What is in your breath

1
What is in your breath

• Terence H. Risby, PhD
• Bloomberg School of Public Health
• Johns Hopkins University
• July 17, 2006
• EnviroHealth Connections
• Summer Institute 2006

2
What are the sources of molecules in breath?

• Any molecule that has a measurable vapor pressure
  can be found in breath
• Breath will contain molecules originating from
  inspiratory air (current or historical exposure)
• Breath will contain endogenously produced
  molecules from normal and abnormal physiology
  that originate from tissues or cells throughout
  the body
• Breath will contain molecules that are directly
  or indirectly derived from foods and beverages
• Composition of breath is an instantaneous product
  of all these processes

3
History of breath analysis

• Water vapor in breath has been used for centuries
  to detect presence of life.
• Classical medicine has used subjective
  impressions of the odors of urine or breath to
  diagnose disease.
• Lavoisier first detected carbon dioxide in breath
  in 1784.
• Earliest modern publications on breath analysis
  date from late 1960s early 1970s

4
Typical concentrations (v/v) of endogenous
molecules found in human breath

• O2, H2O, CO2
• ppm (CH3)2CO, CO, CH4, H2,
• C2H5OH, C6H10S
• ppb HCHO, CH3CHO, C5H10, C5H12
• C2H6, C2H4, NO, CS2, H/Cs
• CH3OH, C2H5OH, COS, CH3SH NH3, CH3NH2,
  (CH3)2S

5
Normal human breath profile
6
Biochemical basis of breath molecules

• H2 carbohydrate metabolism
• C2H5OH gut bacteria
• H/Cs lipid peroxidation/metabolism
• CO heme catabolism
• NO nitric oxide synthase
• C5H12 lipid peroxidation
• C2H6 lipid peroxidation
• C2H6CO decarboxylation of acetoacetate
• NH3 protein metabolism
• CH4 carbohydrate metabolism

• CH3CHO ethanol metabolism
• C5H10 cholesterol biosynthesis
• CH3OH fruit metabolism
• CH3NH2 protein metabolism
• C2H4 lipid peroxidation
• C6H10S garlic
• CH3SH methionine metabolism
• CS2 gut bacteria
• COS gut bacteria
• C2H6S methionine metabolism

7
Method for breath collection or breath sampling
is as important as the method of analysis.

• Breath components will change as a function of
  breathing cycle
• Breath molecules originate from cells throughout
  oral/nasal cavities, the pulmonary system and the
  entire body
• Breath composition will change with breathing
  physiology
• Sampling single breath or multiple breaths

8
Single breath sampling

• Control flow
• Control mouth pressure monitor pressure
  continuously
• Monitor the concentration of carbon dioxide
  continuously

9
Monitoring a single breath in real-time
Critical orifice
CO2 monitor
Monitor
Pressure monitor
Filter
MOUTH
10
Profile of restricted breath
11
Sampling multiple breaths

• Monitor tidal volume of each breath and breathing
  frequency
• Monitor the concentration of carbon dioxide
  continuously, i.e., determine end-tidal and
  steady state concentrations of each breath
• Monitor mouth pressure continuously
• Monitor pulse
• Monitor oxygen saturation
• Sample multiple breaths

12
Sampling tidal breathing
CO2 monitor
Pressure meter
NRBV
Flow meter
Breath
Filter
MOUTH
13
Effects of ventilation on carbon dioxide
14
Most developed field of breath analysis is based
upon metabolites of diagnostic substrates

• Detection of labeled carbon dioxide (C13 or C14)
• Stable isotope mass or optical spectroscopy
• Detection of radioactivity
• Selectivity based upon the diagnostic substrate
  and biological system under investigation
• Knowledge of delivery, reaction and clearance
  rates critical
• Breath sampled at defined time after
  administration of diagnostic substrate

15
Use of C13 labeled substrates

• aminopyrine, caffeine, galactose, methacetin or
  erythromycin liver function
• ketoisocaproate or methionine liver mitochondria
  function
• acetate or glycosyl ureides orocecal transit
  time
• urea H. pylori infection
• triolein fat malabsorption
• glucose insulin resistance
• linoleic acid fatty acid metabolism
• phenylalanine phenylalanine hydrolase activity
• uracil dihydropyrimidine dehydrogenase
  activity

16
FDA approved breath tests

• Breath hydrogen test for carbohydrate metabolism
• Breath nitric oxide test to monitor therapy for
  asthma
• Breath carbon monoxide test for neonate jaundice
• Breath test for diagnosis of H. pylori
• Breath test for heart transplant rejection
• Breath ethanol for intoxication (law enforcement)

17
Oxidative stress status

• Oxidative stress damage to cells, tissues and
  organs caused by reactive oxygen species such as
  02, H202, and OH.
• Reactive Oxygen Species (ROS) initiate or
  exacerbate specific diseases or dysfunctions,
  kill and damage cells.
• Role of ROS in disease and normal aging an
  important and growing field of biomedical
  research.
• Oxidative stress can be quantified through breath
  measurements of biomarkers ethane, ethylene and
  pentane.
• Alternatively, the cellular response to oxidant
  injury can be studied by inducing cellular
  antioxidant defenses and monitoring biomarker
  carbon monoxide.

18
Reactive oxygen species (ROS) are involved in

• diseases of prematurity
• cardiovascular disease
• airway reactivity and pulmonary diseases
• diabetes
• liver disease

• cancer
• Alzheimer's, and Parkinson diseases
• amyotrophic lateral sclerosis
• scleroderma
• infections
• ischemia/reperfusion injuries
• radiation damage

19
Chronic oxidant injury in humans

• Chronic liver or kidney disease
• Smoking
• Effect of antioxidant vitamins
• Feeding studies

20
Breath ethane to monitor vitamin E therapy
21
Maternal cigarette smoking
22
Neonates of mothers that smoke
23
Diet ()
24
Study design
eight weeks
A
eight weeks
three weeks
B
B
eight weeks
C
X
X
Baseline sampling
End of study sampling
25
Change in breath ethane from baseline to end of
study
Diet B
1.5 ppb
0.0 ppb
-1.5 ppb
Diet C
Diet A
26
(No Transcript)
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Oxidative stress and diet restriction
24 month Fisher 344 female rats ad lib 289.710.5
g DR 168.41.9 g

• ad lib 2.31 0.78 pmol/ 100 g min p lt 0.5
• DR 2.32 0.65 pmol/ 100 g min
• ad lib 135 26 pmol / ml CO2 p lt 0.0003
• DR 97 16 pmol / ml CO2

28
Effect of exercise on exhaled breath

• Controlled bicycle exercise
• different work loads
• Ventilation monitored continuously
• MV, O2, CO2, Anaerobic threshold
• Cardiac function monitored continuously
• CO, HR, Stroke volumes, Peak filling emptying
  rates

29
Cardiac output and minute ventilation as a
function of exercise
30
Breath ethane as a function of exercise
31
Breath acetone as a function of exercise
32
Biomarkers of exposure

• chlorinated hydrocarbons
• benzene
• JP-8
• Anesthesia

33
Warfield Air National Guard Protocol

• Goal To quantify individual exposure to JP-8 and
  correlate JP-8 exposure with adverse health
  effects
• Requirements
• Provide a breath sample before work (pre)
• Provide a breath sample after work (post)
• Breath biomarkers quantified in breath were
• hydrocarbons, CO, NO, and total sulfur
  compounds
• Provide a blood sample after work
• Provide a urine sample after work
• Take a computerized neurocognitive test
• Complete and return a questionnaire

34
Categorization of Subjects
63 Volunteers
Task Performed
Smoking Status
17 smokers 46 nonsmokers
13 crew chiefs (CC) 6 fuel cell workers (FC) 6
fuel specialists (FS) 10 mechanics (ME) 28
incidental workers (IN)
35
Aircrafts at Warfield
A10
C130
36
Crew Chiefs
Photograph by Pliel
Duties Perform pre- and post-flight routine
inspections Engine startup times around 15-30
minutes Typically 2 takeoffs each day
37
Fuel Cell Workers
Duties Pull foam in hangar 3 personnel
switching tasks Safety equipment respirator,
boots, cotton coveralls, and
gloves
38
Fuel Specialists
Duties Receive JP-8 on base Check
JP-8 for contaminants and additive
concentrations Refuel aircrafts on the flight
line Safety Equipment worn gloves
39
Pre- and 4 hr post occupational exposure to JP-8
40
Did Total JP-8 Exposure Increase during the day?
41
Route of Exposure to JP-8
42
Health Effects Associated with JP-8 Exposure at
Warfield AGB

• No correlations with liver or renal effects
• Air National Guard individuals performed poorer
  on 20 of the 42 outcomes
• The majority of decreased performances occurred
  on response time measurements
• Performed worse on 3 out of 5 multitasking
  exercises

43
How does Exposure Compare Among Military Bases?
44
Exposure to anesthetics in PACU

• Question
• Can exposure of nurses in recovery room to
  anesthesia be estimated from exhaled breath?

45
Isoflurane in exhaled breath on Friday
46
Isoflurane in exhaled breath on Monday
47
Changes in occlusion pressure
48
Summary of breath analysis

• Modern breath analysis has a history of more than
  35 years
• Breath analysis is non-invasive
• Breath can be easily collected in field, clinic,
  in-patient, OR and ICU
• Breath can be collected from neonate to the
  elderly (mouse to horse)
• Breath can be collected multiple times without
  risk to patient
• Children give breath samples willingly
• Many modern analytical methods have sufficient
  sensitivity to detect breath molecules

49
Exciting new directions for breath analysis

• Portable hand held, real time breath monitors
  using- quantum cascade lasers in the infra red
  mini mass specs mini GCs etc
• Breath profiling
• New diagnostic substrates with different
  selectivities- identifying genetic abnormalities
• Diagnoses in field or developing countries -
  where maintaining blood samples can be difficult
• Diagnoses in neonatal intensive care unit-
  easier to get breath than blood or urine
• Using breath to determine exposure to pollutants
• Breath condensate

50
Collaborators

• Study Subjects Patients, Nurses, Air Force
  Air Guard Personnel
• Former students Cope, Tu, Sehnert, Long, Kazui,
  Andreoni, Fleischer, Solga
• Pediatrics Schwarz, Marban
• Surgery Bulkley, Burdick, Cameron
• Cardiology Lowenstein, Gerstenblith
• Anesthesiology Brown, Merrit, Nyham
• Pulmonary Medicine Orens, Studer
• Oncology Yung, Abrams
• Hepatology Diehl, Solga
• Weight loss, exercise Cheskin, Gerstenblith
• Cognitive Studies Kay
• Aging NIA
• Support NIH, USAFOSR