This invention relates to the analysis of molecules by mass spectrometry. In particular, this invention relates to the analysis of molecules that may possess multiple charges by mass spectrometry by increasing the charge on the molecules.
The analysis of large molecules by mass spectrometry is playing an increasingly important role in modern biological research. Such analysis is facilitated by recent advances in the ability to generate gas-phase ions of these large molecules. Mass spectrometry is particularly suited to the analysis of these biological materials because they are often available only in small quantities, typically as isolates from natural sources. In addition, mass spectrometry is well suited to the area of proteomics, which includes the study of the time-dependent protein complement of an organism.
Large biologically derived molecules are often biopolymers such as proteins, peptides, nucleic acids, oligonucleotides, polysaccharides, and oligosaccharides. The identification and analysis of these biopolymers are dependent on a sufficient amount of fragments, and a sufficient diversity in the fragmentation patterns that may be generated during mass spectrometry. These fragments and fragmentation patterns are interpreted to derive the primary sequence of the biopolymers. Further, the number of and diversity of the fragments generated by mass spectrometry is often dependent upon the number of and diversity of the charges and charge states that may be generated on the molecule. Processes and apparatus for manipulating and modifying these charge states to optimize the observed fragmentation will expand the field of mass spectrometric analysis of biologically derived molecules.
Processes for analyzing molecules by mass spectrometry are described herein. These processes use the technique of generating an initial charge on the molecule to be analyzed, placing that molecule into a gaseous phase, and manipulating or modifying that initial charge.
The initial charge is generated using techniques that include but are not limited to plasma desorption ionization (PDI), field desorption ionization (FDI), electrohydrodynamic ionization (EHI), laser desorption ionization (LDI), matrix assisted laser desorption ionization (MALDI), atmospheric pressure matrix assisted laser desorption ionization (APMALDI), spray ionization techniques, including electrospray ionization (ESI), nano-electrospray ionization, thermospray ionization (TSI), aerospray ionization (ASI), sonic spray ionization (SSI), particle desorption methods such as fast atom bombardment (FAB), fast ion bombardment (FIB), such as secondary ion mass spectrometry (SIMS), and massive cluster impact, and the like.
After the initial charge is generated, an ion storage or trapping procedure is performed. Ions may be stored or trapped in any of a variety of devices including but not limited to linear ion traps, quadrupole ion traps, and the like. In some embodiments of the processes described herein, the initially generated ions are directed to the ion storage device by illustratively using a turning quadrupole, and the like.
In one illustrative embodiment, the initial charge is manipulated by contacting the initially charged molecule, or any charged fragments of that molecule, with at least one multiply charged ionic compound. The initial charge may be a single charge of either polarity or a multiple charge of either polarity. After contacting the initially charged molecule or charged fragment thereof with at least one multiply charged ionic compound, the initial charge on the molecule or fragment thereof is changed. Illustratively, the absolute value of the initial charge is lowered, or the polarity of the initial charge is changed. In the latter case where the polarity of the initial charge is changed, the absolute value of the resulting charge may therefore be greater than the absolute value of the initial charge.
In another illustrative embodiment, the initially charged molecule or charged fragment thereof is contacted with more than one multiply charged ionic compound. After contacting the initially charged molecule or charged fragment thereof with one multiply charged ionic compound, the initial charge on the molecule or fragment thereof is changed. Illustratively, the absolute value of the initial charge is lowered, or the polarity of the initial charge is changed. In the latter case where the polarity of the initial charge is changed, the absolute value of the resulting second charge may therefore be greater than the absolute value of the initial charge. Subsequently, the molecule or fragment thereof having the resulting second charge is contacted with a second multiply charged ionic compound. After contacting the molecule or fragment thereof having the second charge with the second multiply charged ionic compound, the resulting second charge on the molecule or fragment thereof is changed. Illustratively, the absolute value of the second charge is lowered, or the polarity of the second charge is changed. In the latter case where the polarity of the second charge is changed, the absolute value of the resulting third charge may therefore be greater than the absolute value of the second charge.
In one aspect, the polarity of initial charge is changed, and the resulting charge has an absolute value greater than the absolute value of the initial charge. In another aspect, the initial charge has an absolute value greater than one, and the resulting charge has an absolute value of one and the opposite polarity of the initial charge.
In another aspect, the polarity of the initial charge is changed after contacting a multiply charged ionic compound, and the polarity of the resulting second charge is changed back, or reverted, to the polarity of the first charge after contacting a second multiply charged ionic compound. In another aspect, the absolute value of the final charge is greater than the absolute value of the initial charge.
Prior to the manipulation or modification of the charge on the molecule or fragment thereof, or following the manipulation or modification of the charge on the molecule or fragment thereof, an optional mass filtering step is performed on one or more of the ions generated in the processes described herein. This mass selection step may be performed on the molecular ions, including fragment ions thereof prior to contact, or following contact with the multiply charged ionic compounds. This mass selection step may also be performed on any of the multiply charged ionic compounds that are contacted with the molecular ions or fragment ions thereof.
Prior to the manipulation or modification of the charge on the molecule or fragment thereof, or following the manipulation or modification of the charge on the molecule or fragment thereof, an optional mass analysis step is performed on one or more of the ions generated in the processes described herein. The mass analysis step may use any of a variety of techniques including but not limited to scanning from a linear ion trap, scanning from a quadrupole ion trap, time of flight (TOF), ion cyclotron resonance, and the like.
The processes described herein may also include a step of concentrating the charge of the molecule or fragment thereof using any of a variety of techniques including but not limited to atmospheric sampling glow discharge ionization (ASGDI), and the like.
Apparatus for performing the processes described herein are also described. Various configurations are constructed from those components necessary to perform the steps of the processes described herein, including (1) various sources for generating ions or charges on molecules or analytes to be analyzed, (2) various components for storing or trapping those molecular ions or fragment ions thereof so-generated, including components for directing ions to such storage components, (3) various components for optionally filtering or selecting certain masses from those molecular ions or fragment ions thereof, (4) various components for providing a reaction environment for contacting other charged or multiply charged ionic compounds with those molecular ions or fragment ions thereof to modify the charge states of the molecular ions or fragment ions, and (5) various components for analyzing the molecular ions or fragment ions thereof.
The initial charge on a molecule or analyte to be ultimately analyzed by mass spectrometry may be generated by any of the variety of techniques including but not limited to those commonly employed in mass spectrometric analysis, such as plasma desorption ionization (PDI), field desorption ionization (FDI), electrohydrodynamic ionization (EHI), laser desorption ionization (LDI), matrix assisted laser desorption ionization (MALDI), atmospheric pressure matrix assisted laser desorption ionization (APMALDI), spray ionization techniques, including electrospray ionization (ESI), nano-electrospray ionization, thermospray ionization (TSI), aerospray ionization (ASI), sonic spray ionization (SSI), particle desorption methods such as fast atom bombardment (FAB), fast ion bombardment (FIB), such as secondary ion mass spectrometry (SIMS), and massive cluster impact, and the like. The initial ionizing source includes but is not limited to fast atoms such as xenon, argon, and the like, slow electrons, high energy electrons, and clusters such as glycerol clusters, and the like. It is appreciated that the processes described herein are not limited by the method used to generate the ions used in these processes, and further that the ionization method may use either charged or uncharged substances to generate the initial charge on the molecule or analyte.
It is appreciated that depending upon the nature of the analyte or molecule to be detected, either a positive charge or a negative charge may be generated during initial ionization, and therefore the selection of the ionization technique is conveniently chosen after considering the nature of the analyte or molecule. Further, in order for an analysis to take place, a variety of procedures may be followed to introduce a sample containing the analyte or molecule into the apparatus wherein the initial charge is generated. Those procedures include but are not limited to those common to the field of mass spectrometry, such as introduction of the sample as a solution, which is aspirated, volatilized, or atomized, or introduction of the sample as a solid or liquid, which may be adhered to a probe or other solid surface, such as a matrix.
It is also appreciated that certain molecules may form a positive charge more easily than a negative charge, while other molecules may form a negative charge more easily than a positive charge. For example, ambident molecules, such as peptides and proteins, may form with relative ease either a positive ion or a negative ion. However, certain carbohydrates may form positive ions more readily than negative ions. In contrast, certain oligonucleotides and nucleic acids may form negative charges more readily than positive ions.
Peptides and proteins are analyzed by the processes and apparatus described herein, and may be widely varying in both primary sequence and molecular weight. Peptides and proteins, that may be analyzed include but are not limited to bradykinin, ubiquitin, cytochrome C, hemoglobin, and the like. Oligosaccharides and carbohydrates are analyzed by the processes and apparatus described herein, and may be widely varying in both primary sequence and molecular weight. The oligonucleotides and nucleic acids that may be analyzed by the processes described herein are widely varying in both primary sequence and molecular weight. The oligonucleotides and nucleic acids that may be analyzed include but are not limited to oligonucleotides isolated from natural sources and synthetic oligonucleotides, having illustratively from about 5 to about 50 nucleotides. In addition, nucleic acids, including nucleic acids from plants, animals, bacteria, and the like, including very large nucleic acids that have more than about 100 kilobases may be analyzed using the processes and apparatus described herein.
It is understood that any molecule that may be ionized into a gaseous state using any of the techniques described herein or any other technique known to produce ionized molecules in the gas phase, may be analyzed by the processes described herein provided that charge states of both polarities may be formed on the molecule, and that at least one of those charge states has an absolute value greater than one. Though the generation of multiple charges on a molecule or analyte prior to analysis is more often desired in molecules and analytes having high molecular weights, the processes and apparatus described herein are equally applicable to the analysis of smaller molecules, where it is desirable to generate higher charge states or multiple charges thereon. Without being bound by theory, the generation of multiple charges on a molecule may account for an increase in the number of fragments generated, and to the degree of, and the diversity of the fragmentation observed. It is appreciated that increased fragmentation and increased fragmentation diversity may enhance the ability to identify the chemical composition or chemical structure, including the primary sequence of large molecules, such as various biological compounds including proteins, nucleic acids, and polysaccharides.
Depending upon the technique used to generate the initial charge, and depending upon the nature of the molecule to be detected, the initial charge may therefore be positive or negative. In addition, the initial charge on the molecule may have an absolute value of one or alternatively, the initial charge on the molecule may have an absolute value greater than one. In the case where the initial charge is greater than one, the initial charge may be optionally concentrated to a lower selected charge by using a suitably configured concentrating component that uses the technique of ion parking. Ion parking is generally described in U.S. Pat. No. 4,849,628 to McLuckey et al., the disclosure of which is incorporated herein by reference. It is appreciate that the technique of ion parking is advantageously used in processes where the molecular ions or fragment ions form a population of charge states of simultaneously varying values. Such a condition may complicate the interpretation of the resulting mass spectrum. Concentrating the various charge states to a single state will generally simplify the mass spectrum obtained.
Alternatively, using a conveniently placed component such as an atmospheric sampling glow discharge ionization (ASGDI) source, and the like, the absolute value of the initial charge may be reduced one charge at a time, in a stepwise fashion. An ASGDI is capable if reducing the absolute value of both positively charged and negatively charged molecular ions or fragments thereof In one aspect the ASGDI source uses monoanions of perfluorocarbons, such as perfluoro-1,3-dimethylcyclohexane (PDCH). In the case where the initial charge is both greater than one and positive, the initial charge may be optionally lowered to a lower selected charge, including a charge of negative one, by using a suitably configured electron capture dissociation (ECD) component.
In either the case of an initial ionization having an absolute value of one or greater than one, the charge of the molecular ion so generated, may be changed to a charge having an absolute value equal to or greater than one, but of opposite polarity to the charge generated initially. Such a conversion is accomplished by contacting the molecular ion, illustratively in the gas phase, with a multiply charged ionic compound. A great variety of multiply charged ionic compounds may be used for this purpose of polarity change in the processes described herein.
The multiply charged ionic compounds useful in the processes described herein include but are not limited to polyionic compounds having multiple functional groups that may each form a net charge or a plurality of charges under conditions that may be created in typical mass spectrometric analyses and apparatus. Such molecules include but are not limited to compounds having a plurality of amine groups, heteroaromatic amine groups, imine groups, hydrazine groups, guanidine groups, carboxylic acid groups, sulfonic acid groups, phosphonic acid groups, and the like. The multiply charged ionic compounds described herein include polymeric materials, such as dendrimers, and naturally occurring polymeric materials such as nucleic acids and proteins. Illustratively, (5′-d(AAAA)-3′)3−, (melittin+4H)4+, (horse myoglobin+20H)20+, (holomyoglobin+9H)9+, and cytochrome C and apomyoglobin carrying multiple charges are useful in the processes described herein.
It is appreciated that such compounds may possess a mixture of like-charged groups, such as compounds that possess both carboxylic acid groups and sulfonic acid groups, or compounds that possess both ammonium and iminium groups. Further, it is appreciated that a mixture of compounds may be used in the processes described herein for changing the charge on the analyte or molecule to be detected, such as a mixture of a polyamine compound and a polyimine compound.
Multiply charged ionic compounds having a plurality of negatively charged groups include aromatic groups having a plurality of carboxylic acid groups present thereon, including alkylcarboxylates, cycloalkylcarboxylates, arylcarboxylates, such as phenyl-dicarboxylic, -tricarboxylic, and tetracarboxylic acids, arylalkylcarboxylates, such as phenyl-dipropionic, tripropionic, and tetrapropionic acids, and phenyl-diacetic, triacetic, and tetraacetic acids, and the like. In addition, dendrimers having multiple carboxylic acid, sulfonic acid, or phosphonic acid functionalities are useful in the processes described herein.
Multiply charged ionic compounds having a plurality of positively charged groups include aminoalkyl, hydrazinoalkyl, guanidinoalkyl, aminocycloalkyl, hydrazinocycloalkyl, guanidinocycloalkyl, aminoaryl, hydrazinoaryl, guanidinoaryl, heteroaromatic amines, such as pyridinyls, pyrimidinyls, pyridazinyls, pyrazinyls, imidazolinyls, oxazolihnyls, pyrrolyls, pyrazolyls, indolyls, quinolinyls, quinazolinyls groups, and the like. Illustratively, dendrimers having multiple ammonium or iminium groups, including alkyleneammonium and alkyleneiminium functionalities are useful in the processes described herein. Dendrimers useful in the processes described herein include but are not limited to cyclotriphosphazene-phenoxymethyl(methylhydrazono) (PMMH) dendrimers, Polypropylenimine hexadecaamine Dendrimers, Polypropylenimine dotriacontaamine Dendrimers, Polypropylenimine tetraamine Dendrimers, Polypropylenimine tetrahexacontaamine Dendrimers, any of the various polyamidoamine (PAMAM) dendrimers, and any of the various primary-amine terminated dendrimers.
It is understood that many dendrimers are commercially available as a mixture of dendrimers with a single compound having a specified number of chargeable functional groups present thereon. In these cases, it is appreciated that following the preparation of the multiply charged ionic compound from the dendrimer, a mass selection procedure may be advantageously included as a step in the processes described herein. Such a mass selection procedure will have the benefit of decreasing the noise observed in the final mass spectrum and simplifying the interpretation of the resulting mass spectrum. The mass selection process may be accomplished using any of a variety of techniques common to mass spectrometry, including quadrupole ion traps, linear ion traps, time of flight (TOF), and ion cyclotron resonance. In such a step, the lower and higher molecular weight compounds, as well as the compounds that do not have the desired number of chargeable groups present in the multiply charged ionic compound precursor are removed. The resulting purified multiply charged ionic compound may then be contacted with the molecule or analyte. Similarly, in certain aspects of the processes described herein, a mass selection or mass filtering step may be included where the initially generated molecular ions or fragment ions thereof are separated. In these embodiments, only certain charged ionization states, or certain mass ranges are contacted with the multiply charged ionic compounds. A process that uses the principle of tandem mass spectrometry may include such a filtering or selection procedure.
It is also appreciated that during the course of a polarity change on the molecule or fragment thereof, in some embodiments of the invention, aggregates of these ions and the multiply charged ionic compound may form. These aggregates also have the overall effect of changing the charge on the molecule, including changing the charge on the molecule to a value opposite in polarity. However, in these cases the charge is changed by the aggregation of the multiply charged ionic compound with the analyte ion rather than the transfer of either an electron or a proton between the multiply charged ionic compound and the analyte or molecular ion. In either case, the detection of the analyte by mass spectrometry is accomplished by evaluating the masses that are produced in the apparatus. Mustratively, a protein having an initial charge of for example of positive one, may form an aggregate in the ion trap with a multiply charged ionic compound that is for example an aryl(tetraalkylenecarboxylate), where the aggregate has an overall charge of negative three.
In configurations of the processes described herein where there is included a tandem mass spectrometry procedure, also known as MS/MS, it is understood that such aggregates may yet be analyzed by these techniques commonly in use, following a heat treating step, where the aggregate molecule comprising the analyte or molecular ion and the multiply charged ionic compound are separated in a fashion where the analyte of molecular ion retains the increased charge state.
The polarity change of the initial charge on the analyte or molecule or fragment thereof may result in a second charge state that has an absolute value greater than or equal to the absolute value of the initial charge state. In other aspects, the polarity change of the initial charge on the analyte or molecule or fragment thereof may result in a second charge state that has an absolute value less than or equal to the absolute value of the initial charge state. Further, the third charge may have an absolute value greater than or equal to the absolute value of the second charge state. Illustratively, the third charge state has an absolute value greater than the absolute value of the initial charge on the analyte or molecule or fragment thereof.
It is understood that the molecule or analyte may undergo fragmentation at various times during the operation of the processes described herein to produce fragment ions, and at various times during the modification of the charges thereon. The foregoing discussion regarding the subsequent processes and process steps apply equally to these fragment ions as to the molecular ions or analyte ions described above. For example, a protein-based analyte may illustratively fragment upon initial ionization to a plurality of charged ion fragments, each having a first charge that may be the same or different. The unfragmented molecular ion may also be included in this group. Each molecular ion or fragment ion thereof is then contacted with a multiply charged ionic compound, or a mixture of such compounds that change the charge state to a second charged state, the polarity of which is the opposite of the initial charge state. Optionally, this group of now-oppositely charged molecular or molecular fragment ions may be contacted with a second multiply charged ionic compound, or a second mixture of such compounds capable of changing the charge state to a third charge, where the third charge on any given molecular or molecular fragment ion is the same or different, but where each is the opposite polarity of the second charge. Additional fragmentation of the molecular ion or other fragmentation ions produced in the initial ionization may occur following either or both steps wherein the molecular ions or fragment ions are contacted with the one of more multiply charged ionic compounds. In the case of multiply charged molecular ions or fragment ions, further or differential fragmentation is optionally accomplished by including an electron capture dissociation (ECD) procedure, a collision-induced dissociation procedure, and the like.
As described herein, a mass filtering or mass selection procedure may also be introduced in the process following the modification of the initial or even the second charge state on the molecule or fragment thereof. This mass filtering or mass selection procedure is optionally accomplished using a scanning technique, such as by scanning the masses from a linear or quadrupole ion trap, and the like.
It is appreciated that the change between the first and second charge states, and the change between the second and third charge states may occur as a stepwise procedure, wherein a dwell time is introduced into the process to allow a substantial number of molecules or molecular fragments thereof to change charge states. Subsequently, the change between the second and third charge states is performed.
Alternatively, the change among charge states may occur in a contemporaneous manner, wherein a mixture of multiply charged ionic reagents or compounds comprises both positively charged ionic compounds and negatively charged ionic compounds. In this variation of the process, the molecular ion or molecular fragment ion undergoes two changes in polarity, each oppositely directed, in a one-step process. It is appreciated that the selection of positive and negative multiply charged ionic compounds is accomplished in a way to enhance the population of molecular ions or fragment ions that have a desired charge state. This enhancement may be derived through an understanding of the thermodynamics controlling the overall modification of the charge state on the molecular ion or fragment ion thereof. In one aspect, desirable thermodynamics favor the formation of a charge state on the molecular ion or molecular fragment ion that is the same as the initial charge state thereon, passing through a second charge state that is the opposite of the initial charge state, and having a final charge with an absolute value that is greater than the absolute value of the initial charge state. In one illustrative aspect, the second charge state has an absolute value of one. In another illustrative embodiment, both the initial charge state and the second charge state have an absolute value of one, and the third charge state has an absolute value greater than one.
The molecular ions generated in the processes described herein may also be analyzed, where such analysis includes detection of molecular ions or fragments thereof, and measurement of the masses of these molecular ions or fragments thereof that are produced in the processes or by the apparatus described herein. Such detection and measurement may be performed by using any of the variety of techniques commonly employed in mass spectrometric analysis, including but not limited to scanning from quadrupole ion traps, linear ion traps, analysis via time of flight (TOF), ion cyclotron resonance, and the like. Further, the detection and analysis may also include a tandem mass spectrometry configuration.
The processes described herein may be performed by any variety of mass spectrometry equipment that is common to mass spectrometric analysis and has been configured with devices or components that are capable of performing the procedures required by the processes described herein. In general, the apparatus described herein are constructed from components or devices that are capable of performing functions required by the processes described herein, such as (1) various sources for generating ions or charges on molecules or analytes to be analyzed, (2) various components for storing or trapping those molecular ions or fragment ions thereof so-generated, including components for directing ions to such storage components, (3) various components for optionally filtering or selecting certain masses from those molecular ions or fragment ions thereof, (4) various components for providing a reaction environment for contacting other charged or multiply charged ionic compounds with those molecular ions or fragment ions thereof to modify the charge states of the molecular ions or fragment ions, and (5) various components for analyzing the molecular ions or fragment ions thereof. It is appreciated that the apparatus described herein may also be capable of performing other processes in addition to those described herein. Such flexibility possessed by an apparatus should not be understood to suggest that apparatus is excluded from the invention described herein
The second housing component 14 possesses a fourth generating source 40, such as an atmospheric sampling glow discharge ionization source, interfaced with the second housing component 14 through a fourth interface region 42. The fourth interface region 42 is coupled to a fourth general ion optical element 44 which is aimed into the quadrupole ion trap 46. The charges on ions that have been generated in the first housing component 12 and diverted into the ion trap 46 in the second housing component may be changed by the introduction of multiply charged ionic compounds as described herein, and generated illustratively in the first housing component 12, or by ions generated from the fourth ion source 40. The second housing component 14 also possesses a detector 48, such as an electron multiplier, for detecting ions released or ejected from the ion trap 46.
In
In addition,
Illustratively, one source is used to generate an initial charge on the molecular ion. As described herein, the generation of the molecular ion may be accomplished using any of a variety of components that are capable of generating ions using plasma desorption ionization (PDI), field desorption ionization (FDI), electrohydrodynamic ionization (EHI), laser desorption ionization (LDI), matrix assisted laser desorption ionization (MALDI), atmospheric pressure matrix assisted laser desorption ionization (APMALDI), spray ionization techniques, including electrospray ionization (ESI), nano-electrospray ionization, thermospray ionization (TSI), aerospray ionization (ASI), sonic spray ionization (SSI), particle desorption methods such as fast atom bombardment (FAB), fast ion bombardment (FIB), such as secondary ion mass spectrometry (SIMS), and massive cluster impact, and the like.
In one embodiment, the source is a SSI source capable of generating both positively and negatively charged multiply charged ionic compounds. In that embodiment, the molecular ion or fragment ion may be sequentially contacted with each of these ionic compounds generated from the same source. In one illustrative configuration, the SSI source is combined with a linear ion trap. This configuration of the apparatus described herein is well-suited for a flow through process, where the source generates the initial charge on the molecule, and the resulting ions or fragment ions thereof are placed in the storage trap. Subsequently, the SSI source is used to generate a multiply charged ionic compound or compounds having one polarity. These ions are allowed to flow through the linear ion trap, which unlike a quadrupole ion trap is capable of holding only one polarity of charge at a time. After some predetermined time, the SSI source is used to generate a multiply charged ionic compound or compounds having the other polarity. This ion stream is then contacted with the molecular ions or fragment ions in the linear trap.
The apparatus described herein is illustratively configured to include a turning quadrupole capable of directing the ions so produced into a storage device or trap, such as a quadrupole ion trap or a linear ion trap. In order that multiple sources may be conveniently placed on the apparatus, a turning quadrupole may advantageously be used to direct the ions of interest into other components of the apparatus, such as the contacting or reaction vessel component. The various ions generated from the various corresponding sources are directed into the turning quadrupole by an electrostatic lens, a series of electrostatic lenses, an electrostatic lens assembly, such as an Einzellens, or other suitably configured general ion optical element. Initially, the molecular ion or fragment thereof is turned into the contacting component or storage trap.
In an alternate embodiment, the generating component may be configured to include a mass selecting component or mass filtering component, such as a quadrupole ion mass filter, linear ion mass filter, and the like. The mass selecting component will have the effect of purifying the ions, such as the molecular ions or the fragment ions, that are introduced into the contacting component via the turning quadrupole. This purification procedure may be applied to the initially generated molecular ions, or to any of the multiply charged ionic compounds used to modify the charges on the molecular ions or fragment ions thereof during the contacting procedure. Each source may be optionally configured with such a mass selecting component.
Apparatus described herein also include a contacting or reaction vessel component. In some embodiments, the contacting or reaction vessel component is a quadrupole ion trap or a linear ion trap. The contacting or reaction vessel component allows for the molecular ions or fragment ions initially generated, and optionally filtered to be contacted with multiply charged ionic compounds that will change the charges on the molecular ions or fragment ions. It is appreciated, that like the molecular ions or fragment ions that are initially generated, the charges generated on the multiply charged ionic compounds may also be mass filtered or mass selected prior to introduction into the reaction vessel component.
The contacting component includes a trap for the purpose of holding for a predetermined period of time the molecular ions or fragments ions thereof in contact with any one of the multiply charged ionic compounds. In addition, the trap portion of the contacting component has the purpose of holding the molecular ions or fragments ions thereof for subsequent ion concentration or ion parking procedures. In some configurations of the apparatus described herein, the ion storage device or ion trap is a linear ion trap or a quadrupole ion trap. A linear ion trap is a two-dimensional trap that may only store one charge state polarity at a time. A quadrupole ion trap is a three-dimensional trap that may store two charge state polarities at a time.
In apparatus where the generating component and the contacting component are separate, the generating component and the contacting component may be in fluid communication by a suitably placed ion transmission system, such as an electrostatic filter assembly, tube lens, an RF ion guide, and the like, or an electrodynamic ion transmission system such as an hexapole or an octapole, and the like. Alternatively, the tube lens may be configured to include a single mass selecting component that can be used as a mass filter of any substance generated by the various ionizing sources and directed into the ion trap. In this embodiment, the necessity of having a mass selecting component at each source may be thus alleviated.
Further, the contacting component or reaction chamber may also include a mass selecting or mass filtering component, a mass scanning component, and a mass detecting component, such as an electron multiplier. It is to be understood that while the apparatus illustrated in
Illustratively, the mass scanning component may be a TOF component rather than a quadrupole mass filter. In that configuration, a quadrupole ion trap is used to store and contact the molecular ions, and the TOF component is used to analyze the masses resulting from the charge modification. In addition, the mass storage and contacting component may be a linear ion trap, where the multiply charged ionic compounds are contacted with the molecular ion or fragment ion in a flow-type process rather than the batch-type process illustrated by the quadrupole ion storage embodiment. In this configuration, a SSI source is conveniently selected as it has the ability to generate both positive and negative ions, and is particularly suited for this type of flow-through embodiment. However, it is appreciated that various configurations of these components may be constructed to perform the processes described herein.
In some configurations of the apparatus described herein, analyte ions or fragment ions thereof, or molecular ions or fragment ions thereof are detected. Detection is accomplished by any of the variety of techniques common to the field of mass spectrometry, and includes electron multipliers, and the like. Prior to the detection, a mass selection or mass filtering procedure is optionally performed on the analyte ions or fragment ions thereof or molecular ions or fragment ions thereof. Such a mass filtering or mass selection procedure may be accomplished by scanning the masses with a quadrupole. In other embodiments, the mass filtering or mass selection procedure may be accomplished using a time of flight component that in one step permits separation of the masses.
In embodiments having a quadrupole ion trap serving as the contacting component, the ions to be analyzed and detected are released using any of a variety of common techniques, such as resonance ejection and the like, into the detecting component. In one aspect the detecting component is an electron multiplier and the like. In other aspects, the detecting component includes a conversion dynode.
The foregoing discussion sets forth many possible embodiments with various aspects of the invention described herein for both the processes and apparatus. However, it is appreciated that other embodiments arising from combinations of the components described herein or the procedures described herein though not explicitly set forth are nevertheless contemplated to fall within the scope of the invention as described herein. Further, the following illustrative examples are presented to provide an additional understanding of the nature and the spirit of the invention. The examples are intended to be illustrative and should not be considered to limit the invention in any way.
Filing Document | Filing Date | Country | Kind | 371c Date |
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PCT/US03/17022 | 5/30/2003 | WO | 11/29/2005 |