Review paper: Practical approaches to broiler litter odour analysis
Sashikala Maruthai Pillai and Richard Stuetz
UNSW Water Research Centre, School of Civil and
Environmental Engineering, University of New
South Wales, Sydney, NSW, 2052, Australia
1.0 Australian Broiler Production
Meat chicken industry is a one of the major agricultural sector in Australia with
improved meat production of 487,929 tonnes in 1996/97 to 811,591 tonnes in 2006/07
creating AUD$1.4 billion of profit. Australian Bureau of Agricultural and Resources
Economic (ABARE) has confirmed an increase in the consumption of chicken meat
and expecting for more growth from 2009 to 2010 due to reduction in the feed grain
prices (www.abareconomics.com/interactive/09ac_mar/htm/poultry.htm 4th Nov
2009). These meat chickens have high demand both locally and internationally as it is
free of hormones and cheaper in price compared to beef, pork and lamb meats which
use hormones in their productions
t_on_hormones_in_chicken_meat 4th Nov 2009).
Mostly meat chickens in Australian are grown in mechanised tunnel ventilated sheds
having the capability to hold up to 40,000 chickens per shed at a time drawing air
using huge ventilation fans at high speeds across the chicken on thick layer of bedding
material. Chicken growth as broiler ensured with adequate supply of feed and water in
proper feed and water lines in the sheds at all times. A complete growth cycle of
broiler is approximately 7 to 9 weeks and the first harvest occurs around 30-35 day
and the last at 55-60 day before the sheds are cleaned, disinfected and spread with
new bedding material to place next batch chicks to reduce the risk of common ailment
between broiler batches. Bedding material used by integrators varies as freshly collect
material as rice hulls and straw, paper, wood chips, sunflower husks, timber shavings
and sawdust and/or used bedding material from previous batch or composted litter
environment/litter-materials 12 Nov 2009).
However, this industry is very exposed to environmental issues relating to odour
generation from poultry sheds which includes greenhouse gases, particulates and
volatile compounds including volatile organic compounds and ammonia. Broiler
production facilities are categorised as intensive livestock practice areas which means
it has to ensure the environmental impact caused is at a minimum level to the
surrounding area or sensory receptors. Bedding material transforms into litter
consisting of manure, urine, feed, chicken feathers, dust, bedding material and water
at various proportions over the broiler growth cycle (Burne and Rogers 1999) is the
major source of odour. Other sources of odour are the spilled feeding material and
dead chickens which are not eliminated from the sheds at ones. Thou animals
themselves have the potency to generate odours which come from their body, these
emissions considered to cause insignificant annoyance to sensory as the
concentrations are low.
Fundamentally, aerobic and anaerobic biodegradation processes of organic materials
generate odorous gas at rates depending on the production and accumulation of litter
with rapid growth of birds and the natural features within the facility itself (Dorling
1977; McGahan, Kolominskas et al. 2002). Micro-organisms break down the
abundant carbohydrates and proteins in the litter generating offensive odorous
compounds mostly due to oxygen depletion (Mackie, Stroot et al. 1998). Changes in
temperature and humidity, poor bird health, dietary upset, inappropriate drinking lines
and inadequate insulation or ventilation create localised wet litter and birds
contributes to incline in odour generations. Removal and breaking of caked thick litter
after final harvest of chickens from poultry sheds enhance immediate rapid
volatilisation and production of dust.
Establishments of cities closer to the poultry facility (urban encroachment) affect the
life quality of residents and their properties by increasing the potent for odour
nuisance complaints (Gostelow, Parsons et al. 2001; Nicell 2009). Constant and
increase of complaints may cause difficulties to both breeders and authorities
especially when it comes to decide on closure or relocating any livestock operations
which will create loss of employment and income.
Review of livestock odour
Obnoxious odour generations from animal facilities abundantly contribute to air
pollution (Skinner, Lewis et al. 1997; Hobbs, Misselbrook et al. 1999; Mahin 2001;
Powers, Angel et al. 2005; Rappert and MuÌˆller 2005) due to expansion in urban
encroachment and demand for meat, egg and milk intake for human diet (Mackie,
Stroot et al. 1998; Mahin 2001; Centner 2003). Generally, livestock emissions
contains dust, range of micro-organisms and odorants as a result of stocking manure,
bedding materials, ventilation fans, animal and animal feed within the facilities
(Carney and Dodd 1989; Wathes, Holden et al. 1997; Mackie, Stroot et al. 1998;
Ullman, Mukhtar et al. 2004; Rappert and MuÌˆller 2005). These volatiles in the
emissions may instigate great impacts on the environment, global climate change,
health of human and animal and their products (Mackie, Stroot et al. 1998; Schiffman
1998; Seedorf, Hartung et al. 1998; Radon, Weber et al. 2001; Tech 2001; De Boer
2003; Krupa 2003; Nimmermark 2004). It is note worthy than an individual need not
stay near an emission area to be affected by the odorants in the plume of an odour
(Shen and Sewell 1984). Some volatiles from the livestock vicinities are highly stable
throughout the emission and dispersion processes from the point of source to sensitive
Volatiles associated with livestock facilities were mostly isolated, identified and
quantified using diverse analytical or sensory to investigate relationship between
concentration and intensity of perceived odour and to mitigate emissions’ productions
(Schaefer 1977; Yasuhara 1987; O'Neill and Phillips 1992; Misselbrook, Clarkson et
al. 1993; Chen, Liao et al. 1994; Jensen, Persson et al. 2000; Rabaud, Ebeler et al.
2002; Rabaud, Ebeler et al. 2003; Wright, Eaton et al. 2004; Wright, Eaton et al.
2005; Lu, Lamichhane et al. 2008; Trabue, Scoggin et al. 2008). Investigations were
relatively sampling of the air from animal facilities such as at the ventilated fan, swine
buildings, swine lagoons and ponds. Nevertheless, issues on odour tend to rise though
numerous emission reduction technologies were introduced and put into practice
In the case of poultry odour, very limited researches and publications were produced
in the past (Yasuhara 1987)compared to piggery and dairy farm odours. It is important
to note that investigation on emissions produced from poultry production vicinities is
a critical field to be explored (Powers, Angel et al. 2005) as it has experienced
tremendous growth vastly with limited poultry odour research found in the past
leading to uncertainty of the odour assessment and impact on the environment and
human health. Lack of supporting scientific evidence and appropriate measurement
methods of emissions’ concentrations and rates from poultry buildings remain as
significant limitation to assess poultry odour impact thou efforts to minimise effect of
emissions have been proposed (Shen and Sewell 1984; Mahin 2001; Powers 2003;
In 1970s, an early study was conducted comparing organic compounds extracted
from manure using solvent extraction method and air from confinement building
identifying indole, skatole, phenol, p cresol and carboxylic acid using both
instrumental and sensory techniques. However, the technicalities of method of
sniffing were not discussed in detail by author. Author reported fewer compounds and
concentrations in the air compared to manure extract over time and no change in the
proportion of compounds with the change in season.
Similar studies to conducted to identify compounds from poultry manure at different
time length listed out seventy two compounds using freeze vacuum distillation (Table
2.6)(Yasuhara 1987) and twenty seven compounds from composting poultry litter
(Table 2.7)(Turan, Akdemir et al. 2007). However, separation of volatile components
using freeze vacuum distillation method was used selectively targeting compounds
other than aliphatic hydrocarbons and unfortunately created artefacts. It is note worthy
here all volatiles have individual partial pressure to volatilise into atmosphere. Hence,
the lengthy technique of volatile separation is questionable for the resemblances of
ambient environment sampling. Periodical sampling and the nature of the odour
generating resources are important factors in odour emissions data establishment
(Miller and Woodbury 2006).
However, the outcomes from these studies have limited use due to the lacks in the
experimental designs providing inappropriate and incomparable information on
emission levels, description on mass transfer processes and analysis procedures as
they do not replicate the actual environmental status (Rappert and MuÌˆller 2005;
Hudson and Ayoko 2008; Hudson and Ayoko 2008). Point source odour study with
appropriate sampling and analysis method combining both sensory and instrumental is
expected to provide accurate estimation of emissions animal houses to establish
feasible and equitable policies (Dorling 1977; Powers 2003; Powers, Angel et al.
2005). Identification of the source emitting odorous compounds and measuring odour
flow are very important elements to determine the accurate and right treatment for
mitigation techniques (Rappert and MuÌˆller 2005). A researcher with co-worker have
discussed extensively current challenges faced and applicable solutions regarding
poultry production’s emission (Powers, Angel et al. 2005). Power et al (2005) has
identified methods of emissions measurement, location of sampling, periods, duration
and frequency of measurements conducted, influence of weather and dietary as
aspects with wide gaps to be solved. Measurements of odours at ambient condition at
point sources are appropriate for regulatory purposes considering the opportunities for
dilution of emissions before reaching the receptor. Impact of animal nutritional values
as source of emissions is poorly understood.
Table 2.6 Compounds found from poultry manure (Yasuhara, 1984)
Ethanol Ethyl formate
1-Methylethanol Propyl formate
Propanol Butyl formate
2-Butanol Ethyl acetate
2-Methylpropanol 1 –Methylethyl acetate
Butanol Propyl acetate
2-Pentanol 2-Butyl acetate
3-Methyl-2-butanol 2-Methylpropyl acetate
2-Methylbutanol Butyl acetate
3-Methylbutanol 3-Methylbutyl acetate
Pentanol Pentyl acetate
4-Methylpentanol Ethyl propionate
Hexanol 2-Methylpropyl propionate
2-Phenylethanol Butyl propionate
Phenol Ethyl 2-methylpropionate
p-Cresol Methyl butyrate
p-Ethylphenol Ethyl butyrate
Diacetate of 1,3-propanediol Propyl butyrate
Aldehydes 3-Methylbutyl butyrate
Propanal Pentyl butyrate
Butanal Ethyl 3-methylbutyrate
2-Butanone Dimethyl sulfide
2-Hexanone Dimethyl disulfide
2-Heptanone Dimethyl trisulfide
Carboxylic acids Acetamide
Acetic acid Propanamide
Propanoic acid Butanamide
Butanoic acid Nitrogen heterocycles
3-Methylbutyric acid Indole
Pentanoic acid Skatole
2-Methylpentanoic acid 2-Methylpyridine
Table 2.7 Compounds found from poultry litter compost (Turan, 2007)
Aldehydes Carboxylix acids
Acetaldehyde Acetic acid
n-Propanol Methyl ethyl ketone
Pentane Methyl chloride
Cyclohexane Ethyl benzene
Nitrogen heterocycles Carbon dioxide
1.5 Aim of the study
In the past, all analysis of emissions from poultry facilities were conducted using
either sensor or instrumental measurements and emphasise was given more for
specially or single targeted compounds in the emissions to be investigated such as
ammonia and hydrogen sulphide which have been in focus for many years in various
livestock odour studies. It is important to point out that such kinds of studies are not
sufficient enough and do not represent the ‘real’ environment odour emission
estimation and its rate as odour is not caused by one single compound but as a set of
mixed compounds. Investigations to be carried out on odour emissions must be as a
whole at ambient condition rather than just aiming on a particular compound to
improve air quality.
For this purpose, traditionally used solvent extraction or trapping on adsorbent
followed by gas chromatography-mass spectrometry (GC-MS) analysis method which
is the instrumental study alone should be replaced with direct headspace analysis
coupled to gas chromatography-mass spectrometry-olfactometry (GC-MS/O) at the
ambient environment. Similar methods have been used vastly for the analysis of odour
and taste in the perfumery, food and wine industry for many years since 1940s (Van
Ruth 2001; Delahunty, Eyres et al. 2006) and they will be reviewed in following
sections. Such data gathered is expected to assess the variations in emissions
thoroughly from poultry sheds in order to improve waste management to reduce odour
emissions and to provide scientific evidence as guidelines to the local authorities on
odour prevention and abatement framework. This paper aims to review on how
investigation of odorants from broiler litters and its variations in emissions can be
conducted by relating both sensor and instrumental measurement using direct
headspace GC-MS/O technique.
2.0 Perception of odour
Odorants are volatile compounds responsible for creating odour which stimulates
human olfactory system(Gostelow, Parsons et al. 2001; Stuetz and Frenchen 2001).
These odorants must have the ability to volatilise at ambient temperature to ease the
absorption of substance in the mucus layer on the sensitive surface of epithelium. In
addition, these odorants should not be substances existing on the olfactory epithelium
in order to avoid errors in identification of stimuli. Smell receptors in human are build
in the olfactory epithelium in the roof of the nasal cavity. It has approximately five to
six millions of olfactory receptor neurons. The olfactory epithelium consists of three
types of cells a) the olfactory cell b) supporting cell c) the basal cell with each cell
sends information in the form of electrical signal to the olfactory bulb in the forebrain
where it is processed and spread to other parts of the brain that detects and identifies
Human nose is an efficient odour detector compared to any scientific instrument. To
date, no one analytical instrument can measure or evaluate an odour in the manner a
human nose does. The human nose detects and differentiates thousands of volatiles as
low as part per billion in concentration in the ambient air (Guyton, Chiavarelli et al.
1987; Ganong and Coleman 1997). In addition, most importantly, the human nose has
the capabilities to get attracted and rejected according to the perceived stimuli
(Nimmermark 2004). For instance, individual approaches pleasant odours related to
food and taste and aromatherapy but avoids and become aware of identified hazards in
the environment such as spoiled food, smoke and infections. In fact, a study has been
conducted in past on patients diagnosed with probable and questionable Alzheimer’s
disease testing on the efficiency of their olfaction system as an early indicator of that
particular disease (Morgan, Nordin et al. 1995). Sense of smell trains and reminds a
person of awareness and sensitivity of its surroundings.
Intensity of odour is the strength of odour perceived above its threshold level and it is
very much related to its odour concentration (Stuetz and Frenchen 2001; Nimmermark
2004; Nicell 2009). Response of an olfactory receptor depends on the intensity of an
odour. It is the individual perception on the odour’s concentration. A common way of
measuring odour intensity is to compare the intensity of an odour to the intensities of
different but known concentrations of a reference odorant such as the commonly used
n-butanol. However, it can also be described using Fechner’s law and/or Steven’s
power function (Misselbrook, Clarkson et al. 1993). Terms as not perceptible, weak or
strong are used to scale the odour perceptions (Table 1).
Table 1. Odour intensity scaling
Odour concentration Intensity level
Not perceptible 0
Very weak 1
Very strong 5
Extremely strong 6
As mentioned earlier, no one device measures an odour like the human nose. To date,
dynamic olfactometer is the most suitable instrument to measure an odour
concentration by presenting odorous air samples to odour panel in a range of dilution
detecting the presences of an odour. According to European air quality standard draft
(CEN 1999), odour concentration is best expressed in odour units per cubic meter
(OU/m3) or European odour unit (OUE) expresses as OUE/m3. It defines the volume
of diluent needed to dilute a unit volume of odour until the detection threshold of the
odour is gained. Alternatively, odour unit per cubic meter also defines the
concentration of odour in one cubic meter of air as the panel detects the threshold of
the odour. The European air quality standard (CEN 1999) also defines European
Reference Odour Mass (EROM) is equivalent to 123 μg n-butanol evaporated into 1
m3 of neutral gas air. All odours are only detectable at a concentration of 1 OUE/m3.
Odour intensity is a way to compare the strength of an odour perceived with or to
another. At a higher concentration, some odours may be perceived as very weak while
others may be perceived as distinct. The Weber-Fechner law (Eq 2.1) is used to
develop the relationship between intensity and concentration as
I = kw log (C/C o) + Const (Eq 2.1)
with I is the intensity of perceived odour, kw is the dimensionless of Weber-
Fechner constant, C is the concentration of odorant, Co is the concentration of odorant
at the detection threshold and Const is a constant which relates to the use of mean
Threshold is the minimum concentration required by the sensory property to detect an
odour. It is often determined by 50 % of the odour panel consisting of a specified
number of people (5-8 persons) using olfactometer (Voorburg and Kroodsma 1992).
Generally, all volatiles have their own threshold limits. Two levels of threshold
existing in the olfactometry science are the detection threshold and the identification
threshold. The threshold for detection is the minimum concentration needed by an
assessor to identify between sample and blank and need not to identify the odour. The
threshold for identification or recognition is the minimum concentration needed by an
assessor to identify accurately and correctly character of a volatile compound. This is
often difficult as odour exists in the form of mixture and some compounds have the
tendency to mask the other compounds in the mixture (Nimmermark 2004). However,
odour threshold aspect varies among the human population due to nature of the
chemical itself, sensitivity, age, gender, social habits, occupation and state of health of
panellist (Bliss, Schulz et al. 1996; Nimmermark 2004). Mostly women are much
more sensitive and have lower odour threshold detection limits compared to men and
the ability to detect an odour declines with the increase in age especially after 60
Odour character is explanation using words on how an odour smells like (Table 2).
Usually odour descriptors based on source of odour will be provided to panel to help
them to describe the odour perceived. As odour descriptors vary from one to another,
no one descriptor can satisfy or match another completely. The American Society of
Testing and Material (ASTM) has the most collection of descriptor for over 800
compounds (Stuetz and Frenchen 2001). Characteristics of odour can be revealed
using proper methods such as the gas chromatography-mass spectrometry.
Table2. Odour characters and threshold of compounds (Stuetz and Frenchen 2001)
Methyl mercaptan Decayed
Acetic acid Vinegar
Acetone Fruit, sweet
Indole Faecal, repulsive
Ammonia Sharp, pungent
Valeric acid Sweat
Hedonic aspect of an odour is related to its pleasantness or unpleasantness which is
directly related to odour intensity and concentration. Unpleasantness increases
proportional with the increase in odour concentration. While evaluating an odour in
the laboratory for its hedonic tone using an olfactometer, panels are exposed to a
controlled stimulus in terms of intensity and duration. The hedonic scale ranges from -
4 to +4. Negative sign indicated the most unpleasant and the positive sign indicated
the most pleasant odour. Nevertheless, most pleasant odour may still become
unpleasant with the increase in intensity and concentration causing annoyance to
sensitive receptors. The degree of pleasantness or unpleasantness is very subjective as
it is very much influenced by panel’s experiences, psychological and emotional
factors associated with a particular odour.
3.0 Impact of odour
Due to the presence of hazardous volatile compounds and micro-organisms, livestock
odours are becoming serious toxicants affecting human and animal health prominently
than as nuisance. According to the World Health Organisation, health is defined as a
state of complete physical, mental and social well being
(http://www.who.int/about/definition/en/print.html 19th Nov 2009). Health of an
individual may not be predicted merely with the absence or non-appearance of
diseases. Constantly, regulators and environmental groups have shown great concern
on livestock area volatiles as they are classified as hazardous pollutants (Turan,
Akdemir et al. 2007) and have the capability to remain chemically stable thou being
dispersed widely from rural to metropolitan areas. Recent research has suggested
effects of odour on the health of neighbours from large animal production facilities
(Schiffman 1998; Wing and Wolf 2000; Nimmermark 2004) and in some cases it has
the potency to alter human health (Schiffman 1998).
Odour is often regarded as an environmental stressor because of its psychological and
depressive impact to sensitive receptors (Nicell 2009) (i.e. loss of appetite, nausea,
fatigue, vomiting, headache and insomnia). In long time perception stress, illnesses
may lead to heart and blood vessel diseases depending on the decline in the immune
defence. Odorants may also have physical impact on individuals especially impact of
gases present in both high and/or low concentrations and reasonable amount of micro-
organisms causing sensory irritation, tears, asthma-like reaction and allergic
symptoms and reactions.
Exposure to a particular odorant for long time repeatedly may lead to a decrease of
sensitivity to that odorant due to adaptation which makes it possible for human to
react immediately to changes in stimulation. Workers working or exposed to odours
from livestock facilities regularly for long term may have changed or different odour
perception due to high level concentrations of chemicals which might have decreased
their sensitivity and lead to a non-understanding of odour complaints from neighbours
with irregular exposure. However, adaptation is faster and greater to unpleasant than
to pleasant odours. It may cause immediate discomfort reaction from a person due to
past experience thou the intensity of the unpleasant odour is at an acceptable level.
At some point, these odours cause serious uneasiness in human activities and
enjoyments socially and economically such as avoidance of outdoor recreation, mood
swing and difficulties in food preparing, embarrassment while having visitors,
additional washing or cleaning of residence and clothing and decline in business.
Animals’ health and their products are may be affected prominently by emissions
from animal concentrated buildings. S. M McGinn (2003) has reported in decline in
animal production and incurring of pneumonia in cattle due to high content of
ammonia in beef feedlots air.
Add effect on animal and their products
Add effect on ecology
4.0 Odour Analysis Techniques
Odour analysis can be conducted using both sensory and/or instrumental measurement
methods. Selection of the right method for odour measurement depends very much on
the objective/s of the particular analysis. Sensory measurement method which is in
practice for many years is the olfactometer using static or dynamic dilution. The
dynamic olfactometer (Figure 2.7) which uses dynamic dilution method is the best
instrument due to its efficacy to transfer sample to the smelling port with a minimum
impact of sample adsorbed onto the instrument surface (Stuetz, 2001). It was
developed in the 1980s in Netherland. Odour intensity, hedonic tone and characteristic
can be judged using this instrument based on guidelines prepared for olfactometer.
Sample is presented to a group of trained assessors to determine the dilution factor at
the 50 percent of detection threshold in an odour free environment. Trained assessors
are selected from various age limits and gender to produce overall community
representing data rather than focusing to a selected gender or age group people.
Odour samples are usually collected in Tedlar bags from the sites before being
analysed. This air sample sent to the mixing chamber in the olfactometer to be mixed
with the odourless air before being distributed to the smelling ports for the panels to
assess. Dilution of the air sample is subjected to be in descending order to avoid
intensed odour to stay along in the lines of the olfactometer tubings in order to
produce better results by introducing less intensed to more intensed air sample to the
panels. Dynamic olfactometer has been used widely in the waste, livestock and
wastewater treatment compared to any other industry (Carney and Dodd, 1989) to
identify the relationship between the odour concentrations, dust particles and intensity
(Misselbrook, 1993; Hobbs et al, 1998). It is very handy and low in price compared to
very technical instruments. One can handle this instrument easily and does not require
any additional or special equipment for it to function. Data obtained from dynamic
olfactometer can be used to estimate odour emission rates, dispersion modelling and
However, this sensory odour measurement is very subjective and has its drawbacks. It
has to be calibrated regularly to enhance accurate measurement. This step is not easy
for some olfactometers. The accuracy of the results produces often questioned for its
repeatability as results regarding odour strength and odour concentration depend on
the panels’ decision. For this reason, panels have to be trained and screened before
becoming an assessor using standard odorous gas. Commonly used standard odorous
gas is the n-butanol. In addition, detailed quantitative and qualitative analysis of
concentration and components of an odour can not be performed using the dynamic
olfactometer (Lu, 2008). Schaefer (1977) investigated on odour from manure but was
unable to correlate odour intensity with volatile compounds. The most
characterisation which can be carried out is as a whole of the odour mixture rather
than the compounds present in mixture.
Gas chromatography-mass spectrometry (GC-MS) is an instrumental method that
combines the features of gas chromatograph and mass spectrometer to identify
different substances within a test sample for both quantitative and qualitative
purposes. Archer John Porter Martin a researcher who developed gas chromatography
method in the 1940s.GC-MS is becoming a tool of choice for tracking organic
pollutants in the environment. Reliability on GC-MS has increased tremendously even
though the cost of this equipment has fallen significantly. Moreover GC-MS is the
most suitable method for rapid separation of complex mixtures that contributes to
odour formation due to the development of capillary column which gives better peak
resolution. Separation of compounds happens in GC column because compounds have
different affinity to separate (Stuetz 2001).
The GC-MS system is made up of two major sets which are the gas chromatograph
and the mass spectrometer (Figure 2.8). Packed or capillary column is placed in the
GC system. Better separation can be obtained based on (Eq 2.3)
N = 16( T / W ) 2 (Eq 2.3)
with T as retention time and W as peak width on the baseline. Height measurement is
equivalent to (Eq 2.4)
H=L/N (Eq 2.4)
with L as length of the column. There is a correlation between film in the capillary
column and the internal mobile phase or gas which effect the separation between
peaks. This can be formulate as (Eq 2.5)
Phase ratio = column radius / ( 2 x film thickness) (Eq 2.5)
Retention increases when the phase ratio decreases.This gives better resolution of
peaks. Separations of different molecules in a mixture introduced into the GC system
happen as the sample travels the length of the column. Molecules take different
retention time to elute out of the gas chromatograph before it allows the mass
spectrometer downstream to capture, ionize, accelerate, deflect, and detect the ionized
molecules separately. The molecules are break into ionized fragments and detect
using their mass to charge ratio (m/z).
Figure 2.8 GC-MS schematic
It is almost not possible to make an accurate identification of a particular molecule by
gas chromatography or mass spectrometry alone. The mass spectrometry process
normally requires a very pure sample. Gas chromatography using a traditional
detector such as the flame ionization detector (FID) detects multiple molecular that
happen to take the same amount of time to travel through the column which results in
two or more molecules to co-elute. Two different molecules can have a similar pattern
of ionized fragments in a mass spectrometer. Combining the two processes makes it
extremely unlikely that two different molecules will behave in the same way in both a
gas chromatograph and a mass spectrometer. Therefore when identifying mass
spectrum appears at a characteristic retention time in a GC-MS analysis, it typically
lends to increased certainty that the analyte of interest is in the sample.
Selecting the right GC column is very essential. Normally, column supplying
companies provide suggestions and advices on column selection (Table 2.4). Selected
column must have the ability to detect and separate compounds, plus with minimal
effect of oxidation and adsorption of sample while in process. Generally there are two
types of columns for GC use a) packed column and b) capillary column. Packed
columns are glass or stainless steel coil with dimension of 1-5 m total length and 5
mm inner diameter (Figure 2.9). It is filled with the stationary phase, or a packing
coated with the stationary phase. Capillary columns are thin fused-silica capillary with
dimension of 10-100 m in length and 250 um inner diameter with stationary phase
coated on the inner surface (Figure 2.10). Currently many odour studies were
conducted by using capillary column as it provides better separation efficiency than
packed columns (Stuezt, 2001; Sandra, 1980). Common stationary phases in gas-
chromatography columns are polysiloxanes, which contain various substituent groups
to change the polarity of the phase. The nonpolar end of the spectrum is polydimethyl
siloxane, which can be made more polar by increasing the percentage of phenyl
groups on the polymer. For very polar analytes for instance odour or malodorous
compounds, polyethylene glycol is commonly used as the stationary phase. After the
polymer coats the column wall or packing material, it is often cross-linked to increase
the thermal stability of the stationary phase and prevent it from gradually bleeding out
of the column.
Very few detectors can be used to detect odour compounds. The most common one is
the flame ionization detector (FID). FID is robust and sensitive primarily to
hydrocarbons compared to other types of detectors. However, the sensitivity of FID is
less for sulphide compounds (Stuetz, 2001). Gas chromatograph connected to a mass
spectrometer functioning as a detector is indeed common too. Even though the mass
spectrometer is expensive, it has great ability to detect unknown compounds and very
compatible with capillary column used in GC.
Combination of sensory and instrumental
Odour is mixture volatiles at ambient temperature. Human identify the odorants in a
mixed mode and describes odour in different perceptions. Thus, gas chromatography-
mass spectrometry-olfactometry is a great tool to measure volatiles using both sensory
and instrumental measurements. The combination of measuring odour and odorants is
called gas chromatography-olfactometry (GC-O) (Figure 2.11) (Friedrich and Acree,
1998) or gas chromatography-olfactometry-mass spectrometry (Hochereau and
It was first proposed by Fuller and co-workers in 1964 as GC-O and used in the
selection of odour active compounds from a complex mixture. Due to serious problem
in reproducibility and hot effluent from the GC to the odour port, this method was not
widely used then (Van Ruth 2001). However, as these limitations were fixed, GC-O
technique has made a come back starting from 1971. Headspace sampling techniques
suit very conveniently for the GC-MS-O analysis. This system can be used detect all
odorants in a mixed mode but proper sampling and chromatography analysis must be
Chromatograms from GC responses differently to the chromatogram collected from
olfactometry studies. Odour active compound is not necessarily has to be the one with
major peak area from GC chromatogram. Olfactometry studies only detect the
volatiles which are odour potent responding to panel olfactory receptor. It is a reliable
technique to select and identify individually volatiles contributing to odour even at a
very low abundance levels observed on a GC chromatogram. According to Van Ruth
(2001) GC-O data processing are grouped into four classes as:
a) Dilution analysis
Charm and aroma extraction dilution analysis (AEDA) are two different dilution
analysis commonly used to detect potent odour compounds in food as quality control
markers (Song et al, 2008). Charm analyses the detection time of a volatile begins and
ends to be perceived by a panel. This technique usually time consuming and requires
many sniffers. AEDA determines the last dilution in which odour compounds are
detected. Results from AEDA are represented in logarithm of the factor of dilution
against retention index. Both Charm and AEDA are similar as samples are subjected
to dilutions and based on the odour detection threshold principle.
b) Detection frequency analysis
This technique applies detection of threshold with a group of assessors to detect odour
active compounds from undiluted sample at the sniffing port. Assessors evaluate the
sample at the sniffing port while simultaneously measuring the intensity of
compounds. Described peaks or compounds by the assessors are analysed as
frequency of the presence with description recorded for each volatile and not based on
the intensities of the compounds.
c) Posterior intensity analysis
Posterior intensity analysis is not frequently used as it quite complex in task for the
assessors. It records odour intensity as volatiles are eluted from GC system. Scale
used by the assessor differs and complicates the assessment process.
d) Time intensity analysis
Time intensity analysis (OSME) was developed by McDaniel to estimate odour
intensity. OSME means smell in Greek. An extracted sample is injected into a GC
column for the compounds to separate and simultaneously these compounds to two
detectors which are the mass spectrometer and the olfactory detection port. Volatile
eluted from GC column assessed by trained panels at the sniffing port and no dilution
is required. Assessor evaluates the intensity using modes given (i.e light, mild, high
and very high) and describes verbally the smell. Results are computerised as intensity
versus retention time and matches GC chromatogram of the sample examined.
Van Ruth (2001) has reviewed on the methods for gas chromatography-olfactometry.
Author has well discussed techniques to collect and process GC-O data plus
estimation on odour active compounds. The best part of this paper is the author has
also listed the advantages and disadvantages of methods used in the GC-O. It can be
used as a good starting guidance for new researches to select techniques involves the
5.0 Odour sampling techniques
Headspace analysis is a method used to separate volatiles from a liquid or solid prior
to instrumental analysis. It is the simplest way to collect and analyse sample
quantitatively and qualitatively (Valentin, 1975: Kolb and Leslie, 2006).
For the first time in world history, headspace analysis was carried out on vapour
above liquid and was reported firstly by Harger, Bridwell and Raney in 1939 (Snow
and Slack 2002). Static and dynamic headspace investigations were carried out using
gas chromatograph (GC) in 1958 and 1970 respectively. At present, headspace
analysis methods are being conducted in many areas of research especially perfume,
agricultural, livestock, environmental and food and flavour industries.
Attempts were made to categorise the headspace techniques by differentiating
sampling parameters. Nunez et al (Nunez, Gonzalez et al. 1984) has written an
excellent review on preconcentration methods of headspace volatiles for organic
tracing using cryogenic trapping, adsorption or desorption method. Massive literature
review consisting of over 200 reference papers was conducted for this writing.
Emphasis was given more on the thermodynamical equilibrium phase to reduce or
eliminate the lacking in the preconcentration method especially when the sample is a
non fluidic or solid sample. Direct sampling, indirect sampling and closed circuit and
the use of different kinds of solid sorbents to trap headspace volatiles were discussed
briefly. However, the survey was rather confusing the readers. Various points stated
on the headspace technique are becoming invalid or inadequate as this method is
developing technically from time to time.
The headspace method was later classified into 4 classes according to the way of
samples are introduced into the gas chromatography system which were the full on-
line, off-line concentration with on-line desorption on to the column, techniques
involving both off-line concentration and desorption and techniques involving liquid
desorption from a concentration trap (Grob and Habichi, 1985). This seems to give a
better introduction and concept of headspace analysis besides discussing on
advantages and disadvantages of various headspace method, preconcentration
techniques and also adsorbents. Author has pointed out on insufficiently known
parameters of headspace analysis and also alteration to be made to overcome the
Snow and Slack (2002) reviewed the use and categories of headspace method in
modern chromatography. Mainly there are three types of headspace techniques which
are the static (SHS), dynamic (DHS) and solid phase micro extraction (SPME) used
in taste, aroma, food, flavour, environment and pharmaceutical researches (Snow and
Slack 2002). Many researchers have started preferring headspace sampling to any
other types as it is very easy to handle samples using these techniques, simpler, fast
and many are automated too. Major advantage of headspace technique is it reduces
the interferences of solvent in sample analysis and allows more volatile to be analysed
(Chen, Liao et al. 1994). This makes researchers to be much more confident with
using the SHS and SPME for sampling purpose compared to the DHS which involves
use of solvent on the sample (Bart, 2001).
Selecting the right headspace sampling method depends on the purpose of the analysis
and most importantly the nature of the samples. Headspace sampling was reported as
not very suitable for analysis of nitrogen and sulphur compounds (Valentin, 1974).
Minor drawbacks of headspace techniques can be prevented by taking precautions
such as to make sure the syringe is grease free, cleaned after every injection, at a
higher temperature than the sample to enhance the reproducibility of the analysis
(Pillonel, Bosset et al. 2002).
2.5.1 Static headspace (SHS)
This technique is the simplest headspace sampling, solvent free and directly from the
sample matrices for rapid determination of volatiles. It relies totally on volatilisation
of compounds to be separated and identified in the GC system. Static headspace
sampling is being used tremendously in natural products, pharmaceutical, clinical,
food, aroma and forensic analysis due to its sensitivity, stability and reproducibility
(Snow and Slack, 2002). For static headspace sampling (Figure 2.12), a liquid or solid
sample is placed into a vial, sealed, and heated to a specific temperature to reach the
equilibrium before the gas is removed from the vial for analysis. All components that
are volatile at or below the pre-set temperature escape from the sample to form a
gaseous phase above the sample. Upon achieving equilibrium state, the headspace gas
is extracted from the vial and injected into a gas chromatography column which
separates the various components of the sample based on size and/or polarity.
Static headspace is significantly ideal sampling choice. This is because by utilising
this method, tedious, error prone and time consuming sample preparation steps can be
eliminated (Bart, 2001). Dirty environmental samples which are not able to be handle
with syringe can be sampled easily using static headspace technique. Chen and co-
workers (Chen, Liao et al. 1994) have investigated on swine wastewater using
automated static headspace sampling and gas chromatography analysis monitoring
and quantifying the changes in the concentrations of particular odorous compounds
during the aeration. It is reported then that SHS method allowed more volatile
components to be analysed without interference from other heavy components
contained in the sample. Studies have confirmed static headspace sampling coupled
with GC system provides a useful method for monitoring the treatment process
(Chen, Liao et al. 1994). Even volatile compounds that degrade during aerobic
processes can be identified using this technique. Similar to this, Kallio et al (1990)
investigated on the changes of volatiles in roasted ground coffee as an indicator of
storage time using static headspace sampling method coupled with GC-MS system.
Determination was also conducted on various raw vegetable under a non equilibrium
condition which means short time was given to thermostat the sample in order to
avoid the alteration on volatiles (Shinohara, 1991).It reports static headspace method
used was highly reproducible even though equilibrium state was not reached. Mainly
terpenes from parsley and celery, isothiocyanates from wasabi and disulphides from
onion were successfully detected.
However, according to Bart (2001) this technique has its limitations as 1) limited
sensitivity 2) not all sample is being injected into the GC system 3) large molecular
weight compounds are not being missed out 4) has to be calibrated and 5)
quantification limitations when it is used in polymer analysis.
2.5.2 Dynamic headspace (DHS)
DHS uses a purge and trap method to collect the sample on a sorbent material before
the sample is flushed onto the column for GC analysis (Figure 2.13). It is a method
which has been used very intensively in livestock area for odour sampling (Hyttinen
et al, 2008; Trabue et al, 2008; Hobbs et al, 1998).
Using DHS, sample is purged with carrier gas (i.e nitrogen, helium) and heated in a
Teflon vessel. As the carrier gas stream exits the vessel it passes through a thermal
desorption tube filled with an adsorbent material. The outgassed products on the
adsorbent material tubes are transferred to a thermal desorption unit which is inline
with the GC.
The thermal desorption unit heats the individual tubes while a flow of gas is applied
through the tube. Collected materials are flushed from the sorbent material and
collected onto a cold trap within the thermal desorption unit. The entire sample is
purged from the sample tube and collected on the cold trap which will be heated
rapidly. The collected materials are then swept from the cold trap into the GC for
analysis as a volatile analytes.
Van (Van Langenhove, Roelstraete et al. 1985) analysed water samples from all levels
of processes of a wastewater treatment was able to identification hydrocarbons,
oxygen, nitrogen and sulphur containing organic compounds using both DHS and
solvent extraction methods.
Linear aliphatic hydrocarbons, branched aliphatic hydrocarbons, aromatic
hydrocarbons and cyclic and unsaturated hydrocarbons were identified respectively
due to the large number of samples collected and time consuming factor. Hydrophobic
volatiles from the wastewater were loaded on Tenax TA sorbent before thermally
desorbing for GC analysis.
This method is found to be the best way to quantify aroma compounds due to its low
amount of concentrations (Snow and Slack, 2002). However, some volatiles such as
the alcohol and aromatic have very low breakthrough volume compared to other
groups of chemical. Even though volatiles sampled using sorbent material can be
stored till time of analysis is an advantage, breakthrough volumes and the capacity to
desorb completely the absorbed volatiles must be seriously considered (Krost et al,
2.5.3 Solid phase micro extraction (SPME)
SPME uses a coated fused silica fiber to trap and concentrate analytes from a static or
dynamic headspace process (Figure 2.14). It has been developed in the 1993 and used
enthusiastically in environmental, pharmaceutical, food and natural products analysis.
This technique is suitable to be applied to samples with ppb low range concentrations.
The fiber is thermally desorbed to inject the analytes into GC inlet port.
Like SHS, SPME is fast, simple, portable, low in cost and convenient for sampling
(Bart, 2001). Detection of particular compounds such as the phenol group is faster
than the SHS or DHS. On the other hand, direct transfer of absorbed compounds on
the fiber into the GC system is an extraordinary advantage of this system.
SPME and GC-MS were coupled to study the applicability to determine volatile sulfur
compounds and trimethylamine from the headspace of dewatered biosolids (Visan
and Parker 2004). This system samples and preconcentrates at one same step.
Nevertheless, Visan has also reported that previous researchers have found SPME has
its limitations and occurrence of artifacts.
Wright et al (2005) initiated the use of SPME coupled with multidimensional gas
chromatography to identify odour from confined animal feeding operations.
Compounds responsible for odour were tracked and prioritized accordingly.
However, calibrating the instrument for SPME during analysis is a problem. Users
have complained regarding the method to validate the SPME which includes
extraction reproducibility and parameters involved in the system itself (Snow and
In conclusion, headspace methods vary and can not be specified in rigid groups as it is
developing rapidly. In recent years, there is a strong interest in the use of headspace in
various fields. However, trends in instrumentation show improved automation and
ruggedness and perhaps the most versatile of all sampling methods for capillary GC
system (Snow and Slack, 2002).
6.0 Comparison of the use of combined instrument and sensory to instrument or
sensory odour measurement method
As in conclusion, both sensory and instrument measurements are equally important. A
researcher must choose the right and reliable analysing method comparing the pros
and cons in order to achieve the objective/s of a project (Table 2.5).
Table 2.5 Comparison of sensory and instrumental analysis methods.(Gostelow, 2003)
Technique Advantages Disadvantages Applications
Olfactometry Measurements a) No quantitative Annoyance
relate to odours as information on odour prediction.
perceived by compositions.
b) Difficult to relate
liquid phase odorants
c) Lacking of
accuracy compared to
d) Difficult for on
Gas a) Quantitative and a) Expensive. a) Identification
chromatography qualitative analysis of unknown
and olfactometry of complex odour. b) Complex. odorants.
b) Identification of c) Difficult for on site b) Quantitative
unknown odorants sampling. analysis of
with suitable complex mixture.
detectors. d) Often needs
preconcentration of c) Development
c) Measurements sample for targeted of analytical
relate to odours as compounds. sensory links.
humans. d) Analysis of
d) Measurements transformations
can be link to and emissions.
chemical models of e) Design of
odorants odour control
transformations or equipments.
e) High in accuracy
and precision than
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