Valve leak detection

Abstract

This invention relates to a method for detecting leakage in an application valve in an analytical system, the analytical system including an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, the method comprising: (i) applying a sample to the trapping column via the application valve; (ii) applying the sample from step (i) to the detector unit; and (iii) determining at least one sample component within the dead time of the analytical system. The invention also relates to an analytical system including an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, the analytical system being configured to perform the method according to the invention, and relates to methods for quality assurance and related uses.

Description

[0001] This invention relates to a method for detecting leakage in an application valve in an analytical system, the analytical system including an application valve fluidly connected to an eluent pump, a trap column, and a detector unit, the method comprising: (i) applying a sample to the trap column via the application valve; (ii) applying the sample from step (i) to the detector unit; and (iii) determining at least one sample component within the dead time of the analytical system. The invention also relates to an analytical system including an application valve fluidly connected to an eluent pump, a trap column, and a detector unit, the analytical system being configured to perform the method according to the invention, and relates to methods for quality assurance and related uses.

[0002] In chromatographic analysis systems, valves are typically used upstream of the analytical column. Specifically, application valves (such as the rapid valves used in Rapid Liquid Chromatography (Rapid LC)) are found in injection systems that allow switching between sample loading and elution modes. Normally, flow through the column is unidirectional, but in the case of Rapid LC, for example, the sample is loaded in one direction and eluted in the opposite direction.

[0003] Valves (such as quick valves) are spare parts in LC systems that require regular replacement and are critical for proper system function, especially in fully integrated and automated clinical chromatographic and / or mass spectrometric analyzers, for example. Valves fluidly connect an elution pump or loading pump to the analytical column and further to the detector. In the trap and elution setup, the loading pump is fluidly connected to the trap column via an application valve in the loading position in pre-flush mode. In the elution position, the elution pump is fluidly connected to the trap column in backflush mode and elutes the sample into the detector unit. The detector unit can be, for example, a mass spectrometer.

[0004] Due to frequent valve switching events, valves degrade over time, leading to stator and rotor wear, particularly in the valve slots on the rotor. Typically, the application valve switches twice with each injection. Valve wear can cause leaks, resulting in, for example, reduced back pressure, sample material leakage, and / or mobile phase leakage, and may also lead to incorrect fluid connections between different instrument parts. Generally, damaged valves can cause signal intensity reduction, even missing peaks, fluid leakage, and / or increased residue. While fluid leakage may be fairly easy to detect, signal intensity reduction is not. In the case of Rapid LC, damaged valves can lead to undesirable fluid connections between the loading pump, analytical column, and detector, causing sample material to leak directly into the detector unit during the sample loading step. This results in sample material loss from the trapping column and can therefore potentially lead to a loss of sensitivity and accuracy, i.e., potentially erroneous results. If this occurs in a clinical setting, incorrect analytical results can, in turn, have a considerable negative impact on patient health.

[0005] Therefore, to avoid the aforementioned drawbacks, an improved method for detecting leakage in applied valves is needed. The technical problem upon which this invention is based can be viewed as providing means and methods to satisfy the aforementioned needs. This technical problem is solved by the claims and the embodiments characterized below.

[0006] Therefore, the present invention relates to a method for detecting leakage in an application valve in an analytical system, the analytical system including an application valve fluidly connected to an eluent pump, a collection column, and a detector unit, the method comprising...

[0007] (i) Apply the sample to the trapping column;

[0008] (ii) Applying the sample from step (i) to the detector unit via the application valve; and

[0009] (iii) Detect at least one sample component within the dead time of the analytical system.

[0010] Generally, the terms used herein are given their common and conventional meanings to those skilled in the art, and are not limited to specific or customary meanings unless otherwise stated. As used below, the terms “have,” “comprise,” or “include,” or any grammatical variations thereof, are used in a non-exclusive manner. Thus, these terms can refer either to a situation where no further features exist in the entity described in this context besides the features introduced by these terms, or to a situation where one or more further features exist. As an example, the expressions “A has B,” “A includes B,” and “A contains B” can refer either to a situation where no other elements exist in A besides B (i.e., where A is solely and uniquely composed of B), or to a situation where one or more further elements (such as element C, element D, or even further elements) exist in entity A besides B. Furthermore, as those skilled in the art will understand, in embodiments, the expressions “include one” and “include a” mean “include one or more,” that is, equivalent to “include at least one.” Therefore, unless otherwise stated, a description involving one of multiple items refers to at least one such item in one implementation and multiple such items in another implementation; thus, for example, performing an “analysis” refers to performing at least one analysis, and in one implementation refers to performing multiple analyses.

[0011] Furthermore, as used below, the terms “preferred,” “more preferably,” “most preferably,” “particularly,” “more particularly,” “specifically,” “more specifically,” or similar terms are used in combination with optional features without limiting other possibilities. Therefore, features introduced by these terms are optional features and are not intended to limit the scope of the claims in any way. As those skilled in the art will recognize, the invention can be carried out by using alternative features. Similarly, features introduced by “in an embodiment” or similar expressions are intended to be optional features without limiting other embodiments of the invention, without limiting the scope of the invention, and without limiting the possibility of combining features introduced in this manner with other optional or non-optional features of the invention.

[0012] The methods specified below are in vitro methods. In principle, the method steps may be performed in any order deemed suitable by a person skilled in the art, but in one embodiment in an indicated order; furthermore, one or more of the steps (all steps in the embodiment) may be assisted or performed by automated equipment. In addition, these methods may include steps other than those explicitly mentioned above.

[0013] As used herein, unless otherwise specified, the term “about” refers to an indication value having a technical precision generally accepted in the relevant art, in one embodiment involving an indication value ±20%, in another embodiment ±10%, and in yet another embodiment ±5%. Further, the term “substantially” means that there is no deviation affecting the indicated result or use, i.e., a potential deviation will not cause the indicated result to deviate from the specified result by more than ±20%, in another embodiment more than ±10%, and in yet another embodiment more than ±5%. Therefore, “substantially composed of…” means including the specified components but excluding other components, except for materials present as impurities, unavoidable materials present as a result of the process used to provide the components, and components added for purposes other than achieving the technical effects of the invention. For example, a composition defined using the phrase “substantially composed of…” encompasses any known acceptable additives, excipients, diluents, carriers, etc. In one embodiment, a composition substantially composed of one set of components will contain less than 5% by weight, in another embodiment less than 3% by weight, in yet another embodiment less than 1% by weight, and in yet another embodiment less than 0.1% by weight of one or more unspecified components.

[0014] The method for detecting application valve leakage described herein can be performed as a standalone method to check valve sealing, for example, as part of a good practice protocol such as GLP. However, this method can also be integrated into conventional analyte measurement methods, particularly in highly automated settings. In such cases, the method described herein can be performed on the same sample used in the actual analyte measurement; in such cases, the analyte measurement and the method for detecting application valve leakage can be assigned to each other in the sense that the latter's result has a potential impact on the former, i.e., the result of the analyte measurement can be considered unreliable or invalid if an application valve leakage is detected. Therefore, the results of the analyte measurement and the method for detecting application valve leakage can be assigned to each other, making it possible to identify that the two results were obtained using the same application valve. To this end, the method for detecting application valve leakage can be performed after every 50,000 switching events of the application valve, in one embodiment after every 5,000 times, in another embodiment after every 1,000 times, and in yet another embodiment after every 100 times. As those skilled in the art will understand, in such cases, if a valve leak is detected by the method described herein, all analyte measurements prior to the previous execution of the method for detecting the applied valve leak may have to be considered unreliable or invalid; that is, all analyte measurements prior to the previous execution of the method for detecting the applied valve leak will be assigned to the most recent detection. Therefore, it is also contemplated that the method for detecting the applied valve leak is performed with each analyte measurement, such that only one analyte measurement is assigned to a given execution of the method described herein. Furthermore, the method for detecting the applied valve leak can be performed when a valve leak is suspected; such suspicion may arise, for example, from the detection of a predetermined sample component at a non-predetermined location during a previous analytical run. For example, an internal standard with a known elution time may be detected significantly before or after said known elution time. However, suspicion of valve leakage may also arise from smaller-than-expected analyte and / or internal standard peaks. In light of the description herein, the method can be performed on virtually any analytical system including the components indicated below, including analytical systems known in the art and commercially available, such as trap-elution / MS devices, LC / MS or GC / MS devices, etc.

[0015] As used herein, the term "sample" refers to a sample known or suspected of containing at least one analyte. Thus, a sample may be, for example, a sample of a subject, an environmental sample, or a technically manufactured composition of substances, such as intermediates or final products of a manufacturing process, a calibration solution, or an external or internal standard. In embodiments, a sample is or contains a body fluid sample, a sample from a tissue or organ, or a wash / rinse sample, or a swab or smear obtained from an external or internal body surface. In embodiments, a sample contains at least one analyte, as described elsewhere herein. Blood, plasma, serum, urine, saliva, or tear samples are also included. Samples can be obtained using a brush, (cotton) swab, applicator, rinse / wash solution, puncture biopsy device, needle or lancet to puncture a cavity, or by surgical instruments. However, samples obtained by well-known techniques (including, in embodiments, scrapes, swabs, or biopsy tissue from the urogenital tract, perianal region, anal canal, oral cavity, upper respiratory and digestive tract, and epidermis) are also included as samples of the present invention. Cell-free fluids can be obtained from body fluids, tissues, or organs through lysis techniques (such as homogenization) and / or separation techniques (such as filtration or centrifugation). In embodiments, samples are obtained from body fluids known or suspected of containing an analyte. Suitable sample types are selected by those skilled in the art specifically based on the analyte of interest, the availability of sample materials, etc. In embodiments, the sample is a blood sample or a blood-derived sample, such as plasma or serum. It should be understood that samples can be processed, particularly pretreated, for carrying out the methods of the invention. In particular, cells can be removed from the sample using methods and tools known in the art. Furthermore, at least one analyte can be extracted and / or purified from the sample using methods and tools known in the art. Therefore, the term "sample" can also refer to formulations containing or suspected of containing at least one analyte, derived from samples as specified above, for example, diluted, enriched, purified, and / or extracted from samples as specified above.

[0016] As used herein, the term "analyte" refers to any molecule that may or may not be present in a sample and whose presence and / or amount will be detected. Specifically, an analyte can be a small molecule, peptide, protein, oligonucleotide, polynucleotide (such as RNA or DNA), polymer, or other macromolecule. However, typically, an analyte is a small molecule compound having a molecular weight of less than 5000 Da in one embodiment and less than 2000 Da in another embodiment. Thus, in one embodiment, the analyte has a molecular weight of 1 Da to 5000 Da, in another embodiment 10 Da to 3000 Da, and in another embodiment 25 Da to 2000 Da. In embodiments, the analyte comprises or is suspected to be contained in the sample as specified above, particularly samples from a subject, especially bodily fluid samples. Thus, an analyte can be a chemical compound known or suspected to be present in or derived from a subject. Thus, an analyte can be a substrate of an enzyme in a metabolic pathway, an intermediate of such a pathway, or a product obtained from a metabolic pathway. Thus, more generally, an analyte according to the invention can be a metabolite. Metabolic pathways are well known in the art and can vary between species. Preferably, the pathway includes at least the citric acid cycle, respiratory chain, photosynthesis, photorespiration, glycolysis, gluconeogenesis, hexose monophosphate pathway, oxidized pentose phosphate pathway, fatty acid production and β-oxidation, urea cycle, amino acid biosynthesis pathway, protein degradation pathway (such as proteasome degradation), amino acid degradation pathway, and the biosynthesis or degradation of the following substances: lipids, polyketides (including, for example, flavonoids and isoflavones), isoprene (including, for example, terpenes, sterols, carotenoids, lutein), carbohydrates, phenylpropanoids and their derivatives, alkaloids, benzene rings, indoles, indole-sulfur compounds, porphyrins, anthocyanins, hormones, vitamins, cofactors (such as prosthetic groups or electron carriers), lignin, glucosinolates, purines, pyrimidines, nucleosides, nucleotides, and related molecules (such as tRNA, microRNA (miRNA), or mRNA). Therefore, small molecule metabolites typically consist of compounds from the following classes: alcohols, alkanes, alkenes, alkynes, aromatic compounds, ketones, aldehydes, carboxylic acids, esters, amines, imines, amides, cyanides, amino acids, peptides, thiols, thioesters, phosphate esters, sulfate esters, thioethers, sulfoxides, ethers, or combinations or derivatives of the foregoing. The small molecules in the metabolites can be primary metabolites essential for normal cellular function, organ function, or animal growth, development, or health. Furthermore, small molecule metabolites further include secondary metabolites with fundamental ecological functions, such as metabolites that allow an organism to adapt to its environment. Moreover, metabolites are not limited to the aforementioned primary and secondary metabolites and further encompass artificial small molecule compounds.The artificial small molecule compound may be an exogenously provided small molecule that is administered to or taken up by an organism but is not a primary or secondary metabolite as defined above, or may be derived from such a small molecule. For example, the artificial small molecule compound may be a metabolite obtained from a drug via the subject's metabolic pathway, or it may be the administered drug itself. Furthermore, the analyte may be a compound externally added to the sample, such as an internal standard. Additionally, the sample may contain two or more analytes of interest, such as compounds administered to the subject and one or more of their metabolites. Furthermore, the analytes present in the sample and their internal standards may be analytes of interest within a single sample.

[0017] Consistent with the foregoing, the term "sample component" as used herein includes every and every molecule contained in a sample; thus, an analyte (if present) is a sample component; furthermore, compounds forming the sample matrix are sample components. In the context of the methods described herein, in principle any detectable sample component may be used; in embodiments, a sample component present in high abundance in the sample is used; in another embodiment, a sample component detectable by the same detection method as the analyte is used, i.e., particularly a sample component detectable by the detector unit specified below. As understood by those skilled in the art based on the description herein, the sample component used in the methods described herein is, in embodiments, a chemical molecule that is not present in detectable amounts in other compositions (such as eluents) entering the detector unit. Therefore, in embodiments, the sample component is a compound that is specifically detectable in the sample. In embodiments, the sample component used in the methods described herein is an analyte and / or an internal standard used in the determination of the analyte. Where the detector unit comprises an analytical column, the sample component to be used according to the methods described herein may be selected to be either not blocked by the analytical column or only negligibly blocked by it. Therefore, when the detector unit includes an analytical column, the sample component to be used according to the method described herein can be selected so as not to be blocked by the analytical column for more than the dead time of the analytical system.

[0018] As used herein, the term "detection" is used in its usual, general sense. In embodiments, the term refers to drawing attention, such as to an applied valve leak, which in embodiments draws the attention of an operator or decision algorithm. Thus, in embodiments, information about the detection of an applied valve leak leads to at least one of the following decisions: (i) any analysis assigned to the detection has a high probability of error and is flagged as unreliable, its results are not output, and / or are automatically repeated on an analysis system that does not contain the applied valve; (ii) no further analysis is performed on the analysis system unless the applied valve is repaired and / or replaced, and / or (iii) the applied valve must be repaired or replaced.

[0019] As used herein, the term "determination" refers to determining at least one typical characteristic of a sample component to be determined in a sample. Specifically, a typical characteristic is a feature characterizing at least one physical, chemical, and / or biochemical property of a sample component. Such properties include, for example, molecular weight, viscosity, density, charge, spin, optical rotation, color, fluorescence, chemiluminescence, elemental composition, chemical structure, and reactivity with other compounds. The values ​​of said properties can serve as typical characteristics and can be determined using techniques well known in the art. Furthermore, a typical characteristic can be any feature derived from the values ​​of the physical and / or chemical properties of the sample component through standard operations (e.g., mathematical calculations, such as multiplication, division, or logarithmic calculations). In embodiments, at least one typical characteristic allows for the determination and / or chemical identification of the sample component and its quantity. Therefore, in embodiments, the characteristic value also contains information relating to the abundance of the sample component from which the characteristic value is derived. For example, the characteristic value of a sample component can be a peak in a mass spectrometer. Such a peak contains characteristic information of the sample component, i.e., m / z information, and an intensity value relating to the abundance (i.e., quantity) of said sample component in the sample. Furthermore, as mentioned in the method of the present invention, the determination may, in embodiments, include a compound separation step prior to the determination step. In embodiments, the compound separation step produces time-resolved separations of the sample components. Suitable techniques for separation are described below and are known to those skilled in the art. In embodiments, the sample components contained in the sample may be determined qualitatively, quantitatively, or semi-quantitatively. For quantitative determination, the absolute or precise amount of the sample component will be determined, or the relative amount of the sample component will be determined based on values ​​determined by one or more of the typical characteristics mentioned above. In cases where the precise amount of the sample component cannot or should not be determined, the relative amount may be determined. In such cases, it may be determined whether the amount of the sample component present is increased or decreased relative to a second sample containing the sample component in a second amount and / or relative to a second sample component contained in the sample in a second amount (which may be predetermined); thus, the second sample component may be, for example, a sample component known to be present in the sample at a predetermined concentration, such as an internal standard added to the sample. Therefore, quantitative analysis of sample components also includes what is sometimes referred to as semi-quantitative analysis. In embodiments, qualitative analysis is the determination of the presence of a sample component in the sample, which in embodiments exceeds the detection threshold of the detection method. In view of the description below, those skilled in the art will understand that, in the context of the methods described herein, qualitative detection of sample components within the dead time of the analytical system is often sufficient to determine applied valve leakage, although semi-quantitative or quantitative determinations may also be used, for example, according to the methods presented in the various examples herein.

[0020] As used herein, the term "analytical system" refers to a tool system comprising at least the indicated tools operatively connected to each other to allow for the acquisition of detection results. Thus, an analytical system includes at least an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, all of which are specified below. Preferred tools for applying samples to the trapping column and detector unit, and for detecting at least one sample component, are known in principle to those skilled in the art and are discussed in more detail below. How the tools are operatively connected will depend on the type of tools included in the system. In embodiments, these tools are contained in a single device. As described herein, the analytical system is configured to perform steps (i) through (iii) indicated above, in the indicated order in embodiments. Thus, in embodiments, the analytical system includes a controller unit configured to direct the indicated steps to be performed. Furthermore, the analytical system may include further components deemed appropriate by those skilled in the art. In particular, the analytical system may be a liquid analytical system or a gas analytical system.

[0021] In one embodiment, the analytical system includes a sample processing unit comprising a container for a sample. This container may be in direct contact with the sample, or it may be a container for receiving the sample using an additional tool, such as a well plate, onto which one or more samples may be applied. Furthermore, in another embodiment, the sample processing unit includes at least one reservoir for a sample pretreatment agent, such as a diluent, detergent, precipitant, buffer, analyte binder (which may bind to a solid surface, such as magnetic beads), etc. In another embodiment, the sample processing unit includes tools for mixing and / or for adjusting the temperature of the sample or the reaction mixture containing the sample. The sample processing unit may also include tools for separating sample components, such as a centrifuge, magnet, filter, etc. In another embodiment, the analytical system includes a waste container, which may be any embodiment deemed appropriate by those skilled in the art.

[0022] In one embodiment, the analytical system includes a sample loading unit comprising a loading pump and, optionally, an injection valve for sample injection. In one embodiment, the loading pump pumps the sample from a sample container, or in another embodiment, from a sample processing unit, or optionally from the injection valve, to a trapping column. Those skilled in the art select an appropriate pump based on the requirements of the specific analytical system, particularly based on the trapping column used and the associated back pressure, as well as requirements arising from the sample type, diluent, and / or liquid phase composition. The aforementioned pumping is performed via an application valve, or, in another embodiment, via a sample loading port, a fluid connector, and an application port at the loading position of the application valve. Therefore, in one embodiment, the sample loading unit generates a loading pressure of 1 MPa to 120 MPa, or, in another embodiment, 2 MPa to 25 MPa, or, in yet another embodiment, 5 MPa to 15 MPa, or, in yet another embodiment, approximately 10 MPa, in the sample loading port, the fluid connector, the application port of the application valve, the trapping column, and the interconnecting tubing.

[0023] In the implementation scheme, the analysis system includes an input unit. As used herein, the term "input unit" refers to any unit configured to transfer information from another entity to the system (particularly its data processing unit or data storage medium), wherein the other entity may be another data processing device or a user. Thus, an input unit may include a user interface; however, an input unit may also be a storage medium comprising a dataset from which data, such as appropriate parameter values, can be retrieved.

[0024] In an implementation, the analysis system includes a data processing unit. The term "data processing unit" generally refers to any unit suitable for performing or causing the analysis system to perform the steps of the methods described herein, implemented in an implementation using at least one processor. Thus, as an example, the at least one data processing unit may include software code stored thereon, which includes a plurality of computer instructions. In an implementation, when performed on a microprocessor, the computer instructions cause the analysis system to perform the methods specified herein. The data processing unit may provide one or more hardware elements for performing one or more indicated operations, and / or may provide software running thereon to one or more processors for performing one or more method steps.

[0025] In one embodiment, the analysis system further includes a data output unit operatively connected to the detector unit, the evaluation unit, and / or the data processing unit described below. In another embodiment, the data output unit is adapted to output data obtained by the detector unit and optionally from other units of the analysis system. Suitable data output units are known to those skilled in the art and include simple output units, such as indicator signals or displays indicating that at least one sample component (in this embodiment, above a detection threshold) has been detected within the dead time of the analysis system. However, the output unit can also be an interface to an evaluation device, wherein the interface can be any kind of data transmission tool, including, for example, wired connections (such as USB or LAN), wireless connections (such as wireless LAN, Bluetooth, etc.), or indirect connections (such as data transmission via instant messaging, email, etc.).

[0026] In embodiments, the analysis system further includes an evaluation device. As those skilled in the art will understand, the evaluation device may be housed within the same enclosure as other components of the analysis system (e.g., the detector unit), or it may be a separate device. In embodiments, the evaluation device includes a microprocessor programmed to receive output data from the output unit and perform logical operations to provide an evaluation of the output data. The evaluation of the output data may include, for example: correcting the data for values ​​measured in at least one control; statistical calculations (e.g., calculating the average of two or more parallel detection reactions); correcting the data according to a dilution factor; comparing the output data with reference values; peak finding and peak analysis; compiling the data into a list, etc. However, the evaluation data may also include data obtained by applying a decision algorithm (e.g., as specified elsewhere herein). In embodiments, the evaluation device further includes a data storage unit. In another embodiment, the data storage unit contains reference values ​​(e.g., in a reference value database). Furthermore, in embodiments, the data storage unit is adapted to store output data received from the system of the present invention.

[0027] In embodiments, particularly where tools for automatically detecting at least one sample component are applied, the data obtained through the automated determination can be processed, for example, by a computer program to arrive at results supporting a conclusion (e.g., detection of an application valve leak). Typical detection tools are disclosed elsewhere herein in connection with embodiments of the method of the invention. These tools can be operatively linked such that the detection results and their value for the function of the application valve are provided to a user or an automated diagnostic unit. The results can be given as output of raw data for parametric diagnosis, preferably as an absolute or relative amount of the at least one sample component detected. However, the output can also be information about the status of the application valve, or simply confirmation of whether a particular analysis assigned to the method for detecting application valve leaks has been interfered with by the application valve leak.

[0028] The analytical system includes a trapping column. The term "trapping column" is known in principle to those skilled in the art. In embodiments, the term refers to each chromatographic column comprising at least one stationary phase and at least one mobile phase, configured to substantially immobilize at least one analyte from a sample, in embodiments by binding to the stationary phase, for example, through hydrophobic interactions, van der Waals forces, and / or ionic interactions. Those skilled in the art select appropriate stationary and mobile phases for the trapping column based on well-known parameters of the analyte of interest, such as hydrophobicity, charge, dipole moment, eluent composition, and other known parameters. Appropriate trapping columns are known in the art. As understood by those skilled in the art from the foregoing, a trapping column can be used to trap only, i.e., bind the analyte, optionally binding at least a portion of the unbound other sample components. However, a trapping column can also additionally provide chromatography of the analyte, for example, in the case of applying a mobile phase gradient. The trapping column can be used in a trapping and elution mode, i.e., the sample can be applied to the column in a first flow direction, while the analyte elutes from the trapping column in a second, opposite flow direction. Given the description herein, it is irrelevant whether the sample components used to determine in step (iii) are captured (i.e. bound) by the capture column, since in the event of a leak in the application valve, they enter the detector unit via the direct connection (leak) between the application port and / or the sample loading port in the loading position of the application valve and the detector unit port; it will be understood that this also applies if the sample components used in the method are analytes and / or internal standards.

[0029] The analytical system also includes an eluent pump. The term "eluent pump" is known to those skilled in the art and, in embodiments, refers to a pump that pumps eluent or a mixture of eluents from at least one reservoir to a detector unit. In one embodiment, the eluent pump is connected to the eluent pump port of an application valve. Also in one embodiment, in the loaded position of the application valve, the eluent pump port is connected via a fluid connector to the detector unit port of the application valve. Also in one embodiment, in the unloaded position of the application valve, the eluent pump port is connected via a fluid connector to the eluent port of the application valve; thus, in the unloaded position, eluent can flow from the eluent pump to the eluent pump port, via the fluid connector to the eluent port, to the trapping column, then to the application port, then via the fluid connector to the detector unit port, and then to the detector unit. Those skilled in the art select an appropriate pump according to the requirements of a particular analytical system, particularly based on the trapping column used and the associated back pressure value, as well as requirements arising from the sample type, diluent and / or liquid phase composition, the nature of the detector unit, etc.

[0030] The term "detector unit" is known to those skilled in the art. In embodiments, this term includes each tool capable of determining a compound to be determined, wherein the compound to be determined may be a sample component, an analyte, or both as specified above. Thus, a detector unit includes at least one detector for detecting the compound to be determined, the detector being adapted to perform a detection measurement of the compound to be determined. Those skilled in the art select the detector based on the physical, chemical, and / or immunological properties of the compound to be determined. The detector unit is configured to detect at least the sample component in step (iii); if the detector of the detector unit cannot detect the analyte, the detector unit may include a second detector for detecting the analyte. However, in embodiments, the sample component and / or detector are selected such that the detector detects both the sample component and the analyte. Thus, in another embodiment, the sample component is selected as the analyte and / or its internal standard. Sample components and / or analytes that absorb light can be detected by optical tools, such as by UV / VIS spectroscopy; sample components and / or analytes having at least one immunologically detectable epitope can be detected by immunological tools, etc. In embodiments, the detector unit includes at least one mass spectrometry device. The detector unit may further include tools for enriching the analyte relative to other sample components, such as a chromatographic unit comprising at least one chromatographic column, as specified below in the context of chromatography / mass spectrometry in the embodiments. However, in the embodiments, the analyte is transferred directly from the trapping column to the detector, particularly a geospectral device, via an application valve. As those skilled in the art will understand from the description herein, any compound separation step, for example on the chromatographic unit, may be associated with the detection of at least one sample component in step (iii), only in that it may increase the time required for the sample component to become detectable. However, since sample leakage is typically a continuous process, there may not be a definite start time for applying the sample component to the chromatographic unit, especially in the case of long (e.g., >10 s) sample applications, and therefore the retention time of the sample component in the chromatographic unit may not be determined. The detector unit may be connected to the analytical system for the purpose of detecting application valve leakage; that is, the detector unit may be different from the detector unit used to determine the analyte. However, in the embodiments, the detector unit used to detect at least one sample component is the same as the detector unit used to detect the analyte and / or internal standard, especially where the methods described herein are integrated into the analytical method, as described above. Thus, the detector unit may be specifically adapted to determine both the analyte and the sample component. Therefore, in the implementation scheme, the detector unit is selected from the list of the following: mass spectrometry (MS) device, diode array detector (DAD), UV / VIS detector, conductivity monitor, fluorescence detector, refractive detector, radioactive flow detector, chiral detector, evaporative light scattering detector (ELSD), and electro-fogging detector (CAD).In another embodiment, the detector unit includes a mass spectrometry unit, which is a mass spectrometry unit in another embodiment.

[0031] The term "mass spectrometry," often abbreviated as "MS," is known to those skilled in the art. In mass spectrometry, the analyte in a sample is ionized to produce charged molecules or molecular fragments. The mass-to-charge ratio of the ionized analyte or its fragments is then measured. Ionization of molecules can be performed by any method deemed appropriate, particularly by electron impact ionization, fast atom impact, electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), and matrix-assisted laser desorption / sorption ionization (MALDI). Mass spectrometry as described herein encompasses all techniques that allow the determination of the molecular weight (i.e., mass) or mass variable corresponding to the analyte or its fragments. In the implementation scheme, mass spectrometry is used in combination with chromatographic separation steps, particularly as gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), direct infusion mass spectrometry or Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS), capillary electrophoresis mass spectrometry (CE-MS), high performance liquid chromatography-mass spectrometry (HPLC-MS), quadrupole mass spectrometry, any sequentially coupled mass spectrometry, such as MS-MS or MS-MS-MS, inductively coupled plasma mass spectrometry (ICP-MS), pyrolysis mass spectrometry (Py-MS), ion mobility mass spectrometry, or time-of-flight mass spectrometry (TOF). How to apply these techniques is well known to those skilled in the art. Furthermore, suitable apparatus is commercially available.

[0032] The analytical system also includes an application valve. The term "valve" is as understood by those skilled in the art; in embodiments, the term refers to a device for controlling the flow rate and / or direction of a liquid or gas, in a closed system. In embodiments, the valve is an automatic valve, i.e., adapted to be switched without operator interaction; thus, the automatic valve can be switched based on the state of the analytical system, such as, for example, sample application initiation and / or sample dispensing initiation.

[0033] As is known to those skilled in the art, analytical systems typically have at least one valve, and in embodiments, multiple valves. As referred to herein, an "application valve" is a valve of the analytical system adapted to apply a sample to a trap column in a loading position and to detach the sample to a detector unit in a detach position. Therefore, in embodiments, the application valve has at least two switching positions: a sample loading position and a sample detach position. As understood by those skilled in the art, the terms "sample loading" and "sample detach" are used herein with reference to the trap column; that is, sample loading involves loading a sample onto the trap column, and sample detachment involves eluting the sample from the trap column, in embodiments, into the detector unit. The trap column typically applies back pressure to the sample; therefore, the analytical system may additionally include a sample loading unit comprising a sample loading pump, as specified above. In embodiments, the sample is loaded at a pressure of at least 1 MPa in the loading position via the application valve, at least 2 MPa in one embodiment, and at least 3 MPa in another embodiment. In one embodiment, the sample is loaded at a pressure of up to 120 MPa, up to 50 MPa, and up to 20 MPa in another embodiment, through the application valve at the loading position. In another embodiment, the sample is loaded at a pressure of 1 MPa to 120 MPa, 3 MPa to 25 MPa, and 5 MPa to 15 MPa in another embodiment, through the application valve at the loading position. In another embodiment, the application valve is a rapid application valve, i.e., the application valve of a rapid chromatography system, specifically the application valve of a rapid liquid chromatography system.

[0034] In the analytical system, the application valve is fluidly connected to the eluent pump, the trap column, and the detector unit. Therefore, the application valve comprises multiple ports, at least three in one embodiment, at least four in another embodiment, at least five in yet another embodiment, and at least six in yet another embodiment. The term "port," as commonly used, refers to any tool suitable for providing a fluid connection to the indicated unit. In one embodiment, the application valve comprises three to ten ports, four to eight in another embodiment, five to seven in yet another embodiment, and six in yet another embodiment. As mentioned herein, ports can be designated by the name indicating the unit or device to which they are connected; thus, the port connecting the application valve to the elution pump is the elution pump port; the port connecting the application valve to the detector unit is the detector unit port, and any port connecting the trap column to the application valve is the trap column port. Since the trap column can be connected to the application valve at both ends, there can be two trap column ports; in the case of a trap and elution system, the trap column port connected to the loading pump at the loading position can also be called the application port, and the trap column port connected to the elution pump at the sample detachment position can be called the eluent port. In one embodiment, the application port and the detector unit port are directly adjacent to each other on the valve; that is, there is no port between the application port and the detector unit port. In another embodiment, the sample loading port and the detector unit port are directly adjacent to each other on the valve.

[0035] In another embodiment, in the non-leakage application valve, neither the application port nor the sample loading port is fluidly connected to the detector unit port in the sample loading position; however, in the sample disengagement position of the application valve, the application port is fluidly connected to the detector unit port. Also in this embodiment, the sample loading port (i.e., the port through which the sample is applied to the analytical valve) and the application port are pressurized during sample loading, at least 1 MPa in one embodiment, and at least 3 MPa in another embodiment. In this embodiment, the pressure applied to the application port and the sample loading port is higher than the pressure applied to the detector unit port during sample loading. Therefore, when using a loading pump for sample loading, in this embodiment, the pressure of the sample loading pump is higher than the pressure of the elution pump during sample loading.

[0036] The application valve further includes at least one fluid connector, as the term "fluid connector" as used herein refers to any tool that provides a fluid connection between at least two ports of the application valve, in embodiments. Thus, in embodiments, the fluid connector is an internal structure of the application valve. The fluid connector may, in particular, be a recess or cavity in the application valve, having in embodiments a geometry that connects a port to a first adjacent port in a first valve position and to a second adjacent port in a second valve position, wherein the first adjacent port is not the same as the second adjacent port. In embodiments, the application valve has at least two fluid connectors, and in another embodiment, at least three fluid connectors. The fluid connectors may be arranged such that each fluid connector independently connects two ports; thus, the fluid connectors of the application valve are not fluidly interconnected with each other except for potential connections via external elements of the application valve, such as, for example, conduits connecting a port to a first fluid connector to a second port connected to a second fluid connector. That is, in embodiments, the application valve includes a plurality of fluid connectors arranged such that, in a switching position, each port is fluidly connected to one of its adjacent ports but not to its other adjacent port. Exemplary port / fluid connector combinations and geometries are explicitly referenced to the examples provided below. Without any inventive effort, those skilled in the art can obtain iterative schemes of allocation schemes that provide the same functionality as those specifically shown in each example.

[0037] In an embodiment, the application valve comprises two movable elements, specifically a rotor and a stator. Suitable embodiments are known in the art. Relative movement of the rotor and stator of the application valve can cause wear and other damage, potentially leading to new, undesirable fluid connections from the application port and / or sample loading port to the detector unit port, i.e., leakage. In an embodiment, application valve leakage is caused by failure of the elements separating the application port and detector unit port and / or the elements separating the sample loading port and detector unit port.

[0038] The term "leakage" is used herein in the sense known to those skilled in the art; therefore, in embodiments, the term leakage refers to the improper spatial and / or temporal transfer of liquids and / or gases. In embodiments, leakage is the loss of liquid or gas from the analytical system or its components (specifically, its valves). More specifically, leakage of the application valve allows a sample to transfer from the application port of the application valve or from the loading unit port to the detector unit port at the loading position, wherein said transfer, in embodiments, does not include contact between the sample and the trapping column. As those skilled in the art will understand, such leakage (which can be detected by the methods described herein) can be considered as an indication of a specific leak detected, but can also be considered as an indication that the application valve is unsuitable for further use and the risk of other leaks that may occur, such as from the eluent pump port and / or the detector unit port to other ports of the application valve and / or to the outside of the application valve, which may in turn lead to a reduction in analyte peaks. Structurally, leakage can be caused by wear of the application valve components, such as by repeated switching events, by improper changes in the size of the application valve elements (e.g., improper expansion of the application valve housing), by a breakage in the connection between the application port or loading unit port and the detector unit port within the valve, etc.

[0039] As those skilled in the art will understand, every analytical system has an internal volume between the sample (e.g., a sample bound to a trap column) and the detector, however small that volume may be. Because sample components (even those not interacting with any parts of the analytical system) cannot reach the detector before being transported through this internal volume, it is also referred to as the system's "dead volume." As those skilled in the art will also understand, for a given analytical system and a given flow rate, the dead volume of the analytical system can also be described as the "dead time," that is, the time required for the dead volume to be transported through the analytical system. Therefore, the dead time is the shortest time required for any sample component to be transported from the trap column to the detector. Thus, any sample component that arrives at the detector before the dead time has elapsed could not have been transported through the conventional sample transport process within the analytical system. As mentioned herein, the dead volume and dead time begin when the application valve switches from the loading position to the unloading position.

[0040] In the context of the analytical system described herein, the term "connection" refers to a connection that allows for the substantially lossless transport of liquids or gases, in embodiments at pressures as indicated elsewhere herein. Therefore, in such cases, the connection is typically a fluid connection or a gas connection.

[0041] The method includes step (i) applying a sample to a trapping column via an application valve. The sample, as well as the tools and methods for applying the sample to the trapping column, have been described above. As mentioned herein, during step (i), the application valve is in a loading position. Specifically, the sample can be applied to the trapping column via an application port using a tool suitable for overcoming the back pressure of the trapping column. Such a tool can be an operator-operated syringe; however, in embodiments, a loading pump is used, in conjunction with an injection valve. During sample application, at least for a portion of the time required for sample application, the pressure in the application port of the application valve and the sample loading port is higher than the pressure at the detector unit port. Therefore, sample leaking from the application port of the application valve and / or the sample loading port can be transferred to the detector unit port and ultimately enter the detector unit.

[0042] The method further includes step (ii) applying the sample from step (i) to the detector unit. During step (ii), the application valve is in the detached position; in an embodiment, step (ii) begins by switching the application valve to the detached position. As described above, each analytical system has a dead time during which sample components cannot reach the detector via the conventional connection of the analytical system. Nevertheless, applying the sample to the detector unit requires delivering eluent through the trap column and the application valve into the detector. Therefore, any sample material that has entered the detector unit port while the application valve is still in the loaded position will be delivered to the detector, in an embodiment before the dead time ends. Methods for applying the sample from the trap column to the detector unit via the application valve are known in principle in the art and have been described above. In an embodiment, step (ii) includes, with the application valve in the detached position, pumping eluent through the analytical system, via the trap column, the application valve, and into the detector unit.

[0043] The method further includes step (iii) determining at least one sample component within the dead time of the analytical system. The terms sample component, dead volume, and analytical system have been described above. The at least one sample component can be determined using any tool deemed appropriate by a person skilled in the art. In embodiments, the method described herein is integrated into an analytical method, and the at least one sample component is determined using the same detector as the analyte, in the same method. As discussed above, the method may be or may be included in an analytical method for determining the analyte; in such cases, the sample compound detected in step (iii) is, in the embodiments, the analyte and / or internal standard. As will be understood, step (iii) does not necessarily have to include the identification of the at least one sample component; in the context of the method described herein, qualitative determination of the sample component may be sufficient. However, in embodiments, a method for quantitative determination of the sample component is applied.

[0044] Determining sample components within the dead time of an analytical system can be performed by any method deemed appropriate by those skilled in the art. In an embodiment, an average signal is calculated, for example, the average intensity of the MS signal over a predetermined m / z range or value. This can be achieved, for example, as illustrated in the various examples herein, as the average signal intensity over a predetermined time interval. This average signal can be compared to a predetermined reference value. Such a reference value can be, for example, the average of signals measured using multiple analytical systems with known non-leaking application valves. In such cases, for example, the average ratio of the analytical systems of interest can be divided by the aforementioned average signal, wherein, in an embodiment, a ratio substantially 1 would be considered as indicating the absence of application valve leakage, while a ratio significantly higher than 1 would be considered as indicating application valve leakage. Since application valve leakage tends to increase with wear, the ratio can also be used to classify valve leakage, for example, by assigning a ratio value substantially 1 as no leakage, a ratio value between 2 and 5 as the beginning of leakage, potentially automatically suggesting the user contact a service technician, while a ratio higher than 5 indicates significant leakage, potentially automatically stopping all measurements on the affected analytical system and informing the user of significant degradation that may adversely affect measurement results.

[0045] Determining sample components within the dead time of the analytical system can also be performed by measuring the ratio of the amount of sample component eluted within the dead time to the amount eluted within the expected run time. As those skilled in the art will understand, this method can be used, for example, when the sample component to be measured is an analyte and / or its internal standard. The amounts of sample component eluted within the dead time and the amounts eluted within the expected run time can be determined by performing standard peak detection, followed by peak height determination and / or peak integration to determine the amounts. As those skilled in the art will understand, in such cases it may not be necessary to determine the amount as an absolute quantity or concentration, as relative quantities or arbitrary units can also be used. In embodiments, a value of up to 0.1 is considered acceptable, indicating no valve leakage, while values ​​exceeding 0.1 to 1 may indicate the onset of valve leakage, and values ​​exceeding 1, and in embodiments exceeding 2, indicate severe valve leakage, which is unacceptable in embodiments. As those skilled in the art will understand from the description herein, sample component peaks in the dead time of an analytical system can have unusual forms, including plateaus, because sample application may extend to cover a considerable portion of the loading phase, and therefore peak identification algorithms may need to be adapted for this purpose.

[0046] Specific examples of methods for determining sample components are provided in the examples in this article.

[0047] As those skilled in the art will understand from the description herein, determining at least one sample component within the dead time of the analytical system indicates an application valve leak. Therefore, the method may further include step (iv) detecting the presence of an application valve leak if at least one sample component is determined within the dead time of the analytical system, and detecting the absence of an application valve leak if no sample component is determined within the dead time of the analytical system.

[0048] In view of the above, the method may include further steps, such as step (oa), switching the application valve to the loading position before step (i), and / or step (ia), switching the application valve to the unloading position before step (ii). Further steps may, for example, involve washing and / or regenerating the analytical system, specifically trapping columns and optionally any analytical columns.

[0049] Advantageously, in the work that forms the basis of this invention, it was discovered that leakage of the applied valve can be detected by analyzing the sample composition within the dead time of the system.

[0050] The definitions made above apply mutatis mutandis to those below. Further definitions and explanations provided below also apply mutatis mutandis to all embodiments described in this specification.

[0051] The present invention further relates to an analytical system comprising an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, configured to perform the methods specified herein.

[0052] The analytical system, its components, and optional and auxiliary units have been described above. The analytical system may be an analytical system that includes an MS unit as a detector unit, specifically an LC / triple quadrupole mass spectrometry (LC / TQMS) system.

[0053] The terms “apparatus” and “unit” are used substantially interchangeably herein and refer to a collection of tools operatively connected to each other to provide the indicated function, as part of an analysis system in an embodiment. The apparatus may be implemented in a single physical unit or in physically separate units operatively connected to each other. Suitable components and their properties are described elsewhere herein and above in the context of the method. Thus, one or more methods of the invention can be implemented by the analysis system specified herein. Therefore, in an embodiment, the analysis system is configured to perform at least one method as specified elsewhere herein. The analysis system may include further units, as specified above in an embodiment, and / or any other units deemed appropriate by those skilled in the art.

[0054] The present invention also relates to a method for quality assurance in sample analysis, the method comprising:

[0055] (a) Determine at least one analyte in the sample.

[0056] (b) Perform the method described herein for detecting leakage in the applied valve; and

[0057] (c) If a sample component is detected in step (b), the determination in step (a) is marked as invalid.

[0058] The methods used for quality assurance (QA) may be assisted or performed by automated equipment and may be specifically performed on analytical systems described elsewhere herein. These methods may be incorporated into QA processes and / or analyte determination methods.

[0059] The term "quality assurance" is used in its meaning as known to those skilled in the art, and in implementation refers to any measures taken to ensure and / or validate a process (specifically, analyte measurements) meets predefined quality standards. Such quality standards may specifically be ensuring measurement accuracy within predefined error limits, preventing sample cross-contamination, etc. As is known to those skilled in the art, the methods for QA presented herein may be one of many QA measures that may involve the same or different quality standards.

[0060] The term "marking determination" is used broadly herein to include any action that assigns QA-related assessments to at least one measurement and / or thus modifies the process (if necessary). Marking can be specific to the results of the methods described above, or it can be general, such as indicating that at least one QA criterion has not been met. Thus, marking determination can be the result of step (a) that assigns information indicating that the result is invalid, fails QA, etc. However, marking can also be the measurement of step (a) automatically repeated on a different analytical system and / or on the same analytical system after replacing or repairing the application valve, or it can be the result of step (a) that is not output on the output unit.

[0061] The method includes step (a) determining at least one analyte in the sample. Suitable tools and measures for performing step (a) are determined by those skilled in the art without inventive effort, taking into account the analyte of interest, sample type, and other parameters.

[0062] The method includes step (b) performing a method for detecting leakage at the applied valve, as described above. Step (b) may be performed concurrently with step (a), i.e., the sample components may be determined within the dead time as analyte removal from the trapping column begins. As those skilled in the art will understand from the above description, step (b) may be performed each time step (a) is performed; however, step (b) may also be performed after multiple iterations of step (a) with the same or different analytes and / or with the same or different samples. Thus, in a method for QA, step (b) may be performed after step (a) has been performed up to 50,000 times, up to 5,000 times in one embodiment, up to 1,000 times in another embodiment, and up to 100 times in yet another embodiment.

[0063] The method includes further steps (c) and / or (d), wherein step (c) marks the determination in step (a) as invalid if a sample component is detected in step (b). As discussed above, marking determination should be understood as a broad term. Furthermore, if a sample component is detected in step (b), a further action in step (d) may be initiated alternatively or additionally, wherein step (d) causes the analytical system to prevent any further sample application to the application valve. This can be achieved by any tool deemed appropriate by those skilled in the art, for example by preventing switching to the sample loading position, by depressurizing the affected analytical system, for example by deactivating the loading pump, etc. Thus, in the embodiment, further analysis on the analytical system is prevented. Additionally, repair or replacement of the application valve may be initiated. Furthermore, if no sample component is detected in step (b), the sample may also be marked, for example by releasing the result of step (a) to, for example, an output unit, indicating that this particular QA method has been passed.

[0064] The present invention also relates to a method for monitoring and analyzing the performance of an application valve included in a system, the method comprising performing the method described herein for detecting application valve leakage after up to 500,000 application valve switching events.

[0065] Furthermore, the present invention relates to a method for at least one application valve in a management analysis system, the method comprising the step of monitoring the performance of the application valve, and the further step of replacing the application valve if the performance of the application valve is classified as unsatisfactory; wherein, specifically, the performance of the application valve is classified as unsatisfactory if leakage of the application valve is detected.

[0066] Furthermore, the present invention relates to the use of determining sample components within the dead time of an analytical system for detecting leakage in an applied valve.

[0067] The present invention further discloses and proposes a computer program comprising computer-executable instructions for performing the method according to the invention in one or more embodiments appended herein, when executed on a computer or computer network. Specifically, the computer program may be stored on a computer-readable data carrier. Thus, specifically, one, more than one, or even all of the method steps a) to d) indicated above can be performed in the embodiments by using a computer or computer network via the computer program.

[0068] The present invention further discloses and proposes a computer program product having program code tools so that, when the program is executed on a computer or computer network, the method according to the invention is performed in one or more embodiments appended herein. Specifically, the program code tools may be stored on a computer-readable data carrier.

[0069] Furthermore, the present invention discloses and proposes a data carrier having a data structure stored thereon, which, after being loaded into a computer or computer network, such as after being loaded into the working memory or main memory of the computer or computer network, can perform methods according to one or more embodiments disclosed herein.

[0070] This invention further proposes and discloses a computer program product having program code tools stored on a machine-readable carrier, so that when the program is executed on a computer or computer network, it performs methods according to one or more embodiments disclosed herein. As used herein, a computer program product refers to a program that is a tradable product. This 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.

[0071] Finally, the present invention proposes and discloses a modulated data signal comprising instructions readable by a computer system or computer network for performing a method according to one or more embodiments disclosed herein.

[0072] In embodiments, referring to the computer implementation aspects of the present invention, one or more method steps, or even all method steps, of the methods according to one or more embodiments disclosed herein can be performed using a computer or computer network. Therefore, generally, any method steps, including providing and / or manipulating data, can be performed using a computer or computer network. Generally, these method steps can include any method steps other than those typically requiring manual work (such as providing samples and / or performing certain aspects of actual measurement).

[0073] Specifically, the present invention further discloses:

[0074] - A computer or computer network including at least one processor, wherein the processor is adapted to perform a method according to one of the embodiments described in this specification.

[0075] - A computer-loadable data structure, adapted to perform one of the methods described in this specification when the data structure is executed on the computer.

[0076] - A computer program, wherein the computer program is adapted, when executed on a computer, to perform a method according to one of the embodiments described in this specification.

[0077] - A computer program, comprising program means for performing a method according to one of the embodiments described herein when executed on a computer or on a computer network.

[0078] - A computer program, comprising program means according to the foregoing embodiments, wherein the program means is stored on a computer-readable storage medium.

[0079] - A storage medium, wherein a data structure is stored on the storage medium and wherein the data structure is adapted to be processed according to one of the embodiments described herein after being loaded into the main memory and / or working memory of a computer or computer network.

[0080] - A computer program product having program code tools, wherein the program code tools can be stored or stored on a storage medium for performing a method according to one of the embodiments described in this specification if the program code tools are executed on a computer or computer network.

[0081] In summary, the following implementation plan is specifically proposed:

[0082] Implementation Scheme 1: A method for detecting leakage in an application valve in an analytical system, the analytical system including an application valve fluidly connected to an eluent pump, a collection column, and a detector unit, the method comprising...

[0083] (i) The sample is applied to the collection column via the application valve;

[0084] (ii) Apply the sample from step (i) to the detector unit; and

[0085] (iii) Determine at least one sample component within the dead time of the analytical system.

[0086] Implementation Scheme 2: The method described in Implementation Scheme 1, wherein the analysis system is an automated analysis system.

[0087] Implementation Scheme 3: The method according to Implementation Scheme 1 or 2, wherein the analysis system is a liquid analysis system or a gas analysis system.

[0088] Implementation Scheme 4: The method according to any one of Implementation Schemes 1 to 3, wherein the application valve is an automatic application valve.

[0089] Implementation Scheme 5: The method according to any one of Implementation Schemes 1 to 4, wherein the application valve is a fast application valve.

[0090] Implementation Scheme 6: The method according to any one of Implementation Schemes 1 to 5, wherein the application valve has at least two switching positions, namely a sample loading position and a sample ejection position.

[0091] Implementation Scheme 7: The method according to any one of Implementation Schemes 1 to 6, wherein the sample is loaded at a pressure of at least 3 MPa at the loading position via the application valve.

[0092] Implementation Scheme 8: The method according to any one of Implementation Schemes 1 to 7, wherein the application valve comprises two elements movable relative to each other.

[0093] Implementation Scheme 9: The method according to any one of Implementation Schemes 1 to 8, wherein the application valve comprises a plurality of ports and at least one fluid connector.

[0094] Implementation Scheme 10: The method according to any one of Implementation Schemes 1 to 9, wherein the application valve comprises a detector unit port and (i) an application port and / or (ii) a sample loading port, wherein the application port and / or the sample loading port are not fluidly connected to the detector unit port at the sample loading position.

[0095] Implementation Scheme 11: The method according to Implementation Scheme 10, wherein the application port and the detector unit port are fluidly connected at the sample ejection position.

[0096] Implementation Scheme 12: The method according to Implementation Scheme 10 or 11, wherein the application port and / or the sample loading port is pressurized during sample loading.

[0097] Implementation Scheme 13: The method according to any one of Implementation Schemes 10 to 12, wherein the detector unit port is a port fluidly connected to the detector unit.

[0098] Implementation Scheme 14: The method according to any one of Implementation Schemes 10 to 13, wherein the application valve comprises at least six ports, and wherein the application port and the detector unit port are two of the at least six ports.

[0099] Implementation Scheme 15: The method according to any one of Implementation Schemes 10 to 14, wherein the application port or the sample loading port is directly adjacent to the detector unit port on the application valve.

[0100] Implementation Scheme 16: The method according to any one of Implementation Schemes 9 to 15, wherein the application valve comprises a plurality of fluid connectors arranged such that, in a switching position, each port is fluidly connected to one of its adjacent ports but not to its other adjacent port.

[0101] Implementation Scheme 17: The method according to any one of Implementation Schemes 9 to 16, wherein each of the fluid connectors independently connects two ports in the port.

[0102] Implementation Scheme 18: The method according to any one of Implementation Schemes 1 to 17, wherein the application valve comprises a rotor and a stator.

[0103] Implementation Scheme 19: The method according to Implementation Scheme 18, wherein the application valve is a six-port application valve, wherein the stator includes six ports and the rotor includes three fluid connectors, or wherein the rotor includes six ports and the stator includes three fluid connectors.

[0104] Implementation Scheme 20: The method according to Implementation Scheme 19, wherein the first application valve port is connected to the sample loading unit, the second application valve port is connected to the trapping column, the third application valve port is connected to the detector unit, the fourth application valve port is connected to the eluent pump, the fifth application valve port is connected to the trapping column, and the sixth application valve port is connected to the outlet, which is the waste outlet in this embodiment.

[0105] Implementation Scheme 21: The method according to Implementation Scheme 20, wherein, at the sample loading position of the application valve, a first fluid connector is connected to the first and second application valve ports, a second fluid connector is connected to the third and fourth application valve ports, and a third fluid connector is connected to the fifth and sixth application valve ports.

[0106] Implementation Scheme 22: The method according to Implementation Scheme 20 or 21, wherein, at the sample disengagement position of the application valve, the first fluid connector is connected to the second and third application valve ports, the second fluid connector is connected to the fourth and fifth application valve ports, and the third fluid connector is connected to the sixth and first application valve ports.

[0107] Implementation Scheme 23: The method according to any one of Implementation Schemes 1 to 22, wherein the system further includes a chromatographic column.

[0108] Implementation Scheme 24: The method according to any one of Implementation Schemes 1 to 23, wherein the sample components are determined by the detector unit, wherein the detector unit is selected from a list of the following: mass spectrometry (MS) device, diode array detector (DAD), UV / VIS detector, conductivity monitor, fluorescence detector, refractive detector, radioactive flow detector, chiral detector, evaporative light scattering detector (ELSD), and electro-fogging detector (CAD).

[0109] Implementation Scheme 25: The method according to any one of Implementation Schemes 1 to 24, wherein the method is an analytical method for determining an analyte, and wherein the sample compound detected in step (iii) is the analyte and / or an internal standard.

[0110] Implementation Scheme 26: The method according to any one of Implementation Schemes 1 to 25, wherein the leakage of the application valve is associated with the transfer of sample material onto the detector unit when the application valve is in the sample loading position.

[0111] Implementation Scheme 27: The method according to any one of Implementation Schemes 1 to 26, wherein the leakage of the application valve is caused by an increase in the space between the application valve rotor and the application valve stator.

[0112] Implementation Scheme 28: The method according to any one of Implementation Schemes 10 to 26, wherein the leakage of the application valve is caused by a failure of the element separating the application port and the detector unit port and / or the element separating the sample loading port and the detector unit port.

[0113] Implementation Scheme 29: The method according to any one of Implementation Schemes 1 to 28, wherein the leakage of the application valve is caused by wear of at least a portion of the application valve rotor and / or application valve stator.

[0114] Implementation Scheme 30: The method according to any one of Implementation Schemes 10 to 29, wherein the leakage of the application valve is caused by the formation of a fluid connection from the application port to the detector unit port and / or from the sample loading port to the detector unit port at the sample loading position.

[0115] Implementation Scheme 31: The method according to any one of Implementation Schemes 1 to 30, wherein the method is performed after at least one predetermined sample component has been detected at a non-predetermined location during a previous analytical run.

[0116] Implementation Scheme 32: An analytical system comprising an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, the analytical system being configured to perform the method according to any one of Implementation Schemes 1 to 31.

[0117] Implementation Scheme 33: The analysis system according to Implementation Scheme 32, wherein the detector unit is selected from the list of the following: mass spectrometry (MS) device, diode array detector (DAD), UV / VIS detector, conductivity monitor, fluorescence detector, refractive detector, radioactive flow detector, chiral detector, evaporative light scattering detector (ELSD), and electro-fogging detector (CAD).

[0118] Implementation Scheme 34: The analytical system according to Implementation Scheme 32 or 33, wherein the analytical system is included in a chromatography-MS system, or in the implementation scheme an LC / MS or GC / MS system.

[0119] Implementation Scheme 35: An analytical system according to any one of Implementation Schemes 32 to 34, wherein the analytical system is contained in an LC / triple quadrupole mass spectrometry (LC / TQMS) system.

[0120] Implementation Scheme 36: A method for quality assurance in sample analysis, the method comprising

[0121] (a) Determine at least one analyte in the sample.

[0122] (b) Performing the method according to any one of embodiments 1 to 31; and

[0123] (c) If a sample component is detected in step (b), the determination in step (a) is marked as invalid, and / or

[0124] (d) This causes the analysis system to prevent any further sample application to the application valve.

[0125] Implementation Scheme 37: The method according to Implementation Scheme 36, wherein the sample component in step (b) is the analyte or internal standard of step (a).

[0126] Implementation scheme 38: The method according to implementation scheme 36 or 37, wherein step (b) is performed in conjunction with step (a).

[0127] Implementation Scheme 39: A method for monitoring and analyzing the performance of an application valve included in a system, the method comprising performing the method according to any one of Implementation Schemes 1 to 31 after up to 500,000 application valve switching events.

[0128] Implementation Scheme 40: The method according to Implementation Scheme 39, wherein the method is performed after at most 1000 switching events, in Implementation Scheme 100.

[0129] Implementation Scheme 41: The method according to Implementation Scheme 39 or 40, wherein the method includes performing the method according to any one of Implementation Schemes 1 to 31 on each chromatographic run.

[0130] Implementation Scheme 42: The method according to any one of Implementation Schemes 39 to 41, wherein if at least one sample component is determined in step (c) of the method, the performance of the applied valve is classified as unsatisfactory.

[0131] Implementation Scheme 43: A method for managing at least one application valve in a management analysis system, comprising the steps of the method according to any one of Implementation Schemes 39 to 42, and the further step of replacing the application valve if the performance of the application valve is classified as unsatisfactory.

[0132] Implementation Scheme 44: Determining sample components within the dead time of the analysis system for use in detecting leakage in the applied valve.

[0133] All disclosures in all references cited in this specification, and all disclosures specifically mentioned herein, are incorporated herein by reference. Attached Figure Description

[0134] Figure 1: Schematic diagram of an exemplary six-port application valve (quick valve) in the (A) loading position and (B) unloading position in a capture and elution setup; piping connections between units are depicted with solid lines and flow directions are indicated by arrows.

[0135] Figure 2: As shown in Figure 1, but with an injection valve 30 and a loading pump 40.

[0136] Figure 3: As shown in Figure 1(A), but with valve leakage applied; the direction of sample leakage is indicated by the dashed arrow.

[0137] Figure 4: Exemplary photographs of an applied valve unit with wear and leakage, (A) applied valve rotor, (B) applied valve stator; damaged parts are indicated by arrows.

[0138] Figure 5: Chromatograms obtained in the experimental setup of Example 6 using the defective (A) and the new (B) application valves. (C) Pressure diagrams of the loading pump and elution pump for the application valves of (A) and (B).

[0139] Figure 6: An exemplary schematic diagram of a valve leakage detection algorithm.

[0140] The following examples are intended to illustrate the invention only. In any case, they should not be construed as limiting the scope of the invention.

[0141] Example 1: Non-leakage six-port quick valve (Figure 1 and Figure 2)

[0142] As a non-limiting exemplary embodiment, the six-port application valve 20 in the analysis system 10 is discussed: the stator may include six ports 110, 120, 140, 150, 160, 170, and the rotor may include three fluid connectors 210 (FIG. 1), or the rotor may include six ports 110, 120, 140, 150, 160, 170, and the stator may include three fluid connectors 210. In this configuration, in the implementation, the first valve port (sample loading port 160) is connected to the sample loading unit 360, the second valve port (first trapping column port 130, also referred to as the application port 140) is connected to the trapping column 330, the third valve port (detector unit port 120) is connected to the detector unit 320, the fourth valve port (elution pump port 110) is connected to the eluent pump 310, the fifth valve port (second trapping column port 130, also referred to as the eluent port 150) is connected to the trapping column 330, and the sixth valve port (waste port 170) is connected to the waste liquid 370. In this configuration, at the sample loading position of the application valve (Figures 1(A) and 2(A)), the first fluid connector 210 connects the sample loading port 160 and the application port 140, the second fluid connector 210 connects the detector unit port 120 and the eluent pump port 110, and the third fluid connector 210 connects the eluent port 130 and the waste liquid port 170. The sample flow entering the analysis system via sample loading port 160 (e.g., from sample loading unit 360) passes through sample loading port 160, fluid connector 210, application port 140, and reaches trap column 330, where the analyte and possibly other sample components are bound (trapped). Waste liquid flows from trap column 330 to eluent port 150, another fluid connector 210, waste liquid port 170, and into waste liquid 370. As will be understood, in the loading position of application valve 20, there is no fluid connection between sample application port 160 and detector unit port 120, or between application port 140 and detector unit port 120.

[0143] At the sample exit position of the application valve 20 (Figures 1(B) and 2(B)), the first fluid connector 210 connects to the elution pump port 110 and the eluent port 150, the second fluid connector 210 connects to the application port 140 and the detector unit port 120, and the third fluid connector 210 connects to the sample loading port 160 and the waste liquid port 170. The eluent flows from the eluent pump 310 to the elution pump port 110, fluid connector 210, eluent port 150, trapping column 30, application port 140, second fluid connector 210, detector unit port 120, and finally to the detector unit. It will be understood that at the sample exit position of the application valve 20, any sample material that has entered the detector unit port 120 will be delivered to the detector unit 320.

[0144] Example 2: Leaking six-port quick valve

[0145] As shown in Figure 4, repeated switching of the application valve 20 can lead to wear, which creates grooves in the valve material. These grooves can create undesirable fluid connections between the ports 100 and possibly outside the application valve. Figure 3 schematically illustrates how leakage in the loaded position of the application valve can cause sample material to transfer from the application port 140 to the detector unit port 120. The possible flow of the sample material is indicated by the dashed arrows.

[0146] It will be understood that there are other configurations that could lead to the same type of leakage; for example, if sample application port 160 and application port 140 are interchanged in Figures 1 through 3, sample application port 160 will be adjacent to detector unit port 120, and sample material can be transferred from sample loading port 160 to detector unit port 120.

[0147] Furthermore, the wear shown in Figure 4 is not necessarily limited to forming a connection between two adjacent ports. Therefore, it is also envisioned that sample transfer can proceed directly from the sample loading port 160 into the detector unit port 120.

[0148] Example 3

[0149] Acetaminophen was determined using the system setup essentially shown in Figure 2, with the parameters shown in Table 1. The chromatogram shown in Figure 5(A) was obtained using the leaking application valve, while the chromatogram shown in Figure 5(B) was obtained using the new application valve.

[0150] Table 1: Sample Composition and Chromatographic Method Parameters

[0151]

[0152] The results in Figure 5 show that internal leakage in the fast valve caused a decrease in the early elution peak (approximately 2 sec after injection) and the peak height and area at the expected elution time (approximately 24 sec). By replacing the defective fast valve with a new one, the early elution peak disappeared, and the initial and expected peak heights were restored.

[0153] Example 4: Leakage detection via average mean signal (reference)

[0154] The mean signal is calculated over interval A within the dead time of the analysis system, i.e., during the loading phase, interval A:=[0, T / 2], where T is the total number of measurement data points within a cycle of length T.

[0155]

[0156] (Equation (1)), where Let be the mean signal measured during interval A, and let Si be the signal of the i-th data point within that interval. The obtained values ​​are then normalized to the average background signal. The average background signal is calculated in the same way, but based on multiple measurements on different systems with known normal valve function. If this value is large, a fault is detected.

[0157]

[0158] (Equation (2)): Equation used to evaluate the degradation of valve n.

[0159] This method is simple and straightforward, but it may not be distinguishable from other errors, such as the residual signal level increase in interval A, which may also cause n >> 1, but the potential problem is not valve failure.

[0160] As a non-limiting example, when using the method according to this example, if n ~ 1, the valve is considered to be in the initial stage of the degradation process, i.e., in a state where degradation does not significantly affect the results. If n >> 1, the valve is considered defective; for example, if n = 5, significant degradation has occurred or related degradation is about to occur. If n = 2-5, the user is automatically notified and advised to contact a service technician. If n > 5, all pending measurements are stopped, and the user is informed that significant degradation that may significantly affect the reliability of the obtained results has occurred.

[0161] Example 5: Leak detection via peak analysis

[0162] As a further method for leak detection, a peak detection algorithm is applied during the loading phase (i.e., the dead time of the analysis system) and the elution phase, and the height H (or alternative ground area) of the detected peaks is measured. Subsequently, the ratio r of the peak heights in the loading and elution phases is calculated:

[0163]

[0164] Similar to Example 4, a threshold for r is defined to distinguish between normal and abnormal conditions. As a non-limiting example, r ≤ 0.1 is defined as normal, and r > 0.1 is defined as abnormal. This implies a tolerance of approximately 10% permeation. In the example of Figure 5(A) above, r = 1.7 is found, indicating complete valve failure requiring immediate replacement. We hypothesize that valve aging is a gradual process and that the evolution of r over time can be tracked to predict the point at which complete valve failure occurs. Notably, for the exemplary application valve in Figure 5(A) above, no pressure changes were detected at either the loading pump or the elution pump (Figure 5(C)).

[0165] The peak detection method in Example 5 is unaffected by the added background signal. Various peak-picking algorithms are readily available and can be used for the purposes described above. For example, the Scipy signal processing package provides the function "peak_find". This function takes a 1D array and finds all local maxima by simple comparison of adjacent values. A peak or local maximum is defined as any value whose two direct neighbors have smaller amplitudes, and its position and height are retrieved as outputs. The input parameter "peak saliency" can be used to adjust the sensitivity of the function in peak detection. This parameter is a measure of how much a peak stands out from the surrounding baseline of the signal and is defined as the vertical distance between the peak and its lowest contour line. In Example 5, a value of 2000 is effective for detecting the dominant peak and ignoring irrelevant peaks. In general, this value can be fixed or made dependent on determination to obtain increased specificity.

[0166] In summary, the algorithm for the method used in Example 4 on several analytical measurements can be represented as shown in Figure 4.

[0167] Example 6: Valve Leakage Detection Algorithm

[0168] An exemplary, non-limiting algorithm for valve leakage detection based on measurements as described above is schematically illustrated in Figure 6:

[0169] The measured input data is fed into algorithm 600. Preprocessing 610, such as filtering or smoothing, is applied to the input data to remove noise that may interfere with the peak picking process. Peak picking 620 is performed to determine the peak height and / or peak area in the loading and elution stages. If several peaks are found in each stage, the highest peak may be selected for further steps. From the identified peaks, r, i.e., the peak height ratio or peak area ratio, is calculated 630. Based on the obtained r value, an alarm 640 may be issued, for example, if r is not less than 0.01; in such cases, the flow to the affected application valve may be interrupted to prevent any further operation. If the judgment criteria are met, for example, if r < 0.01, no alarm 650 is issued. In both cases, i.e., with or without an alarm, the method may optionally continue by loading the next incoming dataset.

[0170] Figure Labels

[0171]

Claims

1. A method for detecting leakage in an application valve in an analytical system, said analytical system comprising an application valve fluidly connected to an eluent pump, a collection column, and a detector unit, the method comprising... (i) The sample is applied to the trapping column via the application valve; (ii) Apply the sample from step (i) to the detector unit; as well as (iii) Determining at least one sample component within the dead time of the analytical system. The detection of at least one sample component within the dead time of the analytical system indicates leakage in the applied valve.

2. The method according to claim 1, wherein the application valve is an automatic application valve, preferably a rapid application valve.

3. The method according to claim 1 or 2, wherein the application valve has at least two switching positions, the at least two switching positions being a sample loading position and a sample removal position.

4. The method according to any one of claims 1 to 3, wherein the sample is loaded at a pressure of at least 3 MPa at the loading position via the application valve.

5. The method according to any one of claims 1 to 4, wherein the application valve comprises a detector unit port and (i) an application port and / or (ii) a sample loading port, wherein the application port and / or the sample loading port are not fluidly connected to the detector unit port at the sample loading position.

6. The method of claim 5, wherein the application port and the detector unit port are fluidly connected at the sample ejection location.

7. The method according to claim 5 or 6, wherein the detector unit port is a port fluidly connected to the detector unit.

8. The method according to any one of claims 5 to 7, wherein the application port or the sample loading port is directly adjacent to the detector unit port on the application valve.

9. The method according to any one of claims 5 to 8, wherein the application valve comprises a plurality of fluid connectors arranged such that, in a switching position, each port is fluidly connected to one of its adjacent ports but not to its other adjacent port, preferably wherein each of the fluid connectors independently connects to two of the ports.

10. The method according to any one of claims 1 to 9, wherein the sample components are determined by the detector unit, wherein, in an embodiment, the detector unit is selected from the list of: mass spectrometry (MS) devices, diode array detectors (DAD), UV / VIS detectors, conductivity monitors, fluorescence detectors, refractive detectors, radioactive flow detectors, chiral detectors, evaporative light scattering detectors (ELSD), and electro-fogging detectors (CAD).

11. The method according to any one of claims 1 to 10, wherein the method is an analytical method for determining an analyte, and wherein the sample component detected in step (iii) is the analyte and / or an internal standard.

12. The method according to any one of claims 1 to 11, wherein the method is performed after at least one predetermined sample component has been detected at a non-predetermined location during a previous analytical run, or after at most 50,000 switching events of the applied valve, in one embodiment at most 5,000 times, in another embodiment at most 1,000 times, and in another embodiment at most 100 switching events.

13. An analytical system comprising an application valve fluidly connected to an eluent pump, a trapping column, and a detector unit, wherein the detector unit is preferably selected from the list of: mass spectrometry (MS) devices, diode array detectors (DAD), UV / VIS detectors, conductivity monitors, fluorescence detectors, refractive index detectors, radioactive flow detectors, chiral detectors, evaporative light scattering detectors (ELSD), and electro-fogging detectors (CAD). The analysis system further includes a data processing unit adapted to perform or cause the analysis system to perform the method according to any one of claims 1 to 12.

14. A method for quality assurance in sample analysis, the method comprising: (a) Determine at least one analyte in the sample. (b) Performing the method according to any one of claims 1 to 12; and (c) If a sample component is detected in step (b), the determination in step (a) is marked as invalid; and / or (d) Causes the analysis system to prevent any further sample application to the application valve.

15. The purpose of determining sample composition within the dead time of the analytical system is to detect leakage in the applied valve.

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