Method for quantitatively identifying a substance by mass spectrometry

Abstract

The invention relates to a method for the quantitative determination of a chemical substance S from a sample using a mass spectrometer having at least one detector. In line with the invention, a sample which may contain the substance S of interest, or a conversion product of the sample, is analyzed in the mass spectrometer. For the analysis the mass spectrometer is alternately set at least for masses SM1, SM2, so that each of the masses is detected multiple times and all of said masses are detected by the same detector. The masses SM1 and SM2 are fictitious neighboring masses for a mass CM of the substance S with a particular isotope content. The quantity of the mass CM is ascertained by means of calculation from the measured values for the masses SM1, SM2.

Claims

1. A method for the quantitative determination of a chemical substance S contained in or derived from a sample using a mass spectrometer having at least one detector, comprising steps of: providing an ion beam for detection that is free from passage through a split ion detector; operating the mass spectrometer in a non-scanning, alternate peak detection mode; setting the mass spectrometer to detect using a common detector at least masses SM1 and SM2, so that each of the masses SM1 and SM2 is detected at least once, wherein the masses SM1 and SM2 are fictitious neighboring masses which are at defined distances D1 and D2 from a central mass CM, wherein the mass CM is a mass of the substance S with a particular isotope content, wherein SM1 is heavier than CM and SM2 is lighter than CM, wherein the masses SM1 and SM2 are not further masses of the substance S, and wherein the distances D1 and D2 are each shorter than a peak width for the mass CM at prescribed resolution; wherein substantially a full beam of ions hits an ion detector when the system is set to CM and only part of the full beam of ions reaches the detector when set to SM1 or SM2; evaluating from the measured intensity values for masses CM, SM1 and SM2 whether there is interference from the mass CM with other masses; in reliance of the result of the interference evaluating step, determining the quantity of the mass CM by a selected one of: (i) setting the mass spectrometer for the mass CM and detecting the mass or (ii) by means of calculation from the measured intensity values for the masses SM1 and SM2.

2. The method as claimed in claim 1, wherein each of the distances D1, D2 corresponds to the half peak width HWHM of the mass CM.

3. The method as claimed in claim 1, wherein the mass spectrometer is alternately set to detect using a common detector at least the masses SM1, SM2 and CM, so that each of the masses SM1, SM2, CM is detected at least once.

4. The method as claimed in claim 1, wherein the mass spectrometer is alternately set to detect using a common detector at least for the neighboring masses SM1, SM2 of the mass CM and for the mass RM, so that each of the masses SM1, SM2, RM is detected at least once, wherein the mass RM is a mass of the substance S and has a different isotope content than the mass CM, and wherein a measured intensity value for the mass RM is also used for evaluating the interference with the mass CM.

5. The method as claimed in claim 1, wherein components of the sample or a conversion product of the sample are temporally resolved using a chromatographic method prior to the mass spectrometry analysis.

6. The method as claimed in claim 5, wherein the chromatographic method is a gas chromatography method.

7. The method as claimed in claim 1, wherein the mass spectrometer comprises a sector field mass spectrometer, a double-focusing mass spectrometer, or a quadrupole mass spectrometer.

8. The method as claimed in claim 7, wherein the mass spectrometer is set to detect the various masses by adjusting an electrical field.

9. The method as claimed in claim 1, wherein the mass spectrometer includes only a single detector having a detector inlet gap.

10. The method as claimed in claim 1, wherein the mass spectrometer includes at least one of the following ion sources: a) an electron impact ion source, b) a chemical ionization ion source, c) a field ionization ion source, d) a field desorption ion source, e) a fast atom bombardment (FAB) ion source, f) an atmospheric pressure ionization (API) ion source, g) a laser desorption or matrix-assisted laser desorption ionization ion source, h) a photoionization ion source, i) an electrospray ion source, j) a thermospray ion source, k) a plasma desorption ion source, l) a secondary ion (SIMS) ion source, m) a thermal desorption ion source, and n) an inductively coupled plasma (ICP) ion source.

11. The method as claimed in claim 1 wherein the width of the detector substantially matches the width of the ion beam.

12. The method as claimed in claim 1 wherein the width of the ion beam is the same for each of the settings CM, SM1 and SM2.

13. The method as claimed in claim 1 where possible interference ions could enter the detector when setting the MS to the defined distances D1 or D2.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) Further features of the invention can be found in the description in other respects and in the claims. Exemplary embodiments of the invention are explained in more detail below with reference to drawings, in which:

(2) FIG. 1 shows a simplified illustration of an apparatus for carrying out the method according to the invention, namely a mass spectrometer having an upstream gas chromatograph and a connected computer system for evaluating the accruing data,

(3) FIG. 2 shows a detector with an inlet gap and a two-dimensional illustration of the transiting ion beam of the detected ions in accordance with a particular set mass,

(4) FIG. 3 shows an illustration similar to FIG. 2, but for a different set (adjacent) mass, so that in this case a portion of the ion beam is kept back (shadowed) from the gap,

(5) FIG. 4 shows an illustration similar to FIG. 3, with the same ion beam, but with the mass spectrometer set for an opposite adjacent mass, the ion beam being shadowed to an even greater extent,

(6) FIG. 5 shows an illustration of adjacent mass peaks with reciprocal interference, namely a tetradioxin and a tetrafuran,

(7) FIGS. 6 to 12 show schematic illustrations of (chromatographic) peaks for the masses Q0 and R0 and of peaks Q1 and Q2 adjacent to the peak at the position PQ0,

(8) FIGS. 13 to 15 show illustrations similar to FIGS. 6 to 12, but with the addition of adjacent masses R1, R2 to the mass R0.

DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS

(9) In order to carry out the method according to the invention, a mass spectrometer MS is used in this case which, as shown in FIG. 1, may be of customary design, namely with an inlet system ES, an ion source IS, a mass analyzer MA and a detector D. Upstream of the inlet system ES is a device for chromatographic separation, for example a gas chromatograph GC or a liquid chromatograph LC. The signals arising on the detector D are processed and conditioned by a computer system CS. Preference is given to an implementation with a gas chromatograph GC, an EI ion source, a double focusing mass analyzer and a detector with an inlet gap.

(10) What is intended to be examined is a particular pollutant content in a food sample, for example. The food sample is pretreated in a known manner. The ingredients are temporally resolved in the gas chromatograph GC, so that with a particular dwell time a target substance (pollutant) is predominantly supplied to the inlet system ES. Typically, the target substance is known and only the quantity thereof needs to be determined. An example of this inherently known method is cited in EPA 1613. Reference is hereby made to this document in its entirety.

(11) The mass analyzer is set to a position PM0 for a mass M0 of the sought pollutant, so that the relevant ions theoretically hit the detector D in FIG. 2 centrally, see the dashed line 20 therein as a continuation of the central, relatively long arrow 21, which represents the ion beam from the mass M0. Naturally, the ions enter the detector D with a certain (rate) scatter and in so doing pass through a collector gap 22. In practice, various gaps or slots or openings may be provided at this point. The collector gap referred to is usually the inlet gap of the detector. This function can also be performed by an outlet gap of the mass analyzer. Similarly, an outlet gap in the mass analyzer and a collector gap in the collector may be provided in succession. To simplify matters, only the collector gap 22 is mentioned in this case. What is important in this connection is the possible shadowing of a portion of the ion beam on a gap in this region of the mass spectrometer. The quantity of ions reaching the detector D is shown in FIG. 2 by the two rectangles 23, 24.

(12) The mass analyzer also contains the ion beam from the mass M0. During this, the mass analyzer MA is adjusted by a difference D1 for a different mass, in this case for an adjacent heavier mass position PM1, see FIG. 3. Theoretically, all ions from the mass M0 hit precisely the left-hand edge of the collector gap 22 or of the detector 10. The statistical scatter of the ions gives rise to a distribution such that one portion of the ions reaches the detector D, see rectangular area 26, while the other portion of the ions cannot pass through the collector gap 22, see hatched area 27.

(13) Next, the mass analyzer is adjusted by an amount D2 for a somewhat lower position PM2 than the mass position PM0, see FIG. 4. In this case, the adjustment is made to the extent that the position PM2 is opposite the position PM1 and even outside of the collector gap 22 or of the detector D. In FIG. 4, a quantity of ions entering the detector D is obtained in line with a rectangle 29 and a quantity of masked-out ions is obtained in line with the hatched rectangle 30.

(14) With reference to the inlet gap 22, the position PM1 is preferably a half gap width next to the position PM0. Usually, the width of the collector gap 22 is tuned to the resolution of the mass spectrometer and is mechanically adjustable. The adjustment by said half gap width to the left then corresponds to the adjustment of the mass position by a half peak width HWHM (= FWHM), see also FIG. 5. The amount D1 therefore corresponds to the half gap width and also to the half of the (full) peak width FWHM in this configuration.

(15) In practice, the gap width is set once and then not altered again as far as possible, at any rate not during the determination of the substance. Only the mass which is set on the mass spectrometer is changed, for example by changing the voltage of the electrical sector in a double focusing mass spectrometer. This change can be made very quickly.

(16) The position PM2 in FIG. 4 is situated more than a half gap width next to the position PM0 only for the purposes of illustrating the different adjustment options. Preferably, the position PM2 is set such that it differs from the position. PM0 by the same amount as the position PM1. This is not absolutely necessary for the application of the invention, however.

(17) The adjustment of the mass analyzer for differing mass positions PM1, PM2 also affects the effective resolution of the appliance. Assuming that there is a resolution R of 10 000 for the setting shown in FIG. 2, the masking-out of the half ion beam shown in FIG. 3 results in an increase in the effective resolution R to 20 000. A further shift, for example in a similar manner to FIG. 4, results in shadowing of 75% of the ion beam and accordingly in an effective resolution of R=40 000.

(18) Similarly, the mass transferred in a quadrupole mass analyzer can be adjusted by a portion of the peak width, for example such that the response to an undisturbed peak decreases to 50% of the response in the peak center.

(19) The various masses PM0, PM1, PM2 are selected in succession and repeatedly. The presence of interference for the mass M0 can be derived from the intensities IM1, IM2, measured at the positions PM1 and PM2.

(20) FIG. 5 shows the simulated peaks in a mass scan using two closely adjacent masses, namely m/z=319.90 for (2, 3, 7, 8 tetradioxin), m/z=319.94 for (2, 3, 7, 8-13C tetrafuraninternal standard labeled with 13C atoms).

(21) What can be seen is an example of the determination of the half peak width indirectly, namely as peak width (FWHM) at half peak height. Other kinds of determination of the half peak width are possible and also known.

(22) The two peaks coincide with one another in the lower region, so that quantitative determination of a target mass from one of the two masses without corrective measures produces an incorrect result. The ascertained quantity as the area below the peak is greater than the quantity which is actually present, because ions from the adjacent mass are included in the detection of the target mass. In order to avoid or correct this the method according to the invention is used. The adjacent masses M1 and M2 are detected in addition to the examined target mass M0. The results are used for carrying out different computation steps and comparisons. In an approximate division, two essential steps can be distinguished from one another:

(23) a) checking the target mass M0 for interference with adjacent masses,

(24) b) quantitatively determining the target mass and the proportion of the pollutant in the sample.

(25) As shown by the illustration in FIGS. 6 to 15, the quantitative determination of a substance involves up to six different masses being detected and being used for further calculations (more are possible but not preferred):

(26) Typically, these are the target mass (quantification mass) QM, with the exact mass position PQ0 (central mass) and the associated, adjacent mass positions PQ1 and PQ2, and the comparison mass RM with the associated exact mass position PR0 and the adjacent mass positions PR1 and PR2. In the prior art (cf. EPA 1613), only the ratio of IQ0 to IR0 is used for qualifying the target mass. The quantification is then based on IQ0 alone or on IQ0 and IR0, relative to a calibration standard.

(27) Since the sought pollutant is known, the distribution of the masses with the different isotope contents within said pollutant is also known. The different masses/isotopes have an almost constant statistical distribution relative to one another in the pollutant. In the event of discrepancies between the relative intensities and this distribution, it can therefore be assumed that measurement errors or interference with other masses is/are present.

(28) As shown in FIG. 6, a simple method is used to detect the (total of four) intensities IQ0, IQ1, IQ2 from QM and IR0 from RM. It is possible and even simpler to measure without IQ0. A comparison is performed between the two intensities at the positions PQ1 and PQ2, which are preferably at the same distances from the position PQ0. If the intensities are essentially the same, it is assumed that there is no interference. The intensity IQ0 can then be calculated from IQ1, IQ2 or from both, as desired by the user. In the simplest case, for which the expected intensities of an interference-free peak are=2IM1=2IM2=IM0, the most reliable calculation of IQ is: IQ0=IQ1+IQ2. Good results can also be attained, with IQ0=2IQ1 or IQ0=2IQ2, however.

(29) An additional check for interference can be attained by comparing the intensities IQ1 and IQ2 with the intensity IR0 of the comparison mass RM. Naturally, it is also possible to carry out the conventional approach for comparing measured or calculated values IQ0 to IR0 (and this approach must be carried out if the method disclosed in EPA 1613 is to be followed). If no interference is indicated, the target substance can be quantified from the intensity IQ0 alone or from IQ0 and IR0 together.

(30) FIG. 6 shows the (chromatographic) peak areas for the mass intensities IQ1 and IQ2 as triangles of the same size so as to illustrate interference which is not present. Merely for the purpose of simplification, the triangle for IQ0 is the same size as that for IR0.

(31) The experiment may involve the measurement of an internal standard for a similar compound (for example the target substance, in which ail carbon atoms have been replaced by 13C, the heavier and usually less frequent carbon isotope) which is considered to be usually free of interference. In this case, it would be sufficient to measure the intensities of the isotope peaks in the internal standards which correspond to QM and RM in the target substance. The results are used for calculating the relative isotope rate. This allows the content of the target substance in the sample to be ascertained (usually on the basis of a previously performed quantification calibration) and for the purpose of ascertaining a possible compliance with limit values in the case of pollutants. Finally, all validated, measured data can be added for the quantification. This improves the overall accuracy of the calculation.

(32) FIG. 7 shows the possible relationships between the four masses shown in FIG. 6. The following ratios can be calculated and assessed:

(33) a) IQ1 to IQ2 (triangular areas b/a); if the resultant number is significantly different than 1, there is interference;

(34) b) IR0 to IQ1 (triangular areas c/b) and IR0 to IQ2 (c/a); if these two results are different, there is interference;

(35) c) IR0 to the sum of IQ1+IQ2 (c/(a+b)); the resultant ratio is intended to match the known isotope pattern of the known pollutant if there is no interference.

(36) Similar and equivalent calculations can easily be derived from the teachings of these examples.

(37) Levels of significance can be determined from principles of ion statistics or can be prescribed by experienced users. By way of example, a typical, expected measurement accuracy for the intensities for the instrument is +/10%. In this case, a ratio of 1.1 to 0.9=1.22 with a deviation of less than 25% from the basic value would not be regarded as an indication of interference. If the expected intensity accuracy is +/20%, for example when the value is closer to the detection limit, a ratio of approximately 1.5 would still be acceptable.

(38) FIG. 8 shows interference. As indicated above, the different masses are detected and the results compared with one another. It is possible to see the larger area b for IQ1 in comparison with the smaller area a for IQ2. Accordingly, IQ0 at the mass position PQ0 has interference on the right at the position PQ1. The ratio of IQ2 to IR0 may therefore be in order, while the ratio of IQ1 to IR0 does not correspond to the statistical value. Furthermore, the ratio IQ1 to IQ2 is significantly different than 1. Finally, the ratio of IQ0 to IR0 is also different than the expected value. Assuming that interference is present only on one side, namely at the position. PQ1, the other value, that is to may IQ2, can be used for the quantification. The absence of interference for IQ2 can be assumed if the ratio of 2IQ2 to IR0 corresponds to the expected (statistical) isotope ratio.

(39) The mass AM may also be influenced by interference. This case is illustrated in FIG. 9. IR0 (size of the triangle c) therein is significantly above the value which can be expected statistically. By contrast, the ratio of IQ1 to IQ2 is correct, which means that there is probably no interference for IQ0 and the value can be used for quantification. IQ0 can be adopted from direct measurement or by calculation from IQ1 and IQ2, as described above.

(40) Further possible measurement results are shown in FIG. 10. IQ0 is much larger than could be expected statistically. However, there is no imbalance, which means that IQ1 and IQ2 are approximately the same. The fact that there is interference therefore results only from comparison of the intensities for QM with the intensities for RM.

(41) Interference relating to a plurality of masses is shown in FIG. 11. None of the ascertained ratios meets the expectation, this also applying to IQ2 to IR0 (a/c). Assuming that the smaller values are not subject to interference, the measured value IQ2 (the area a) could be used for quantitative determination.

(42) A special case is also shown in FIG. 12. In this instance, there is interference on the values IQ1 and IQ2 and on the measured IQ0. The associated areas a, b and c, like the metrologically or computationally ascertained area for IQ0, are larger than could be expected statistically. Quantitative determination of the pollutant is not possible with the measurements. In the unfavorableor improbablecase, IQ1 is approximately as large as IQ2, which means that no interference is assumed for the measured values and they are used for the quantification, unless a comparison with IR0 is performed by RM at the position PR0.

(43) The time or quantity of samples available for the measurement is usually highly limited. This applies particularly under chromatographic conditions, with GO peaks which are only a few seconds wide, for example. This limits the measurement cycles to as few masses as possible in order to allow maximum dwell times for the detected masses. Secondly, the determination of further masses can avoid the risk of unrecognized or quantification-disturbing interference. This is discussed in the section below.

(44) In the example in FIG. 13, six masses are detected, with the intensities IQ0, IQ1, IQ2 for the quantification mass and the corresponding group of three containing the intensities IR0, IR1, IR2 for the comparison mass.

(45) The additional values IR1 and IR2 allow further ratios to be calculated and compared with the values which can be expected statistically, for example the ratios of the areas a to d and b to e. This would allow the situation shown in FIG. 12 to be checked in more detail. It is also possible for sums to be related to one another, for example the areas (d+e)/(a+b). The setpoint values thereof can be compared with additionally measured internal standards. FIG. 14 shows an illustration of measured values with which no interference is associated.

(46) FIG. 15 in turn shows the instance of interference for IQ0, specifically in the right-hand half thereof, that is to say with reference to IQ1. The ratio of IQ2 to IR2 (area a to d), which, with knowledge of the ratio to the overall intensity, can be used for quantification, corresponds to the value that can be expected.

(47) In FIGS. 7 to 15, some of the triangular areas are linked by arrows. Each arrow represents the calculation of a ratio for the associated areas a to e. Dotted arrows indicate interference, while continuous arrows mean that no interference can be assumed.

(48) It is worth mentioning that for a prescribed overall detection timeat least for the case of no interferencein a method in which IM1 and IM2 are measured instead of IM0, only half of the total number of ions detected are used for the calculation. Better ratio values can be ascertained if the measurement of the target mass and the intensity IM0 thereof is not omitted. In other words: additional information and certainty can be obtained with minimal involvement.