated, indicated by reference numeral 126. The actual setting may be determined and / or calculated based on the expected retention time and the tailing factor. The actual setting may be determined by comparing the peak value (particularly the signal intensity) and the background. For example, the signal intensity and the background may be compared by defining and / or using at least one threshold at the beginning of the peak and / or at least one threshold at the end of the peak. The actual setting may be determined by making a prediction based on a plurality of data sets.

[0106] The initial setting may include a retention time for a wider time range than the actual setting. Specifically, the initial setting of the measurement window 120 may be selected to be so wide that it is ensured that the peak corresponding to the target analyte is within the time range. Subsequently, the initial setting of the measurement window 120 may be optimized in view of the measurement results, which allows the width of the measurement window to be reduced and / or the measurement window 120 to be positioned, particularly in view of changes in the LC column (e.g., due to aging) or other changes. The reference measurement may include or may be a known MRM transition, wherein the method includes optimizing their measurement. The determination of the actual setting may include adapting and / or changing the initial setting of the measurement window according to at least one subsequent measurement. The timing of the measurement may be fixed. However, due to changes in the peak position caused by the ongoing analysis, particularly in the case of repeated execution of method steps a) to d), the retention time may shift and may be adapted based on a pre-measurement. The method may include adjusting the timing of the MRM plan according to a parameter that varies over time. Therefore, the optimized retention time can be used to optimize the planned MRM measurement, thereby making time for more MRM transitions.

[0107] The method comprises measuring the transition of the analyte in the sample with the liquid chromatography mass spectrometry device in step d) (indicated by reference numeral 130 ). The measurement may be triggered 128 by a user, for example by entering at least one input into at least one human machine interface of the liquid chromatography mass spectrometry device 111 .

[0108] The method also includes determining measured peak information of the transition of the analyte using the actual setting of the measurement window 120 in step d), which is represented by reference numeral 132. The measured peak information can be at least one information of a peak of a chromatogram corresponding to and / or related to the target analyte measured in step d) using the actual setting of the measurement window 120. The measured peak information includes one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, in particular half-height full width, peak shape, tailing factor and / or any type of peak fitting and filtering. The determination of the measured peak information may include evaluating the actual measurement. The evaluation may include performing at least one data analysis, and the at least one data analysis includes performing at least one peak search algorithm and / or performing at least one peak fitting algorithm.

[0109] The method may further comprise updating 134 the data set 112 by adding the measurement of the transition of the analyte to the data set 112. In particular, in case method steps a) to d) are repeatedly performed, the data set 112 may be updated after performing step d) so that the subsequent step a) is performed using the updated initial settings. Thus, the updating 134 may be performed permanently.

[0110] Method steps a) to d) may be performed repeatedly. In step a), the most recent measurement may be used as a reference measurement. In step b), the reference peak information of the transition of the analyte may be determined using the most recently determined actual setting as the initial setting. After multiple repetitions of method steps a) to d), in step a), the average of multiple previous measurements may be used as a reference measurement. The average may be determined using at least five previous measurements. Additionally or alternatively, particularly in the case of large variations between runs, a moving average of multiple previous measurements may be used as a reference measurement. Additionally or alternatively, the maximum amount of previous measurements used to calculate the average may be limited. Using more than one measurement ensures that the reference data is corrected for outliers or sample effects.

[0111] The width of the measurement window 120 may narrow with the number of repetitions. Thus, the measurement window 120 becomes even better and / or more useful to clear more MS detection windows. A detection window may be a time range within which a mass spectrometer must perform a measurement of a sample.

[0112] FIG. 1B shows a further flow chart of the method according to the present invention, wherein, in addition to the embodiment shown in FIG. 1A , the method may further include at least one in-situ adjustment step 136 . In situ The adjustment step 136 may be performed during step d). In situIn the adjustment step 136, the intensity of the transition of the analyte can be monitored during the measurement and compared with at least one predetermined or predefined threshold level. The liquid chromatography mass spectrometry device 111 may include at least one additional database 138, which is configured to store at least one definition of the threshold level and / or threshold. For example, the at least one threshold level and / or the at least one threshold can be defined by the percentage change of the signal intensity relative to the background. Additionally or alternatively, at least one absolute value can be used as a threshold for determining the number of excesses or undershoots. The additional database 138 can be configured to receive (represented by reference numeral 142) input information from the database 114, such as the value of the threshold level. Therefore, the threshold level can be data-driven. If the intensity drops below the predefined threshold level, the acquisition of the transition can be stopped. The predetermined or predefined threshold level can be defined by multiplying the signal-to-noise ratio by a factor X, which is also known from the beginning. In situ The adjustment step can be implemented as a feedback loop 140 with automatic real-time adjustment of measurement parameters. For example, a measurement (i.e., a specific MRM) can be started at a time known based on a previous measurement. The running time of the measurement can be determined based on the actual measured intensity of this MRM. When the intensity drops below a predetermined or predefined threshold level, the acquisition of the MRM can be stopped, and residence time can be allowed for other MRMs running simultaneously. This method may be reliable for clear peaks. However, problems may arise when the signal-to-noise ratio is low, for example, small peaks that vary greatly in separate signals. For these situations, it may be advantageous that a predetermined or predefined threshold level can be given by the end of a fitting curve (e.g., a Gaussian curve). The fitting curve allows the definition of peak regions, such as the starting point of a peak and the end point of a peak. In particular, a certain peak height at the end point of a peak (represented as the end of the fitting curve) can be used as a threshold level. In principle, it is problematic to use a fitting curve during measurement because there is no complete signal at this stage. However, it is possible to use a fitted curve such as a Gaussian curve, since the parameters of the Gaussian curve or other fitted curve can be determined from the one or more previous measurements. This can allow all parameters of the fitted curve to be determined with only a single iteration. Additionally or alternatively, the fitted curve can be determined from the measurement of an internal standard. This can allow the fitting process to be accelerated. Additionally or alternatively, a combination of fitting results for different peaks of the same analyte can be used to enhance the robustness of the fitting result. The fitted curve, in particular the Gaussian curve, can be independent of the background or less dependent on the background, and is therefore advantageous even under high noise.

[0113] The method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) can be performed by at least one computer. In particular, the method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) can be performed fully automatically. In particular, the method can be fully or partially computer-implemented, in particular implemented on a computer (such as a processor) of a device for multi-transition monitoring.

[0114] 2A and 2B show a comparison of the conventional method shown in FIG. 2A and the method according to the invention shown in FIG. 2B , such as described with respect to FIG. 1A and FIG. 1B . In particular, the intensity I (in %) is plotted as a function of the retention time RT (in seconds). In FIG. 2A and FIG. 2B , the upper graph shows the chromatogram for a new LC column and / or initial conditions, while the lower graph shows the chromatogram for an aged LC column and / or other changes that may occur in the experimental performance of the system. For the conventional method shown in FIG. 2A , a large measurement window is used in order to compensate for potential changes during the service life of the LC column (indicated by arrow 144 ). For example, the width Δt of the measurement window may be Δt = 20 s, and the position of the measurement window may be between 20 s and 40 s. In FIG. 2B , the measurement window 120 can be selected to be narrower than in FIG. 2A , such as Δt = 10 s. In addition, due to the repetition of method steps a) to d) and the use of the method based on pre-measured and In situ Adjusting the permanent reallocation of the measurement window 120 (indicated by arrow 146 ), the measurement window 120 can be maintained constant or can even be narrowed.

[0115] 3A and B show a further comparison of the measurement windows of the conventional method shown in FIG. 3A and the method according to the present invention shown in FIG. 3B . Specifically, the functional relationship between the intensity I (in %) and the retention time RT (in seconds) is depicted. In addition, in FIG. 3A and FIG. 3B , seven measurement windows and their widths are shown, each window corresponding to a respective peak in the chromatogram. For the conventional method shown in FIG. 3A , a large measurement window is used in order to compensate for potential changes during the service life of the LC column. In contrast, in FIG. 3B , the measurement window 120 is narrowed compared to FIG. 3A . Due to the fact that the conventional method based on the pre-measurement and In situ Permanent reallocation of the adjusted measurement window 120 is possible using such narrow measurement windows. No safety margin is required. The measurement windows show less overlap. A higher sampling rate per peak may result in more points per peak.

[0116] FIG. 4 highly schematically shows an apparatus 110 for multi-transition monitoring according to the present invention. The apparatus 110 comprises the at least one liquid chromatography mass spectrometry apparatus 111 configured for multi-transition monitoring. The liquid chromatography mass spectrometry apparatus 111 may be or may comprise at least one high performance liquid chromatography (HPLC) apparatus or at least one micro liquid chromatography (µLC) apparatus. The liquid chromatography mass spectrometry apparatus 111 may comprise a liquid chromatography (LC) apparatus and a mass spectrometry (MS) apparatus, wherein the LC apparatus and the MS are coupled via at least one interface. The LC apparatus may comprise at least one LC column. For example, the LC apparatus may be a single column LC apparatus or a multi-column LC apparatus having a plurality of LC columns. The LC column may have a stationary phase through which a mobile phase is pumped to separate and / or elute and / or transfer target analytes. The LC column may be replaceable, for example after a predefined or predetermined time and / or number of runs and / or other suitable counters. The mass spectrometry apparatus may be or may comprise at least one quadrupole mass spectrometry apparatus. The interface coupling the LC device and the MS may include at least one ionization source configured for generating molecular ions and for transferring the molecular ions into a gas phase.

[0117] The apparatus 110 further comprises a database 114 configured to store a data set 112 comprising the at least one reference measurement 116 of at least one transition of at least one analyte. For a description of the data set 112 and the database 114, reference is made to the above description of FIGS. 1A and 1B .

[0118] The device 110 includes at least one evaluation device 148. The evaluation device 148 is configured to determine at least one reference peak information of the transition of the analyte using the initial setting of the measurement window 120. The measurement window 120 is defined by the time range of the retention time. The evaluation device 148 is configured to determine the actual setting of the measurement window 120 taking into account the reference peak information. The determination includes adjusting the time range. The evaluation device 148 is configured to determine the measured peak information of the transition of the analyte in the subsequent measurement of the transition of the analyte using the liquid chromatography mass spectrometry device 111 using the actual setting of the measurement window 120.

[0119] Reference numerals list

[0120] 110 Device for multi-transition monitoring

[0121] 111 CMS Device

[0122] 112 Datasets

[0123] 114 Database

[0124] 116 Reference Measurement

[0125] 118 Step b)

[0126] 120 Measurement window

[0127] 122 Evaluation Reference Measurements

[0128] 124 Automatic Analysis

[0129] 126 Reallocate measurement window

[0130] 128 Trigger

[0131] 130 Measuring Transition

[0132] 132 Determine the measured peak information

[0133] 134 Updates

[0134] 136 In-situ adjustment steps

[0135] 138 Additional databases

[0136] 140 Feedback Loop

[0137] 142 Receiving Input

[0138] 144 Arrow

[0139] 146 Arrow

[0140] 148 Evaluation device.

Claims

1. A method for performing multi-transition monitoring using a liquid chromatography-mass spectrometry device (111), the method comprising the following steps: a) determining at least one data set (112) from at least one database (114), said data set (112) comprising at least one reference measurement (116) of at least one transition of at least one analyte performed with said liquid chromatography mass spectrometry device (111), wherein in step a) one or both of an average or a moving average of a plurality of previous measurements is used as the reference measurement (116); b) determining at least one reference peak information of the transition of the analyte using an initial setting of a measurement window (120), wherein the measurement window (120) is defined by a time range of retention times; c) determining an actual setting of the measurement window (120) taking into account the reference peak information, wherein the determination comprises adjusting the time range, wherein the actual setting is a setting of the measurement window (120) determined by taking into account the reference peak information determined in step b); d) measuring the transition of the analyte with the liquid chromatography mass spectrometry device (111), and determining measured peak information of the transition of the analyte using the actual setting of the measurement window (120).

2. The method according to claim 1, wherein the method further comprises: The data set (112) is updated (124) by adding the measurement of the transition of the analyte to the data set (112).

3. The method according to claim 2, wherein method steps a) to d) are repeatedly performed, wherein in step a), the most recent measurement is used as a reference measurement (116), and wherein in step b), the reference peak information of the transition of the analyte is determined using the most recently determined actual setting as an initial setting.

4. A method according to any one of claims 1 or 2, wherein the method further comprises at least one in situ adjustment step (136), wherein the in situ adjustment step (136) is performed during step d), wherein in the in situ adjustment step (136), the intensity of the transition of the analyte is monitored during the measurement and compared with at least one predetermined or predefined threshold level, wherein if the intensity drops below the threshold level, the acquisition of the transition is stopped.

5. A method according to any one of claims 1 or 2, wherein one or both of the reference peak information or the measured peak information includes one or more of the following: peak maximum, retention time, peak start time, peak end time, peak width, peak shape, tailing factor, or any type of peak fitting and filtering. The method of claim 5 , wherein one or both of the reference peak information or the measured peak information comprises a full width at half maximum.

7. The method according to any one of claims 1 or 2, wherein the method comprises at least one initial calibration step, wherein one or both of the reference measurement or the initial setting are determined in the initial calibration step.

8. The method according to claim 7, wherein the initial calibration step is one or more of the following: performed during or after at least one quality control run of the liquid chromatography-mass spectrometry device (111), performed during or after at least one internal standard sample run.

9. A method according to claim 7, wherein the start of the initial calibration step is triggered by one or more of the following: a change of a column of the liquid chromatography-mass spectrometry device (111), after a predefined or predetermined time, or after a predefined or predetermined number of runs or other suitable counters.

10. The method according to any one of claims 1 or 2, wherein the initial setting comprises a wide time range for retention times compared to the actual setting.

11. The method according to any one of claims 1 or 2, wherein the method comprises repeating method steps a) to d), wherein the width of the measurement window (120) narrows with the number of repetitions.

12. The method according to any one of claims 1 or 2, wherein the reference measurement and the measurement of the transition of the analyte are performed under substantially the same conditions.

13. The method according to any one of claims 1 or 2, wherein the method steps b) to c) and the determination of the measured peak information of the transition of the analyte in step d) are performed by at least one computer.

14. A device (110) for performing multi-transition monitoring of at least one analyte in a sample, the device comprising: - at least one liquid chromatography mass spectrometry device (111) configured for multi-transition monitoring; at least one database (112) configured to store at least one data set (114) comprising at least one reference measurement (116) of at least one transition of at least one analyte, wherein an average and / or a moving average of a plurality of previous measurements is used as the reference measurement (116); - at least one evaluation device (148), wherein the evaluation device (148) is configured to determine at least one reference peak information of the transition of the analyte using an initial setting of a measurement window (120), wherein the measurement window (120) is defined by a time range of a retention time, wherein the evaluation device (148) is configured to determine an actual setting of the measurement window (120) taking into account the reference peak information, wherein the determination comprises adjusting the time range, wherein the actual setting is a setting of the measurement window (120) determined by taking into account the determined reference peak information, wherein the evaluation device (148) is configured to determine measured peak information of the transition of the analyte in a subsequent measurement of the transition of the analyte performed using the liquid chromatography-mass spectrometry device (111) using the actual setting of the measurement window (120).

15. The apparatus (110) according to claim 14, wherein the apparatus (110) is configured to perform the method according to any one of claims 1 to 13.

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