MXPA98007444A - Simultaneous analysis of multiple samples and apparatus for im - Google Patents

Abstract

Apparatus and analytical methods to process multiple samples in a simultaneous manner. Radiation, such as laser light, desirably includes plural wavelengths (12, 14) across multiple samples (32a-d) in a simultaneous manner, such as by directing a beam of radiation along a single path through all the samples. The response to each wavelength is monitored by monitoring an induced effect, different from the intensity of the applied radiation itself (42). It obtains useful proportions of the signal to the noise with a low absorbency in each sample. A sample is desirably of a known composition, and serves as an inter-calibration standard.

Description

SIMULTANEOUS ANALYSIS OF MULTIPLE SAMPLES AND APPARATUS FOR THE SAME The present invention relates to methods and apparatus for analyzing a plurality of samples, such as gas samples, by their exposure to radiation, such as laser light. Analytical test methods involve the transmission of radiation through a sample of the material to be tested, commonly referred to as an "analyte". For example, some photometric tests measure the amount of light absorbed by a sample at a particular wavelength associated with a particular substance. If a particular chemical strongly absorbs red light, the amount of the substance in the sample can be determined by directing a beam of red light through the sample. A photodetector measures the amount of red light that remains in the beam after passing through the sample. The higher the lighting cnt, the less red light remains in the beam. Many variations of this basic scheme are known through the use of different wavelengths of radiation. Because the amount of light that reaches the photocell depends on the operation of a light source, such as the amount of illumination provided by a lamp, a reference beam can be directed from the lamp along the same path when the sample is removed, to provide a reference or calibration reading, or along a separate path to a separate photodetector, to provide a cnuous calibration reading. Also, where the analyte can cin several different substances, each of which absorbs light at a different wavelength, the sample can be tested at each of these different wavelengths to determine the amount of each substance. Typically, photometric measurements of this type are used to monitor the amounts of different chemical substances, that is, different elements or compounds in the sample. As described in U.S. Patent No. 5,394,236 to Daniel E. Murnick, another measurement technique can be used to determine the amounts of particular atomic isotopes present in a sample. Isotopes are different forms of the same chemical element, which have an atomic nucleus of different mass. For example, naturally occurring carbon consists predominantly of 12C, that is, carbon having an atomic mass of 12 atomic mass units ("amu"). Other carbon isotopes are 13C and 14C, which have masses of 13 or 14 masé, atomic units, respectively. The 1 C is radioactive, while the 13C and the 12C are stable non-radioactive materials.

Certain preferred methods taught in the '236 patent involve directing one or more light beams through a sample that includes multiatomic fractions, such as carbon dioxide molecules or ions cining different isotopes. Preferred methods include the same step of providing the analyte in a condition such that some of the species carrying isotope in the analyte are present in excited states. Normally, the analyte is kept in this excited condition by keeping the analyte in an ionized gas or "plasma". When at least some of the electrons in the molecules or ions are at higher energy levels than the energy levels occupied in the earth or normal stage of the isotope-bearing species. These excited states have associated "transition energies" corresponding to the energy released on the transition from the excited state to a lower state, or absorbed after the inverse transition, from the excited state to another higher energy state. More preferably, the species carrying isotope are multiatomic fractions, such as ions or multiatomic molecules. The transition energies are different for isotope-bearing species that incorporate different isotopes, as for example, 13C02 and 12C02. In the preferred methods according to Murnick's' 236 Patent, radiation, such as light incorporating plural wavelengths corresponding to the transition energies of excited isotope-carrying species. The incorporation of different isotopes to the sample is applied. Light at each wavelength interacts with species that include an isotope, and does not substantially interact with species that include the other isotope. "By measuring the response of the analyte to the radiation applied at different wavelengths, you can determine the amounts of the different isotopes present in the sample In the particularly preferred methods according to the '236 patent of Murnick, the response of the sample is measured by monitoring the changes in the electrical impedance of the plasma, caused by the light at the different lengths of wave, commonly referred to as the "opto-galvanic effect." As disclosed in the '236 patent, light can be provided at plural wavelengths by one or more laser devices in a single beam with light at different wavelengths that varies at different frequencies, for example, light at a wavelength associated with 13C02, can be activated and deactivated at a first frequency. modulation, while light at the wavelength corresponding to 13C02 can be activated and deactivated at a second modulation frequency. The electrical signal corresponding to the opto-galvanic effect includes two separate components, one at the first modulation frequency representing the amount of 12C02, and another at the second modulation frequency representing the amount of 13C02. These can be electronically separated from each other, and are measured to provide a pair of signals representing the relative amounts of the two isotopes. Preferred methods according to the '235 Patent of Murnick provide numerous advantages over other methods employed for the determination of the amount of different isotopes in a substance. The methods and apparatus according to the '236 Patent can be reapplied to many different analytes for many different purposes. However, an especially useful application of these methods is medical testing. Different medical and scientific procedures require the determination of the relative amounts of the different isotopes. In certain medical tests, a test compound includes a rare isotope such as 13C in the compound. The test compound is administered to the subject. The amount of the rare isotope that appears in the body fluids of the subject or in their respiration, depends on the ability of the subject to metabolize or process the test compound. Accordingly, the amount of the rare isotope or the ratio of the rare isotope, such as 13C, to the more common isotope, such as 12C, indicates the ability of the subject to metabolize the test compound. One of these tests involves the administration of urea labeled with 13C, to the subject orally. If the subject has heliobacter pylori bacteria present in the gastrointestinal tract, 13C will be incorporated into the carbon dioxide produced by the patient, and will be exhaled as part of the patient's breathing. Accordingly, the ratio of 13C to 12C in patient respiration indicates whether heliobacter pylori is present or not. Other breathing tests involve the administration of other labeled compounds, or with carbon isotopes, or with isotopes of other elements. Apparatus and methods for the isotopic analysis of substances face different conflicting requirements. Analytical devices should be able to process as many samples as possible per unit of time. Typically, the camera demonstrates that it contains the sample during the analysis, it is a permanent component of the instrument. Therefore, a delayed process of purging the sample chamber and introducing a new sample between each test of a series of tests must be performed. Even when the real test can be done quickly, the total production or the speed of the sampling process of the instrument is limited by this procedure. Although it would be possible to increase the test speed by duplicating the test instrument, this solution would be expensive. Moreover, it would introduce an additional source of variation in that the test readings would require a cross calibration to compare the characteristics of the different instruments with each other, in such a way that a reading obtained in an instrument would be directly comparable with the data obtained in another instrument. Even when only one instrument is used, its calibration may be out of phase or changed from time to time. To provide a useful comparison between the samples, the instrument must be recalibrated repeatedly by testing known samples. In turn, this further reduces the time available to test real samples. These problems are particularly important in the case of tests where the analyzes of different samples are compared with one another. In certain medical tests, plural samples of bodily fluids are taken from a particular subject at different times. For example, in the urea respiration test as discussed above, samples of respiration can be collected before administration of the labeled urea, and one or more times after administration. The evaluation of the test may involve comparison between the "before" and "after" samples. It is important that any effects of variation among the instruments, or variation of a single instrument from time to time, do not amplify or diminish the differences between the plural samples. In accordance with the above, there have been substantial needs for improvements in methods and apparatus for testing analytes by exposure to radiation.

Compendium of the Invention The present invention solves these needs. One aspect of the present invention provides a method for analyzing an analyte, which includes the steps of maintaining a plurality of samples separated from the analyte. The method according to this aspect of the invention further includes the step of directing radiation that includes a wavelength corresponding to a transmission energy of each of the species through plural samples, by directing one or more beams of radiation through all the samples in an order upstream to downstream. Typically, the samples are kept in separate chambers arranged on a trajectory, and in beam is directed along the trajectory, to pass through all the cameras in sequence, and in this way simultaneously expose all the samples to the radiation in an essentially simultaneous way. The methods according to this aspect of the invention also include the steps of monitoring the interaction between the applied radiation and the samples, by monitoring an induced effect caused by the radiation applied to the samples. As used in this disclosure, the term "induced effect" means a phenomenon other than the change in the intensity of the radiation at the applied wavelengths. Induced effects include the optoacoustic effect, - the stimulated fluorescence, and the opto-galvanic effect. The supervision of an induced effect can provide useful proportions of the signal to noise, even when only a small fraction of the radiation applied by each sample is absorbed. More preferably, the applied radiation suffers little or no net change in intensity as it passes through each sample chamber. The samples at the downstream end of the path receive essentially the same radiation intensity as the samples at the upstream end. Moreover, any variation in absorption by the samples at the upstream end of the path produces only a minute change in the intensity applied to the samples at the downstream end. For all practical purposes, the intensity of radiation applied to the samples at the downstream end can be considered independent of the possibility of absorption of the samples at the upstream end. The step of directing the beam may further include the step of reflecting the beam through the cameras, such that the beam passes both in the upstream and downstream directions through the chambers one or more times. This further reduces the differences in the intensity of radiation applied between the samples at the upstream end of the path, and the samples at the downstream end of the path. As a comparison, where the interaction is monitored in a conventional manner, by monitoring the intensity of the radiation applied after passing through the sample, the signal representing the interaction of the radiation applied to the sample is the difference between the intensity of the applied radiation and the intensity of the radiation after passing through the sample. Any noise or fluctuation in the applied radiation appears as noise in the signal representing the interaction. This noise obscures the signal that the interaction represents. To provide a useful ratio of signal to noise, each sample must absorb a substantial amount of the applied radiation, and the amount of radiation absorbed by each sample must vary substantially depending on the composition of the sample. For these reasons, common photometric instruments typically do not direct a single beam of light through plural samples in series. The ability to direct light through plural samples in series in the methods according to this aspect of the invention, leads to very significant benefits. Because a single beam can be directed through several samples in a simultaneous manner on a single optical path, the number of samples processed per unit time, or the production speed of the instrument, can be multiplied several times. This can be done using a simple optical configuration, which includes only an optical path. Because several samples can be exposed to radiation in a single simultaneous operation, the variation in the operation of the radiation-producing elements of the instrument will not affect the comparisons between these samples. In a particularly preferred embodiment, plural samples simultaneously tested using a single beam of light may include samples taken from a single patient in a medical test, such as, for example, breath samples taken from a single patient at different times, such as before and after the administration of a test substance. This allows a particularly accurate comparison between the results for the different samples. Preferably, one of the plural samples is a sample of a known composition. The results observed within the known composition serve as a calibration reference. In this configuration, the instrument is calibrated each time a sample is measured. Any change in the characteristics of the incident radiation beam is detected. Accordingly, the results observed within known samples can be corrected to compensate for any changes. Because the calibration can be performed in a simultaneous manner with the tests of the unknown samples, the production of samples does not decrease substantially. More preferably, the samples are kept in a condition where at least one species to be detected is in an excited state, and the wavelengths of the applied radiation correspond to the transition energy of each of these species in their excited state. Preferably, the samples are maintained in plasmas. The step of supervising an induced effect preferably includes the step of supervising the opto-galvanic effect caused by the applied radiation. The term "opto-galvanic effect" refers to the change in the electrical impedance of a plasma, caused by the applied radiation. The opto-galvanic effect provides an easily measurable electrical signal, even where the plasma absorbs only a small portion of the applied radiation. Moreover, where the applied radiation includes a wavelength c [ue that corresponds to a transition energy of a species in an excited state, each sample will emit some radiation at that wavelength through a process known as stimulated emission. The relationship between the amount of radiation emitted and the amount absorbed will depend on the properties of the plasma, such as the proportion of atoms or molecules of the species that are in the excited state. The net effect on the beam passing through each sample may be a decrease in intensity, or an increase in intensity. Preferably, however, the amount of radiation emitted is slightly greater than the amount absorbed, such that the net increase in beam intensity caused by the passage through a sample compensates for the attenuation caused by the passage of the beam through the walls of the sampling chamber. In other words, the sample itself can provide an intensity gain of slightly more than unity, while the sample and the camera together can have an intensity gain of about one unit. More preferably, the radiation beam directed through the plural samples includes a plurality of wavelengths corresponding to the transition energies of a plurality of species that may be present in the analyte samples. Desirably, the method also includes the step of comparing the responses for each sample in each wavelength, with the response of the same sample in the other wavelength, to measure the relative abundances of the different species in each of these samples. For example, where the different wavelengths correspond to the transition energies of the species that incorporate different isotopes, the method can provide a measure of the relative abundances of the different isotopes in each sample.

In accordance with a further aspect of the invention, a method for analyzing analytes can include the steps of maintaining plural samples of the analyte, and directing radiation that includes plural wavelengths corresponding to the transition energies of a plurality of species through the plural samples, in such a way that the radiation passes from a common source of radiation through all the samples in a substantially simultaneous manner, and therefore, all samples will be exposed substantially to the same radiation, despite any change or variation in the operation of the radiation source. Methods of conformance with this aspect of the invention further include the steps of monitoring the response of the samples to the radiation, to determine a response for each of the wavelengths, and comparing the responses for each sample at each wavelength. , with other responses from the same sample to the wavelengths to produce a measure of the relative abundances of the species in each of these samples. This step can be done by determining a response ratio between the magnitudes of the responses of each sample, at different wavelengths. Here again the samples may include at least one reference sample having a known composition, and at least one unknown sample. The method may include the step of adjusting the measure of relative abundance for each unknown sample, based on the responses for the reference sample. This step can be done by calculating a proportion in the aforementioned response ratio for the unknown sample, and the response rate for the standard sample. As discussed further below, these radio-electric calculations can cancel the effects of changes in instrument conditions. The methods according to this aspect of the invention may include the other features discussed above. Accordingly, the step of monitoring the responses of the samples to the applied radiation may include the step of monitoring an induced effect. Here again, the samples can be kept in a condition where the species to be determined are in an excited state. The step of directing radiation through the plural samples in a simultaneous manner, may include the step of directing a beam of radiation through the plural samples, such that the same beam passes through all the samples in sequence. . Methods of conformance with the above aspects of the present invention more preferably include the step of loading the plural samples into plural chambers in a simultaneous manner. When the samples are gaseous, the loading step may include the steps of evacuating a plurality of sampling chambers in a simultaneous manner; admit different samples in the individual evacuated chambers in a simultaneous manner, and bring the different samples up to a previously selected pressure, removing portions of each sample from the respective chambers in a simultaneous manner. Other aspects of the present invention provide an analytical apparatus. The apparatus according to one aspect of the invention includes a plurality of sampling chambers configured along an optical path in an upstream to downstream order, each sampling chamber having an upstream end and a downstream end, and transparent walls at the ends upstream and downstream. The apparatus may include a frame, and the cameras may be permanently mounted to the frame in alignment with one another along the optical path. The apparatus further includes an element that introduces the analyte into at least one of the chambers. The apparatus also includes a radiation source at one or more wavelengths of previously selected analysis, and an element for directing this radiation in a beam along the upstream extension upstream of the path, through all the sampling chambers. Additionally, the apparatus includes an element for monitoring an induced effect caused by the radiation, to thereby monitor the response of the analyte disposed inside each chamber to this radiation. More preferably, the light source may include one or more laser devices. The apparatus desirably includes an excitation element to apply energy to the analyte disposed within each chamber, to carry the species contained within this analyte to the excited states. The excitation element may include an element for applying electrical energy, such as radiofrequency energy, to the samples contained in the different chambers. Accordingly, the excitation element may include one or more radiofrequency coils connected to a common source of excitation energy, such as a common radiofrequency energy unit. The apparatus may further include a loading element for loading the samples in the different chambers. The charging element can be operated in cycle, to load all the cameras with different samples in a single cycle. Preferably, the loading element includes an element for charging a standard analyte of a known composition in one or more of the chambers in each cycle. Alternatively, one of the chambers may have a standard analyte permanently sealed therein. The apparatus according to this aspect of the present invention can be used to perform the methods discussed above. These and other objects, features, and advantages of the present invention will become clearer from the detailed description of the preferred embodiments stipulated below, taken in conjunction with the accompanying drawings.

Brief Description of the Drawings Figure 1 is a diagrammatic view illustrating portions of the apparatus in accordance with one embodiment of the invention. Figure 2 is an additional diagramatic view illustrating additional portions of the apparatus illustrated in Figure 1.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT The apparatus according to one embodiment of the invention includes a frame 10, a first laser device 12, and a second laser device 14 mounted on the frame. Each of the laser devices is a conventional gas laser device. As described, for example, in the aforementioned '236 patent, a conventional gas laser device includes a tube filled with a gas mixture, discharge electrodes adapted to create an electrical discharge inside the tube, and optical components such or Brewster or polarization windows, and a partially reflective exit mirror. The first laser device L2 has a tube filled with a mixture of 13C02 in a carrier gas, such as a helium-nitrogen mixture. Its optical components are configured in such a way that the light at a wavelength corresponding to an ion transition energy of 13C02 in an excited state, desirably at approximately 11,200 nanometers, is amplified inside the tube. The first laser device 12 also includes an energy source 13 for applying an excitation potential between the electrodes, and creating a discharge inside the tube. Accordingly, the laser device 13 is adapted to emit the first beam 16 of infrared light at the first wavelength, which corresponds to an ion transition energy of 13C02 in the excited state. The second laser device 14 includes similar structures, but its gas tube is filled with a mixture containing 12C02 in the inert carrier gas, and its optical components are configured to amplify the light at a second wavelength, desirably about 10,600. nanometers, which corresponds to a transition wavelength of 12C02 ions in the excited state. The second laser device 14 also includes a power source (not shown) adapted to apply an excitation voltage across the electrodes in the tube. These known elements are adapted to cooperate with each other, to emit a second beam 18 of infrared radiation consisting essentially of light at a second wavelength corresponding to the ion transition energy of 12C02 in the excited state. The apparatus further includes a steering mirror 20 adapted to direct the beam 18, and a combination optics 22 adapted to combine the two beams into a single beam 24. The optical components, including the combination optics 22, are configured to attenuate the beam from the second laser device 14 (at the second wavelength associated with 12C02), to a greater degree than they attenuate the beam from the first laser device 12 (at the first wavelength associated with the 13C02) . For example, the combining optics may include a partially transmitting and partially reflecting element configured in such a way that the beam from the second laser device 14 is transmitted through the element, while the beam from the first laser device 12 is reflected from the. The characteristics of this element can be selected in such a way that the transmitted beam suffers substantially greater attenuation than the reflected beam. Accordingly, if the first and second laser devices produce approximately equal intensities, the combined beam 24 will have substantially greater intensity at the first wavelength than at the second wavelength. As discussed further below, this configuration compensates for the greater abundance of the species associated with the second wavelength in the samples to be analyzed. A mirror is adapted at the upstream end 26 to receive the combined beam 24, and to direct it along an optical path coincident with the axis 28. A mirror is provided at the downstream end 30, at the downstream end of the path 28, to reflect the beam back to the upstream end, that is, back to the mirror 26. All mirrors and optical components of the laser device are mounted, directly or indirectly, on the frame 10, and therefore, they are kept in alignment with each other by the frame. Additional conventional optical components, such as collimator lenses, filters, and the like, can be incorporated into the laser devices 12 and 14, or they can be placed along the different trajectories of the beam. These may be used, in a conventional manner, to provide a well focused collimated beam along path 28. Four sample cells 32 are mounted on frame 10 on optical path 28. Sample cell 32a is a container substantially closed which defines an internal volume, and which has a gate 34a connected to the internal volume. The sample cell 32a has a transparent upstream end wall 36a, and a transparent downstream end wall 38a. The term "transparent" is used herein in the ordinary sense to indicate that the end walls transmit a substantial proportion of radiation to the first and second wavelengths. However, even the transparent end walls typically attenuate the radiation to some degree. The cell is aligned in such a manner that the end walls 36a and 38a extend generally perpendicular to the upstream direction downstream of the optical path 28. The sample cell 32a is formed of one or more dielectric materials. For example, the entire cell, including the end walls 36 and 38, may be formed of quartz or other crystals. The other cameras 32b, 32c, and 32d have similar characteristics. The chambers are formed in a row, on the common axis 28 of the trajectory, giving the end walls of each chamber towards the directions upstream and downstream. A coil 40 is provided in proximity to each chamber 32. Each coil is electrically connected to a separate excitation and detection unit 42. Each excitation and detection unit includes a conventional source 43 of radio frequency ("RF") alternating potential, connected in a circuit with the associated coil 40. Each unit 42 also includes a conventional detector 45 for monitoring the current and voltage across the coil in the circuit, and for providing a signal representing the electrical impedance of a gas discharge within the associated chamber 32. The signal outputs from the excitation and detection units 42 are connected by conventional electronic elements, symbolized by a busbar 48, to a signal processing unit 50. The signal processor 50 is adapted to convert the analog signals from the units of detection 45, in digital signals. The signal processor, therefore, includes conventional amplification, filtering, and analog-to-digital conversion equipment. The digital outputs from the signal processor 50 are connected by means of the digital data bus 52 to a control computer 54. The control computer may include generally conventional computer elements, such as a central processing unit, control devices, etc. data storage, including random access memory and mass storage memory, as well as an internal data bus. The computer is also equipped with output control units 56 and 58 adapted to be connected to the control inputs of the power supplies 13 and 15 of the laser devices 12 and 14. The output control units can be interface cards conventional computer, and can be connected to the control inputs of the laser devices, through conventional control links. The computer also connects to the output communication equipment 60, such as a display screen, a printer, a data storage device such as a disk drive or a tape drive, or a computer network. The output device is configured in such a way that the results derived by computer 54 can be displayed in human readable form, stored for later retrieval, or both. The link is configured in such a way that the computer 54 can command the laser devices 12 and 14 to vary their light output. Normally, this is done by varying the energy input to the laser device. In accordance with the foregoing, the internal power supplies 13 and 15 of the laser devices are adapted to receive commands from the computer, and to vary the energy supplies towards the electric discharge in accordance with those commands. The apparatus further includes a sample handling and driving system illustrated in Figure 2. This system includes a vacuum pump 70, which may incorporate a conventional vacuum tank, and a vacuum manifold 72 connected to the suction gate of the bomb. A standard operating manifold 74 is also provided. The gate 34a of the first chamber 32a is connected to a subsystem of the first chamber 76. The subsystem 76 includes a node directly connected to the gate 34a of the chamber, and a pressure sensor. 78 connected to the node 77. A main evacuation valve 80 and an injection valve 82 are also connected to the node 77. The injection valve 82 is a solenoid operated valve adapted to rapidly cycle on / off, and configured to move from completely open until completely closed within a few milliseconds. In turn, the injection valve 82 is connected to a gate of a multi-port manifold valve or "air lock" 84. An additional gate valve 84 is connected to a calibration valve 86, which in turn it is connected to a conventional source isolation valve 88. The conventional source isolation valve 88 is connected to a source 90 of a standard gas having known concentrations of 13C02 and 12C02. The source can be a conventional tank filled with standard gas. The tank is equipped with conventional pressure regulating devices adapted to provide the standard gas under a preselected pressure, desirably about 2 psi (about 14 KPa). An additional gate of the multiple gate valve 84 is connected to a sample valve 92, which in turn is connected to the standard manifold 74. Yet another manifold valve gate 84 is connected through a bypass valve. needle 94, with a node 96, which in turn is connected to the main evacuation valve 80, and to a side of a pump isolation valve 98. The pump isolation valve is connected to the vacuum manifold 74. The outlet of the vacuum pump is connected to the waste. The gate 34b of the second chamber 32b is connected to a generally similar subsystem 176, which includes the pressure sensor 178, the main evacuation valve 180, the injection valve 182, the multiple gate valve or air lock 184, the calibration valve 186, sample valve 192, needle valve 194, and pump isolation valve 198 connected to vacuum manifold 72. However, calibration valve of local system 176 is connected to standard manifold 74 Also, the sample valve 192 of the subsystem 176 is connected to a needle inlet valve 200. The needle inlet valve is connected in turn with a small sharp hypodermic needle 204. The needles of the different local subsystems 176, 214, and 216 are mounted on a common actuator 212 to move relative to the frame 10. To allow movement of the needles, each needle is connected by a flexible capillary tube 202 to the associated inlet valve 200. A container of sample 206 is mounted on frame 10 of the apparatus. The sample container 206 has a generally cylindrical receptacle 208 adapted to receive a container containing a gas sample, such as a breath sample to be analyzed. The sample containers of the different subsystems can be formed as portions of a turntable or other conveyor for moving containers. The container 210 may be a respiration collection device of the type described in U.S. Patent No. 5,361,772, the disclosure of which is incorporated herein by reference. As further disclosed in the '772 patent, this container includes members defining a chamber and a pierceable septum formed as part of an end wall of the container. The cell 32c (Figure 1) is connected to a local subsystem 204 identical to the subsystem 176, while the cell 32d (Figure 1) is connected to an additional local subsystem 216, also identical to the subsystem 176. All the valves are linked to the computer 54 (Figure 1) by means of conventional control interfaces incorporated in the valves and / or in the computer, such that the computer can order each valve to open or close. Also, the pressure sensors 78, 178, and the corresponding pressure sensors of the subsystems 214 and 216, are connected to the computer 54 through other conventional interconnection equipment, in such a way that the computer can receive the data from the pressure sensors. In a process according to one embodiment of the invention, the system is evacuated by the vacuum pump 70, and purged with the standard gas from the source 90. In the purge process, the computer can drive different valves in sequence, to connect all the portions of the system to the vacuum pump and to the standard gas source. After the purge, the system initiates the cyclic operations. Each cycle includes the step of evacuating the cameras and pressure sensors; loads the cameras and pressure sensors with gases, and adjust the pressure inside the different chambers to a previously selected pressure. In the evacuation stage of each cycle, the main evacuation valves 80, 180, and the pump isolation valves 98, 198 are opened, while all other valves are closed, such that all the chambers 32a-32d they are evacuated in a simultaneous way. This operation continues until the pressures in the chambers fall below a previously selected evacuation pressure, desirably about 0.3 Torr. The pressure detected by the sensor 78 associated with the first cell 32a can be used as representative of all chamber pressures in this stage. When it falls below the previously selected evacuation pressure, the evacuation stage is completed. Next, gas is charged in all the cells simultaneously. The valves 86 and 88 are operated to connect the standard source 90 to the injection valve 82 of the first subsystem. The actuators 212 move all the needles 204 of the subsystems 176, 214, and 216, towards the container 206, thereby coupling the pierceable septum of a sample container 210 with each needle 204. The valves 192, 200, and 184, is operated to connect the needle 204 of the system 176 to the injection valve 182, while the corresponding valves of the systems 214 and 216 are also operated in the same manner. Accordingly, the injection valves 82, 182, and similar valves of the systems 214 and 216 are connected to the gas sources to be administered in the chambers 32. Then the computer drives each of the injection valves repeatedly for an interval of impulse previously selected on each repetition. After each repetition, the computer acquires the signal from the associated pressure sensor. If the pressure indicated by the sensor indicates that a particular subsystem exceeds a previously selected load pressure, the computer system terminates the cyclic operation of the injection valve of that subsystem. In this operation, the injection valves of the different subsystems are treated in an independent manner. The repeated cycling of one subsystem may end before the others. In the next step, the multiple gate valves 84, 184 of the different subsystems are operated to connect each injection valve 82, 182 through the associated needle bypass valve 94, 194 and the pump isolation valve 98, 188, with the vacuum pump 70. In this stage of the operation, the main evacuation valves 80, 180 are closed. The system again repetitively drives the injection valve 82, 182 of each subsystem, while continuously reading the signal from the associated pressure sensor of each subsystem. When the pressure indicated by the sensor of a particular subsystem reaches a desired set point pressure, the cyclic operation of the injection valve is terminated. Because the needle bypass valves 94, 194 introduce a relatively high resistance to flow, each pulse of the injection valves produces only a small change in pressure inside the associated chamber 32. Therefore, this stage serves as Fine adjustment of the pressures in the different chambers. At this point, inside the chamber is at the set point pressure appropriate to be tested as discussed below. In a variant of this cyclic process, the charging step is replaced by a reference gas charging step. In the reference gas loading step, the standard gas source 90 is connected through the multi-gate valve 84 and the sample valve 92 of the subsystem of the first chamber, with the manifold 74, and is also connected by means of the multiple gate valve 84 with the injection valve 82 of the subsystem of the first chamber. At the same time, the calibration valve 186 and the multi-gate valve 24 are actuated to connect the manifold 74 to the injection valve 182 of each subsystem 176, 214, and 216. The injection valves 82, 182 are kept open, while the calibration valve 86 of the subsystem of the first chamber is repeatedly driven. The computer 54 monitors the pressure in the first chamber by monitoring the reading from the sensor 78. When this reaches the previously determined charging pressure, the charging step ends. After the sample gas loading step, pressures inside the chambers are adjusted by evacuation through the needle bypass valves 94, 194, as discussed above. This sample gas loading step can be used during a reference cycle, as discussed further below. The instrument can be operated in alternating reference and sample cycles. In each reference cycle, all the chambers 32 are filled with standard gas from the source 90, and are adjusted to the previously established point pressure selected in the manner described above. During each sampling cycle, the chamber 32a is filled with the standard gas, while each of the chambers 32b, 32c, and 32d receives a sample of a different unknown gas. When the gases are samples collected from a medical patient, the unknown gases supplied to the chambers 32b, 32c, and 32d, may be samples collected from the same patient at different times. The unknown gases may be breathing samples collected from the patient before a test compound labeled with 13C is administered, in a first time after that administration, and a second time after administration. The excitation and detection units 42 supply radio frequency energy to the coils 40, thereby converting the gas in each chamber into a plasma. The computer commands the laser device 12 to provide the beam 16 with light at the first wavelength corresponding to the transition energy of 13C02, modulated at a first modulation frequency, desirably from about 50 to about 100 Hz, and commands the laser device 14 which provides beam 18 with light at the second wavelength corresponding to the transition energy of 12C02, modulated at a second modulation frequency, desirably from about 100 to about 200 Hz. Preferably, the modulation frequencies are not integral multiples of each other. Each unit 42 detects the electrical impedance of the plasma in the associated camera 32, and provides a signal representing this impedance to the computer 54 through the signal processor 50. All the units 42 are operated to detect the signals associated with all the cameras 32a-32d in a simultaneous manner. The impedance signal for each chamber will include a first component of magnitude S12 at the first modulation frequency, which represents the opto-gallagic effect of the light at the first wavelength, and a second component of a magnitude S12 at the second modulation frequency , which represents the opto-galvanic effect of light at the second wavelength. Light at the first wavelength interacts with 13C02, but does not interact in a substantial way with 12C02. The magnitude of the first signal S13A for the camera 32a is given by: S13A = P13AM13AW13A <; D where: P13A is the partial pressure or molecular concentration of 13C02 inside chamber 32a; 13A is the energy of the beam at the first wavelength, and consequently, the energy in the first beam 16 from the laser device 12; M13A is a proportionality constant that depends on factors such as the magnitude of the opto-galvanic effect for the particular transition associated with the first wavelength, the configuration of the camera 32a, and the sensitivity of the detector on the unit 42a, associated with the camera 32a. The proportionality constant M13A also depends, to some degree, on the proportion of 13C02 in the excited state inside the chamber 32a, which in turn depends on the excitation energy supplied to the coil 40a, and on the coil configuration .

In a similar way, the magnitude of the second signal for the camera 32b is given by: S12A = P12AM12AW12A (2) wherein: P12A is the partial pressure or molecular concentration of 12C02 inside chamber 32a; W12A is ^ a beam energy at the second wavelength, and consequently, the energy in the second beam 18 from the laser device 14; M12A is a proportionality constant that depends on factors such as the magnitude of the opto-galvanic effect for the particular transition associated with the second wavelength, the configuration of the camera 32a, and the sensitivity of the detector in the unit 42a, associated with the camera 32a. The proportionality constant M12A also depends, to some degree, on the proportion of 12C02 in the excited state inside the chamber 32a, which in turn depends on the excitation energy supplied to the coil 40a, and the configuration of the coil. For typical samples encountered during the use of the instrument, the concentration of 12C02, P 2A 'is several times greater than the concentration of 13C02, pi3A- Therefore, to provide S13A and S12A signals of a comparable magnitude, the beam energy W12A at the second wavelength associated with C02 must be less than beam energy 13A at the first wavelength associated with C02. The configuration of the optical components discussed above, which attenuates the beam from the second laser device to a greater extent than the beam from the first laser device, provides the desired energy ratio in the combined beam. Combining equations (1) and (2), the R13 / 12A ratio of 13C to 12C in the gas inside chamber 32a is given by: 13A = S13AM12AW12A R13 / 12A - 12A S 12AM13AW13A The reconfiguration of equation (3) gives: 1 = S13AW12A KA (4) R13 / 12AS12AW13A Where KA is an additional proportionality constant equal to the quotient of M12A and M13A. The same equations are applied with respect to each of the other cameras 32b, 32c, and 32d, sub-index A being replaced to indicate the corresponding variables that apply to cameras b, c, and d. Because the combined beam of light passes through all the chambers simultaneously, and because the net absorption inside each chamber is negligible compared to the energy in the beam, all chambers receive substantially the same proportion of optical energy at the first and second wavelengths. Therefore: W 12 __ W12A W 12B = W12C = W12D Equations 4 and 5 give the relation: S13AW1 KA = S13BW12 KB = S13CW12 K = S13DW12 KD R13 / 12AS12AW13 R13 / 12BS12BW13 R13 / 12CS12CW13 R13 / 12DS12DW13 (6) Dividing between 12, and reconfiguring the W13 terms R13 / 12B = S13BS12A __ _ ^ B X) R13 / 12A S13AS12B KA R13 / 12C = S13CS12A = _ ^ C (8) R13 / 12A S13AS12C KA R13 / 12D = S13DS12A = _ ^ D (g) R13 / 12A S13AS12D KA For each standard cycle, all the chambers are filled with the standard gas, and therefore, the left side of each of equations 7, 8, and 9 is the unit.

Therefore, each of the proportions KR Kr Kr KA KA KA they can be determined from the signals observed in a calibration cycle. When measurements are taken on a sample cycle, a standard gas is used in chamber 32a, and therefore R13 / 12A is the known ratio R3.3 / 12S 'where the subscript s indicates the standard gas. The ratio R13 / 12 ^ e 13c to 12c For the unknown sample in each chamber 32b, 32c, and 32d, it can be deduced from the known ratio R13 12S '^ as observed signals S, and the proportions of the calibration factors K determined on a calibration cycle. The computer 54 performs the calculations specified by the above equations, and provides the results through the output device 60. With the understanding that the K calibration factors remain constant for the different cameras, the results do not depend on the levels of radiation energy 12 and 13. Put another way, the radio-electric calculation of equations 7 to 9 adjusts the value of each unknown analyte, based on the results obtained for the reference analyte known in chamber 32a. That is, the calculation requested by equations 7 through 9 involves the calculation of a "double ratio" between (i) the proportion of the response magnitudes for the two wavelengths for the unknown sample in a particular cell, and (ii) ) the ratio of the response quantities for the two wavelengths for the standard sample cell 32a. For example, in equation 9 the proportion S13DS12A S13AS12D is the double proportion between (i) the proportion of the response magnitudes for the unknown sample S13D / S12D, and (ii) the proportion of the response quantities for the. sample standard S13A / S12A. The effects caused by the changes in the applied radiation and other variations in the system, cancel each other in the calculation of the double proportion. The standard gas acts as an internal calibration standard during each sampling cycle, and the results of this internal calibration are incorporated in the double proportion. Preferably, the laser devices used in the system are stabilized to minimize the variation in the wavelengths of the light emitted by the laser devices. This variation in wavelength may occur, for example, as the temperatures of the laser devices change. The variation in the wavelength of the light emitted by a laser device will alter the opto-galvanic effect of the light from this laser device. The farther the wavelength of light is from the exact transition energy, the lower the opto-galvanic effect. For a first approximation, this effect is corrected by the system in a manner very similar to the variations in the energy level of the laser devices. Therefore, to the extent that the variation affects all K parameters in equations 7 through 9 in a similar way, the variation will not affect the calculated value of the isotopic ratio in the unknown sample. However, it is still desirable to maintain each laser device at a substantially constant wavelength. The decarcating tubes of the laser devices must be kept under controlled temperatures. For example, the apparatus may incorporate a container for containing a fluid, preferably a liquid, and a temperature controller to maintain the fluid at a constant temperature, and circulate the fluid within the container. The discharge tubes of the laser devices can be mounted inside the container, and can be bathed in the fluid. Also, the wavelengths of the laser devices can be stabilized by means of a feedback control configuration where the opto-galvanic effect caused by the radiation is monitored from each laser device in the standard gas, and each laser device is tune in response to the results of this monitoring, to maintain this optogal effect at a constant level.

Numerous variations and combinations of the features described above may be used without departing from the present invention. For example, although the use of a single beam path as described above is preferred, so that radiation is directed through all the samples in order, other optical configurations could be used to direct the light from a source through plural samples in a simultaneous manner. For example, optical elements, such as couplers or beam splitters, which direct light from one or more beams of the laser device on plural paths to several samples simultaneously can be used. Provided that these optical elements maintain the fixed deviation portions of the optical energy applied to each sample, the composition of each sample can be determined in a manner similar to that described above, using a reference sample simultaneously exposed with the others samples, as an internal calibration standard. Also, the apparatuses and methods according to the present invention can be used for other analyzes than the analysis of the C02 isotopic content discussed above. The trajectory of the beam through the sample vessels does not need to be a straight line; the trajectory can be doubled if the appropriate optical components are provided to deflect the beam. Since these and other variations and combinations of the features discussed above can be used, the above description of the preferred embodiments should be taken by way of illustration rather than by way of limitation of the invention, as defined in the claims.

Claims (26)

1. A method for analyzing an analyte, which comprises the steps of: (a) maintaining a plurality of samples separated from the analyte, including at least one reference sample having a known composition, and at least one unknown sample; (b) directing radiation that includes first and second wavelengths corresponding to the transition energies of a first species and a second species, respectively, that may be present in the analyte, through the plural samples, in such a way that the radiation passes from a common source of radiation through all the samples in a simultaneous manner, whereby, all samples will be exposed to radiation of substantially the same spectral composition; and (c) monitoring the response of the samples to the radiation, to determine a response for each of the wavelengths; characterized by the step of: (d) calculating a double proportion equal to a ratio between (i) a response ratio of the unknown sample between the magnitudes of the responses of the unknown sample to the first and second wavelengths for each sample unknown, and (ii) a standard sample response ratio between the magnitudes of the standard sample responses to the first and second wavelengths for that standard sample.

2. A method as claimed in the claim 1, wherein the step of directing the radiation includes the step of directing one or more radiation beams through all the samples in an order from upstream to downstream.

3. A method as claimed in the claim 2, wherein the step of directing radiation includes the step of providing all wavelengths in a single beam of radiation, and directing this single beam of radiation through all the samples in the order of upstream to downstream .

4. A method as claimed in the claim 3, wherein the step of providing all wavelengths in a single beam includes the step of combining a plurality of beams of different spectral composition, from a plurality of sources, to form the single beam.

A method as claimed in claim 2, wherein the step of directing one or more radiation beams includes the step of reflecting each beam, such that each beam passes: through the samples in a current order down to upstream, as well as in an order from upstream to downstream.

6. A method as claimed in any of claim 2 or claim 3 or claim 4 or claim 5, wherein the step of monitoring the responses is performed by monitoring an induced effect in each of the samples.

7. A method as claimed in the claim 6, wherein the step of maintaining the samples includes the step of keeping the samples in a condition where each species is present in an excited state, the wavelengths corresponding to the transition energies of the species in those excited states.

8. A method as claimed in the claim 7, wherein the step of maintaining the samples includes the step of maintaining each sample in a plasma, and the step of monitoring the induced effect includes monitoring the electrical impedance of each plasma, in order to detect in this way the opto-galvanic effect induced by the radiation .

9. A method for analyzing an analyte, which comprises the steps of: (a) generating a first beam of radiation having a first wavelength corresponding to a transition energy of a first species in the analyte, and a second beam of radiation having a second wavelength corresponding to a transition energy of a second species in the analyte; (b) combining the first and second beams to form a beam composed of radiation, - (c) directing the composite beam of radiation through the analyte, and (d) monitoring an induced effect on the analyte caused by the radiation, to determine an answer of the analyte at each wavelength; characterized in that the step of combining the radiation in the second beam is attenuated to a greater degree than the radiation in the first beam.

A method as claimed in claim 10, wherein the step of generating the first radiation beam is performed by the operation of a laser device containing the first species, and the step of generating the second beam of; Radiation is done by, the operation of a laser device that contains the second species.

A method as claimed in claim 10, wherein the step of generating the first radiation beam is performed by the operation of a laser device containing the first species, and the step of generating the second radiation beam is performed by the operation of a laser device that contains the second species.

12. A method as claimed in claim 11, wherein the first and second species are carbon dioxide species having different isotopic composition.

13. A method as claimed in claim 12, wherein the first species is 13C02, and the second species is 12C02. 1 .

A method as claimed in the claim 9, wherein the step of directing the composite beam through the analyte is performed by the direction of the composite beam through plural samples of analyte, by the direction of the composite beam through plural samples of the analyte, by the direction of the beam composed through all the samples in an order from upstream to downstream.

15. A method as claimed in claim 9, wherein the step of combining the beams is performed by applying the beams to a partially reflecting mirror, such that the second beam is transmitted through the mirror, while The first beam is reflected from the mirror.

16. An analytical apparatus for determining the composition of an analyte, which comprises: (a) an irradiation element for providing radiation at first and second wavelengths corresponding to the transition energies of first and second species in an analyte, and directing this radiation along the trajectory, in such a way that the radiation passes through the analyte; and (b) a detection element for monitoring an induced effect in the analyte, in order to monitor the response of the analyte to the radiation at each wavelength, characterized in that the radiation element includes an element to produce a first beam that it includes radiation at the first wavelength, and a second beam that includes radiation at the second wavelength, and combining the optics to combine the first and second radiation beams, to form a composite beam, adapting the combination optics to attenuate the second beam to a greater degree than the first beam.

17. An apparatus as claimed in the claim 16, which further comprises: (a) a framework; (b) a plurality of cameras mounted in the frame along a path, each camera having upstream and downstream end walls that give upstream and downstream directions along the path; and (c) an element for charging the analyte in the chambers.

18. An apparatus as claimed in the claim 17, which further comprises an excitation element for maintaining an analyte in each chamber in an excited condition, such that one or more species of the analyte are present in the excited states.

19. An apparatus as claimed in claim 18, wherein the excitation element includes an element for maintaining the analyte in each chamber as a plasma.

20. An apparatus as claimed in claim 19, wherein the sensing element includes an element for detecting the electrical impedance of the plasma in each chamber.

21. An apparatus as claimed in claim 17, wherein the loading element includes an element for charging separate analytes in a plurality of the chambers in a simultaneous manner.

22. An apparatus as claimed in claim 21, wherein the loading element includes a source of a standard analyte, and an element for charging the standard in one of the chambers, and charging the analytes of unknown composition in other of the cameras.

23. An apparatus as claimed in claim 17, wherein the path has upstream and downstream ends, the chambers being configured in an order of upstream to downstream on the path, and wherein the irradiation element includes an element to direct the composite beam along the path from the upstream end to the downstream end, and a mirror to reflect the composite beam from the downstream end to the upstream end.

24. An apparatus as claimed in claim 16, wherein the combining optics includes a partially reflecting mirror, and an element for directing the first and second beams on the partially reflecting mirror, such that the second beam is transmitted to through the mirror, while the first beam is reflected from the mirror.

25. An apparatus as claimed in the claim 24, wherein the element for providing the beam includes a first laser device that includes the first species, to provide the first beam, and a second laser device that includes the first species to provide the second beam.

26. An apparatus as claimed in the claim 25, wherein the first and second laser devices are C02 laser devices containing C02 of different isotopic compositions.