CA2248321C - Condensate colorimetric nitrogen oxide analyzer - Google Patents

Condensate colorimetric nitrogen oxide analyzer Download PDF

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Publication number
CA2248321C
CA2248321C CA002248321A CA2248321A CA2248321C CA 2248321 C CA2248321 C CA 2248321C CA 002248321 A CA002248321 A CA 002248321A CA 2248321 A CA2248321 A CA 2248321A CA 2248321 C CA2248321 C CA 2248321C
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exhalate
inlet
conduit
outlet
reagent
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CA2248321A1 (en
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Benjamin M. Gaston, Iv
John F. Hunt
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Nitromed Inc
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61BDIAGNOSIS; SURGERY; IDENTIFICATION
    • A61B5/00Measuring for diagnostic purposes; Identification of persons
    • A61B5/08Detecting, measuring or recording devices for evaluating the respiratory organs
    • A61B5/083Measuring rate of metabolism by using breath test, e.g. measuring rate of oxygen consumption

Abstract

Disclosed is a device for determination of the content of higher oxides of nitrogen in exhaled breath condensate which comprises a conduit (7) having an exhaling condensing portion with an inlet and an outlet (the inlet can be configured to fit with a mechanical respirator, or for direct use by a patient, an inlet assembly providing one-way ingress (2) of ambient atmosphere so the device can be associated with the inlet of the conduit exhaling condensing portion); a coolant jacket (6) coaxially surrounding said exhaling condensing portion; a gas release port; and, in an enclosed fluid communication with said conduit exhaling condensing portion outlet, a sample assay assembly (9) comprising the following: (a) a translucent analysis chamber attached to the outlet to receive condensate fluid and having a reagent port;
(b) a reagent chamber in enclosed fluid communication with the reagent entry port; and (c) a pliable element connecting the oulet and the analysis chamber and forming a portion of the reagent chamber such that flexion of the pliable element closes communication between the owlet and the analysis chamber and contracts the reagent chamber so as to deliver to the analysis chamber controlled amounts of condensate and reagent. The device is disposable and-inexpensive, and is used to collect human exhaling for colorimetric assay of liquid and gas phase nitrogen oxides to assist in evaluation of airway inflammation.

Description

CONDENSATE COLORIMETRIC NITROGEN OXII)E ANALYZER

BACKGROUND OF THE INVENTION
Conventional ~csescment of severity of air~vay diseases, including actllm~
consists of measures of pulmonary mPch~nics which are effort dependent and thus not suitable for many pediatric p~ti~nts~ mecll~nir~lly ventilated patients, or patients with neuromuscular disease. See Crapo R., Pulmonary-function testing. N. Engl.
J. Med., 331; 1:25-30; 1994; and American Thoracic Society/European Respiratory Society: Respiratory m~ch~ni-~s in infants - physiologic evaluation in health and disease,Am. Rev. Respir.Dis., 1993; 147:474-96. CurrentlythePeakExpiratory flow meter is the only method commonly available for home use by ~ ics to monitor the state of their ain,vay disease. This test suffers from being an insensitive in-lic~tor of airway infl~mm~tion, and is effort dependent. Other physiologic parameters require e~l,ensi~e m~rhines and experienced operators to obtain reproducible results. Klein R., Fritz G. Yeung A., McQuade E., Mansell A., Spilolllc~ic Pa~ lls in Childhood Asthma: Peak Flow Col"paled with Other Indices, Pediatr. Pulmonol., 1995, 20:372-379.

Endogenous production of nitrogen oxide in the human airway has been shown to be inc.~ased in patients with asthma and other infl~mm~tory lung ~lice~ces.
Gaston B., Drazen J., Chee, C., et al. Expired nitric oxide concellt~aLions are elevated in patients with reactive airways disease. Endothelium. 1993; 1:S87;
Gustafsson, I.E., Leone A.M., Persson M.G., Wiklund N.P., Moncada S., Endogenous nitric oxide is present in the exhaled air of rabbits, guinea pigs and humans. 1991; 181; 2:852-857. However, measulc-llenl of gas phase nitric oxide CA 0224832l l998-09-04 WO 97/35519 PCTrUS97/04848 (nitrogen monoxide) per se in the parts per billion range found in the airway requires the use of mass spectrometry or chemiluminescent technology, which are cumbersome, exE,ensive and not reasonably suited for home use.

Moreover, measurement of aqueous phase higher oxides of nitrogen compounds such as S-nitrosothiols, nitrite and nitrate, which are not nitrogen monoxide but which are likewise elevated in concentrations in subjects with infl~mm~tory lung disease, has required that patients be hospitalized for invasive airway sampling procedures, including airway intubation and bronchoalveolar lavage.
Gaston, B., Reilly J., Drazen J., e~ al., Endogenous nitrogen oxides and bronchodilator S-nitrosothiols in human airways. Proc. Natl. Acad. Sci., 1993;
90: 10957-10961 .

SIJMMARY OF THE INVENTION
The invention provides a device for determination of the content of higher oxides of nitrogen in exhaled breath con-~enc~te which comprises a conduit having an exhalate condencine portion with an inlet and an outlet (the inlet canbe configured to fit with a mech~nir~l respirator or, for direct use by the patient, an inlet assembly providing one-way ingress of ambient atmosphere to the device can be associated with the inlet of the conduit exhalate condensing portion); a coolant jacket coaxially surrounding said e~h~l~te condensing portion; a gas release port; and, in enclosed fluid communication with said conduit exh~i~t~ con~Pn~ing portion outlet, a sample assay assembly comprising (i) a translucent analysis chamber ~ttar~eA to the outlet to receive condensate fluid and having a reagent entry port, (ii) a reagent ch~mber in enclosed fluid communication with the reagent entry port and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber such that flexion of the pliable element closes commllnir~tion between the outlet and the analysis chamber and con~lacl~ the reagent chamber soas to deliver to the analysis chamber controlled amounts of con-len~te and reagent.
The device is disposable and inexpensive, and is used to collect human e~h~l~te for colorimetric assay of liquid and gas phase nitrogen oxides to assist in evaluation of airway infl~mm~tion.

WO97/35519 - PCTrUS97/04848 In a ~rcfellcd embodiment the device further co-l~ iscs an inlet assembly providing one-way ingress of ambient atmosphere to the e~rh~ql~te condensing conduit inlet. This is ideal for the patient who is breathing directly into the device, rather than the situation where the device is comlec~ed to a mechanical respirator.

In a preferred embodiment the device further comprises, between the inlet assembly and the exh~lqte condensing conduit portion, a filter capable of removing particulate matter from exhaled air which pas~s therethrough.

In another plcfel~d embo-liment, the device further co-np-ises a coolant material in the coolant jacket.

In another p,cfe--ed embodiment, the device further co-npri~s a reagent composition capable of ~letecting a nitrite or nitrate in an e~h~lqtion condensate sample.

In another ple~cllcd embodiment7 the device further COIll~liSCS a reagent-permeable membrane in the reagent entry port.

In another plcfe~-cd embodiment, the device further col..~rises a rupturable membrane in the reagent entry port.

In another prererlcd embodimetlt, the inlet assembly further includes therein a filter capable of removing ambient nitrogen oxides from e~h~lq-fP passing therethrough.

In another plefcl~cd embodiment the exhalate con-l~ncing conduit portion comprises a single lumen conduit having a lumen divqm~tP-r sufficient to cause substqntiqlly no resicPn~e to the flow of e~h,qvl~t~ thclethlollgh.

~ In another preferred embodiment the e~h~ql~te condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diq-met~r sufficient to cause subst~nti~lly no resict~nce to the flow of elrh~l~te therethrough.

W O 97135519 rcTrusg7/o48 The purpose of this invention is to condense exhaled lung gas and vapor for aqueous phase nitrogen oxide analyses. This will assist in the evaluation of thehuman airways' production of nitrogen oxides in both gas and several liquid phase states, including nitrite and nitrate, with potential for multiple other studies to help delineate the lung's redox enviroml.ent and the airways' degree of infl~mm~tion.The investigations which have resulted in the present invention in~lic~te that easily measured liquid phase exhaled nitrogen oxides, nitrite and nitrate, are likewiseelevated in ~cthm~tic subjects during periods of infl~mm~tion (see abstract enclosed, unpublished data). Utility in the clinical setting for inexpensive and domiciliary evaluation of airway infl~mm~tion in various disease states, including ~thm~, is the primary purpose of the device of the present invention and its utility has been proven, inter alia, as demon~tr~ted herein.

The invention is comprised of cold tolerant materials and consists of two tubing units, one inside the other. Surrounding the inner tube or set of tubes, and contained by the outer tube, is a chemical substance which has a high specific heat, and therefore, once frozen, can ~n~int~in rr~e~ing temperatures for an extended period of time. The disposable unit can be frozen in a standard size home freezer and then conn~cted together with collection and analysis instruments as a compact integral unit.

Attached to the proximal portion of the unit is a port through which the subject breathes. This consists of two one-way valves which direct atmo~heric air or selected gases to the patient's lungs during inspiration, and channel ~Yh~l~d gas down a condensing tube. Gas moves in only one direction through the condensing apparatus. Inserted between the breathing port's mouthpiece and the condensing chamber is a microporous filter which traps all small particles(such as saliva or sputum), is illlperllleable to liquids, but allows gas and vaporized fluids of less than 0.3 microns in diameter to pass. This acts as a saliva trap and may also act as a filter for the larger fluid particles which may be aerosolized in the larger airways.

W O 97/35519 PCT~US97/04848 The distal end of the condensing chamber tube(s) is ~tt~fhed to a collecting apparatus which utilizes gravity to trap condensed fluid. At the botLom of this trap is a clear plastic analyzing chamber in which the sample is sequentially warmed,sealed, reacted with col~ ..el-ic reagents and analyzed spectrophotometrically by the non-disposable colorimeter.

The patient breathes comfortably in and out through the mouthpiece. Lung fluid vapor collects on the inner surface of the inner tube(s) of the condensingapparatus starting immediately. Gravity carries the larger droplets down the tube, these droplets recruiting other small droplets on their trip to the collecting vial distally. Alternatively, after a fixed period of tidal breathing, the condensed fluid can be e~ ,ssed down the inner tube with a device similar to a syringe plunger.
Aqueous phase nitrite and nitrate can be measured by standard colorimetric assays, and can be reasonably quantified by simple tests pe,ror",ed by patient~ in theirhomes.

Brief D~ )tion of the D~ a~

Figure 1 is a side elevation view, partially in section, of the device of the invention.

Figure 2A is a side sectional elevation view of a preferred embodiment of the sample assay assembly of the device with the non-disposable analyzer, and includes a top view of the conne~ pincer valve.

Figure 2B is a top view of the pincer which flexes the pliable element of the sample assay assembly.

Figure 3 is a histogram showing the results of the nitrite assay pelrolllled as described in E~xample 2.
.

Figure 4 is a histogram showing the results of the nitrate assay pel rorllled as described in Example 2.

Detailed Description of the Invention The device of the invention has several advantages, including:

1. This is the only device which measures airway infl~mm~tion with a simple home assay, without invasive procedures or expensive and cumbersome testing equipment.
2. Ease of transportability and use of the apparatus.
3. Clinical application for individual patients in their own home or in the clinic or hospital setting to test for or monitor status of asthma or otner causes of airway infl~mm~tion. May be used to predict disease exace~ ions, allowing time for successful early intervention.
4. Clinical applicability in the setting of mech~nir~lly ventilated patient to easily monitor airway infl~mm~tion.
5. Clinical applicability for measuring airway infl~mm~tion in many children too young to cooperate with standard pulmonary function testing, but who can cooperate with tidal breathing through a tube. Similar potential in p~tientc with neurom~crul~r disease in whom standard tests are uninterpretable.

The device can be made with easily available materials. The cooling substance in the condencing ch~mber can be water, or a viscous substance with a high specific heat. The device can also be made, as described above, without refrigerant, utili7ing a gas/membrane or gas/aqueous reaction compartment.

Referring now to Figure 1, mouthpiece 1 is formed as an integral element with low resistance one-way valve 2 allowing inspiration of air into the device of the invention resulting from inhalation through it by the user. Low resict~nre one-way valve 2 optionally contains potassium perm~ng~n~te and a charcoal filter to remove ambient nitrogen oxides. Mouthpiece 1 and low reSict~nre one-way valve 2 are further integral with low resist~nr,e one-way valve 3 to direct all expired gas from the user into the rem~ining elements of the device. Integral with and downstreamof low resi~t~nre one-way valve 3 is low resict~nre filter 4, cont~ining elements which removes all particulates greater than .03 ~M in diameter.

Connected downstream to low resistance filter 4 is condensing çh~mber 5 which consists of either a single, long inert plastic tube of sufficient diameter to be of low resistance to air expired into the device, or a bundle of shorter, parallel, inert plastic tubes of suf~lcient number to provide ~ninim~l reci~t~nre to airflow.
Condensing chamber 5 is in continuity with, i.e., receives air arising from filter 4.
Coolant chamber or jacket 6 contains an aqueous solution of sufficient osmolality to cause freezing point depression to just above the minimum temperature of the average household freezer, e.g., in a range of :~: 15~F, preferably below 0~F.

Condensing chamber 5 is completely sealed from exposure to coolant chamber 6. Essentially coolant chamber 6 coaxially surrounds condensing chamber 5 so as to control the temperature within con~encing chamber 6. Condensing chamber 6 is continuous with a conduit that divides to condensate delivery conduit 7 and exhaust port 8. A reagent i,l,pr~dnaled mesh, such as filter paper, can optionally be situated in conduit 7 for reagent combination with condensate prior to their arrival in sample analysis assembly 9. F~h~uct port 8 co",~,i~s a one-way,low-resict~nre valve through which air, in which the condensate has been analyzed, is expelled from the device of the invention.

Sample analysis assembly 9 is comle~led to the outlet of condel ~ing chamber 5 through condensate delivery conduit 7 and is desclil,cd in detail with ~ ert,nce to Figure 2A.

Analyzer 10 which is not a component of the disposable device of the invention, includes a wavelength-filtered light source and photometer bulb, for measuring the specific absorbance of reacted reagent in sample assay assembly 8,and a col"l)uler both for calculating and displaying the co"~_sl,onding nitrogen oxide W O97/35519 PCT~US97/04848 content of the sample in assembly 8 and for calibrating the absorbance/col1ce~ alion relationship based on standard samples is shown as an association of conventional elements in essentially~'block form. Elements 1 through 9 of the device of the present invention form a disposable unit which is stored under refrigeration, preferably frozen, until used. In contrast, analyzer 10 is not disposable and ism~int~in~d at room temperature.

The disposable unit comprising elements 1-9 is removed from frozen storage and placed as a unit in connecting housing 11, which is ~ hed to analyzer 10 in such a way that analysis assembly 9 fits snugly into housing 11. "Housing" serves to hold the disposable and non-disposable units together at the approyl iale orientation, and to prevent the subject user's hand from becoming affected by the cold of the disposable device while holding the ~;~pa~dlus during gas sampling.
Sample analysis assembly 9 is surrounded by a thermost~tic~lly controlled heating coil 12 ~tt~hed to housing 11 and analyzer 10 which brings the sample and test reagents to 25~C. Heating coil 12 is part of the non-disposable unit.

Referring now to Figure 2A, illustrated in h~tr~ed line is the outlet of condensate delivery conduit 7, which is in enclosed fluid contact with sample assay assembly 9, sample assay assembly 9 colllplises continuous with a round, hard-plastic analysis ch~mber 13, reagent chamber 14 and pliable element 15. Conduit 7 and assembly 9 are brought into juxtaposition and are conn~cted by pliable element 15, which is a soft-plastic component that extends beyond the tli~metPr of conduit 7. At the bottom of analysis chamber 13 is an opening 16 in which is positioned a fluid-permeable membrane or a thin, plastic rupturable membrane.

Reagent(s) for cololhllt~ric nitrogen oxide salt analysis could be a diazo reagent, or such a reagent in conjunction with nitrate redll~t~ce and appropriate cofactors to enable both nitrite and nitrate to be measured simultaneously. Other reactions, such as reduction of nitrite and nitrate to nitric oxide in v~n~lium chloride, followed by reaction with oxyhemoglobin to achieve a colorimetric shift in the Soret band, and including heterolytic cleavage of S-NO bonds with mercuric chloride followed by diazo analysis could also be employed.

W O 97/35519 PCT~US97/04848 When exhaled, the con~lenc~tP of eYh~led air passed into analysis ch~mber 13.
Thereafter, the user squeezes pliable element 15, using pincer 17 (shown in Figure 2). When pincer 17 is closed against pliable element 15, this causes the volume of reagent chamber 14 to declease such that reagent therein is forced upwardly through the permeable or rupturable membrane into analysis chamber 13 to combine and react with the condensate. TherearLer, the reaction product of this çhPmir~l reaction is analyzed by colorimetry using the non-disposable colorimeter which passes a path of light through a fixed path of analysis chamber 13.

Potential modifications to the system include:

1. Attachment of an assembly distal to the inspiratory portion which would contain a fiber paper through which the saliva free exh~l~tç is channelled.
The filter paper would be impregnated with reagents which react with nitrogen oxides and subsequently change color. A patient could breathe through the fîlterpaper for a given period of time and the degree of color change could be measured against a chart or with a small colorimetric device.

2. Direct ~tt~chment to the distal end of the condensing chamber of a filter paper i~ r~gnaled with dried reagents which will react with the aqueous phase nitrogen oxides to yield a change in color of the paper which can easily be measured in a similar fashion to modification #1 above.
3. Direct att~hment of a chamber cont~ining liquid reagents through which the exhalate is bubbled, allowing exhaled nitrogen oxides to directly react with reagents, producing a Illeasulable color change.
4. Att~rhmellt of a ~lehllmi~lifying chamber to the inspiratory portion to eliminate environment~l liquid phase nitrogen oxides.

EXA~PLE 1 Expired Nitric Oxide Durin~ Tidal Br~athinp in Humans WO 97/35519 PCT~US97/04848 Recent insi~ht~ regarding the use of expired nitric oxide (NO-) concell~rdtion measurement as an index of airway infl~mm~tion cannot be applied to children tooyoung to perform vital capacity (VC) maneuvers. We have Ihel~îole developed assays which require only tidal breathing. Sequential colorimetric measurements were made during quiet breathing by reacting dried air expirate (DAE) with Griess reagent under acid conditions. Tidal expired gas NO- production rates (V~,O.) were also measured by chemihlminescence over 5 minutes using a 765 liter sealed plethysmograph. Eight control subjects and 6 patients with asthma and cystic fibrosis (CF) were studied. DAE NO- was detected in all normal subject samples with mean (iSD) accumulation curve slope = 40 ~17 nmoles/liter/breath (p ~ 0.05 compared with air controls). By comparison, the mean [dissolved nitrogen oxides (NO,~)] collected in a precipitation tube (PT) (-80~C) from size-filtered lung water produced by normal subjects during quiet breathing was 0.8 ~tO.1 ~M. Tidal VNo/body surface area (specific VNO. [VNO~SP]) was 0.21 :tO.043 ,ul/min/m~ in normal subjects. Consistent with previous reports based on VC measurements, patients with asthma had higher VNO.SP (0.31 iO.014 ~I/min/m2, p<O.O1) and PT
NOX (16 i4.0 ,uM, p < 0.005). Further, p:~tient~ with CF also had high VNO~SP (0-34 + 0.84 ,ul/m2/min, p<O.005). We report for the first time 1) the detection of NO-in expired air through the use of colorimetric assays; 2) the effort-independentdetection of VNO; and 3) increases in these tidal breath conce~ ions in p~tien with asthma and CF. We speculate that tidal-breathing based t~ niques will have clinical utility both as measures of airway infl~mm~tion applicable to home use and as effort-independent measures of airway disease in the pediatric pulmonary function laboratory.

Exam~le 2 Device Evaluation in ~Iuman Clinical Study Asthma is an inflamm~tory disease which leads to airway obstruction.
Current tests of disease severity are based on measures of ob~ u-;lion, alterations of which are late findings in asthma exacerbations. Recent studies reveal that expression of inducible nitric oxide synthase (iNOS) is increased in the airway W O 97/35519 PCTrUS97/04848 epithelial and infl~mm~tory cells of patients with ~cthm~ The activity of iNOS is known to be reflected in high collcenllalions of nitric oxide (NO ~ ) gas measured in the expired air of ~cthmatir subjects. We studied other aqueous phase NOX in ~cthm~tic aspirate in an effort to develop a simple test for airway infl~mm~tionapplicable to home use.

Subjects 6 normal subjects no smoking history no history of acute, chronic, or recurrent cardiopulmonary, determatologic, gastrointestin~l, or neurologic disease.

13 ~cthm~tic patients 3 or more episodes of reversible bronchosp~m I/E ratio <0.5 FEV,/FVC C0.75 at study entry Methods Saliva free, 0.2 micron particle si~-filtered expired vapor was obtained from subjects breathing quietly through a condensing apparatus.

Nitrite (NO2-) was assayed cololh-~eLlically using Griess reagent.

Nitrate (NO3-) was assayed by chemih~minr-scence after reduction in vanadium chloride.

St~ti~tir~1 Methods Mean NO" concelllla~ions were compared using unpaired t-testing. Results are presented as mean, ~ standard error. P values of C0.05 were considered significant.

Results As shown in Figure 3, nitrite concentrations were significantly higher in ~cthm~tic subjects (1.877 ~LM ~ 0.36) than in controls (0.902 ~M + 0.08) (p < 0.01).

As shown in Figure 4, nitrate concentrations were likewise significantly elevated in ~cthm~t~c subjects (9.48 ,uM i 1.25) relative to controls (5.27 llM
0.51) (p<O.Ol).

Discussion Tests of ain,vay obstruction, such as the peak flow meter, are currently the primary modality for objectively ~Cce~ssing severity of infl~mm~tory airway disease.
These tests have several limitations, including being effort-dependent, and thus are not suitable for many children and neuro-muscularly h~n-lir~pped ind}viduals. They also poorly reflect the degree of underlying infl~mm~tion.

Utilizing recent insights regarding the biochemistry of higher oxides of nitrogen, we have developed a simple, non-invasive assay for airway infl~mmation.
Tests such as this could be performed at home, and may prove to be useful in longitu~lin~l evaluation and management of ~ct~m~ including dosing of anti-infl~mm~tory medication.

Conclusion Aqueous phase nitrogen oxides, measured in condensed exhalate, are significantly elevated in ~Ct~ tic patients, and can be used to distinguish between normal and infl~mPd airways.

Claims (66)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A device for determination of a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;
a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises:
(i) a light transmissive analysis chamber attached to the outlet to receive the exhaled breath condensate, wherein the light transmissive analysis chamber comprises a reagent entry port, (ii) a reagent chamber in communication with the reagent entry port, and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber, wherein flexion of the pliable element closes communication between the outlet and the analysis chamber and contracts the reagent chamber to deliver to the analysis chamber controlled amounts of condensate and reagent.
2. The device of claim 1 which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
3. The device of claim 1 which further comprises a coolant material in the coolant jacket.
4. The device of claim 1 which further comprises a reagent composition for detecting a nitrite or nitrate in the exhaled breath condensate.
5. The device of claim 1, wherein the reagent entry port comprises a reagent-permeable membrane.
6. The device of claim 1, wherein the reagent entry port comprises a rupturable membrane.
7. The device of claim 1 wherein between the inlet and the outlet is an inlet assembly which further comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
8. The device of claim 1 wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
9. The device of claim 1 wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
10. The device of claim 1 which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
11. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;

a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises:
(i) a light transmissive analysis chamber attached to the outlet to receive exhaled breath condensate, wherein the light transmissive analysis chamber comprises a reagent entry port, (ii) a reagent chamber in communication with the reagent entry port, wherein the reagent entry port comprises a reagent-permeable membrane, and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber, wherein flexion of the pliable element closes communication between the outlet and the analysis chamber and contracts the reagent chamber to deliver to the analysis chamber controlled amounts of condensate and reagent.
12. The device of claim 11, which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
13. The device of claim 11, which further comprises a coolant material in the coolant jacket.
14. The device of claim 11, which further comprises a reagent composition for detecting a nitrite or nitrate in the exhaled breath condensate.
15. The device of claim 11, wherein, between the inlet and the outlet, is an inlet assembly which comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
16. The device of claim 11, wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
17. The device of claim 11, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
18. The device of claim 11, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
19. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;
a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises:
(i) a light transmissive analysis chamber attached to the outlet to receive exhaled breath condensate, wherein the light transmissive analysis chamber comprises a reagent entry port, (ii) a reagent chamber in communication with the reagent entry port, wherein the reagent entry port comprises a rupturable membrane, and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber, wherein flexion of the pliable element closes communication between the outlet and the analysis chamber and contracts the reagent chamber to deliver to the analysis chamber controlled amounts of condensate and reagent.
20. The device of claim 19, which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
21. The device of claim 19, which further comprises a coolant material in the coolant jacket.
22. The device of claim 19, which further comprises a reagent composition for detecting a nitrite or nitrate in an exhalation condensate sample.
23. The device of claim 19, wherein between the inlet and the outlet is an inlet assembly which further comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
24. The device of claim 19, wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
25. The device of claim 19, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
26. The device of claim 19, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
27. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet, wherein the inlet comprises a mechanical respirator;

a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises:

(i) a light transmissive analysis chamber attached to the outlet to receive the exhaled breath condensate, wherein the light transmissive analysis chamber comprises a reagent entry port, (ii) a reagent chamber in communication with the reagent entry port, and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber, wherein flexion of the pliable element closes communication between the outlet and the analysis chamber and contracts the reagent chamber to deliver to the analysis chamber controlled amounts of condensate and reagent.
28. The device of claim 27 which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
29. The device of claim 27, which further comprises a coolant material in the coolant jacket.
30. The device of claim 27, which further comprises a reagent composition for detecting a nitrite or nitrate,in the exhaled breath condensate.
31. The device of claim 27, wherein the reagent entry port comprises a reagent-permeable membrane.
32. The device of claim 27, wherein the reagent entry port comprises a rupturable membrane.
33. The device of claim 27, wherein between the inlet and the outlet is an inlet assembly which further comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
34. The device of claim 27, wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate theregthrough.
35. The device of claim 27, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
36. The device of claim 27, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
37. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;
wherein the inlet comprises a dehumidifying chamber;
a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises:

(i) a light transmissive analysis chamber attached to the outlet to receive the exhaled breath condensate, wherein the light transmissive analysis chamber comprises a reagent entry port, (ii) a reagent chamber in communication with the reagent entry port, and (iii) a pliable element connecting the outlet and the analysis chamber and forming a portion of the reagent chamber, wherein flexion of the pliable element closes communication between the outlet and the analysis chamber and contracts the reagent chamber to deliver to the analysis chamber controlled amounts of condensate and reagent.
38. The device of claim 37, which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
39. The device of claim 37, which further comprises a coolant material in the coolant jacket.
40. The device of claim 37, which further comprises a reagent composition for detecting a nitrite or nitrate in the exhaled breath condensate.
41. The device of claim 37, wherein the reagent entry port comprises a reagent-permeable membrane.
42. The device of claim 37, wherein the reagent entry port comprises a rupturable membrane.
43. The device of claim 37, wherein between the inlet and the outlet is an inlet assembly which further comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
44. The device of claim 37 wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate theregthrough.
45. The device of claim 37, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
46. The device of claim 37, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
47. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:
a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;
a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises a filter paper or fiber paper.
48. The device of claim 47, wherein the filter paper or fiber paper comprises a reagent composition for detecting a nitrite or nitrate in the exhaled breath condensate.
49. The device of claim 47, which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
50. The device of claim 47, which further comprises a coolant material in the coolant jacket.
51. The device of claim 47, wherein, between the inlet and the outlet, is an inlet assembly which comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
52. The device of claim 47, wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
53. The device of claim 47, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
54. The device of claim 47, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
55. The device of claim 47, wherein the inlet comprises a mechanical respirator.
56. The device of claim 47, wherein the inlet comprises a dehumidifying chamber.
57. A device for determining a content of higher oxides of nitrogen in exhaled breath condensate comprising:

a conduit for receiving a flow of exhalate, said conduit having an exhalate condensing portion with an inlet, an outlet and a gas release port coupled to the outlet;

a coolant jacket coaxially surrounding said exhalate condensing portion; and a sample assay assembly in communication with said conduit exhalate condensing portion outlet, wherein the sample assay assembly comprises at least one liquid reagent.
58. The device of claim 57, wherein the liquid reagent can detect a nitrite or nitrate in an exhalation condensate sample.
59. The device of claim 57, which further comprises, between the inlet and the exhalate condensing conduit portion, an inlet assembly comprising a filter for removing particulate matter from exhaled air which passes therethrough.
60. The device of claim 57, which further comprises a coolant material in the coolant jacket.
61. The device of claim 57, wherein between the inlet and the outlet is an inlet assembly which further comprises a filter for removing ambient nitrogen oxides from exhalate passing therethrough.
62. The device of claim 57, wherein the exhalate condensing conduit portion comprises a single lumen conduit having a lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
63. The device of claim 57, wherein the exhalate condensing conduit portion comprises a plurality of conduit tubules having a collective lumen diameter sufficient to cause substantially no resistance to the flow of exhalate therethrough.
64. The device of claim 57, which further comprises an inlet assembly between the inlet and the outlet for providing one-way ingress of ambient atmosphere to the exhalate condensing conduit inlet.
65. The device of claim 57, wherein the inlet comprises a mechanical respirator.
66. The device of claim 57, wherein the inlet comprises a dehumidifying chamber.
CA002248321A 1996-03-28 1997-03-25 Condensate colorimetric nitrogen oxide analyzer Expired - Fee Related CA2248321C (en)

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US08/623,643 1996-03-28
US08/623,643 US6033368A (en) 1996-03-28 1996-03-28 Condensate colorimetric nitrogen oxide analyzer
PCT/US1997/004848 WO1997035519A1 (en) 1996-03-28 1997-03-25 Condensate colorimetric nitrogen oxide analyzer

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CA2248321A1 (en) 1997-10-02
EP0897284A1 (en) 1999-02-24
WO1997035519A1 (en) 1997-10-02
AU2346197A (en) 1997-10-17
AU724417B2 (en) 2000-09-21
US6033368A (en) 2000-03-07
US6419634B1 (en) 2002-07-16
EP0897284A4 (en) 1999-05-19

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