This paper has the aim of highlighting the importance of establishing suitable quality protocols applicable to electronic noses used as environmental odour monitoring systems and to describe some different approaches that can be adopted in order to perform quality checks on such instruments, thereby reviewing the relevant standards and publications in this field, and critically discuss the pros and cons related to their practical applicability.

For this purpose, the paper is divided into three parts.

The first part (Section 2) represents a state-of-the-art overview of how, in the last 15 years, electronic noses for environmental odour monitoring have evolved from a purely scientific and research level to become proper air quality monitoring tools used by plant operators as well as by local authorities for the management of odour issues through quantification of exposure or identification of emission sources. This state-of-the-art does not claim to provide an exhaustive review of all literature studies describing the application of electronic noses for environmental odour monitoring, nor to describe all currently available different electronic nose technologies, for which other extensive review papers already exist [4,15,16,48,49], but it is limited to those works proving the evolution of such instruments from research objects to regulatory tools.

The second part (Sections 3 and 4) reviews the relevant existing technical standards and guidelines that directly refer to electronic noses or that could possibly be applied to them. In more detail, Section 3 gives a short historical overview of the standardization attempts that have been made in Europe related to electronic noses, although not all necessarily referring exclusively to their environmental applications. Then, Section 4 briefly describes the relevant technical standards referring to other instruments intended for air quality monitoring, i.e., automatic measuring systems (AMSs). Although electronic noses are not AMSs, their implementation for emission and ambient air monitoring purposes, arises the question of the possibility to assimilate them with AMSs

Finally, the last part of this review (Section 5) represents a critical discussion of the possible approaches for the development and the application of standardized quality protocols for electronic noses intended specifically for environmental odour monitoring. In this section, possible alternative or complementary schemes for electronic noses qualification are proposed and discussed. As previously mentioned, such quality protocols are fundamental in order to properly characterize the instruments in terms of performance characteristics, measurement uncertainty, and effective applicability.

It is important to highlight that this paper only focuses on electronic noses used for monitoring odour as a whole, and not for the detection or identification of odorant substances.

#### **2. State-of-the-Art of Electronic Nose Applications as Environmental Odour Monitoring Tools**

The aim of this section is to give a brief overview of relevant examples of electronic nose applications for environmental odour monitoring, thereby focusing on those works that aimed to promote them as effective air quality monitoring tools. As previously mentioned, this section does not claim to give an exhaustive review of electronic nose applications for environmental odour monitoring, which have been already discussed in a recent review paper [15], but it has the aim only to describe their evolution in time from research objects to potential regulatory tools.

Historically, one of the first important works dealing with the use of an electronic nose for odour analysis and identification was reported by Nicolas et al. in 2000 [50]. This work describes the use of very simple instruments based on SnO2 sensors for measurements around real sources of malodour in the environment such as compost facilities, waste water treatment plants, rendering plants, etc. giving promising classification results with discriminant analysis and principal component analysis. The authors also highlight the influence of atmospheric conditions on the sensor responses and the necessity to carry out repeated training over time in order to compensate sensor drift. In a more recent work [51], the same authors describe the application of five electronic noses, each comprising six Metal Oxide Semiconductor (MOS) sensors from Figaro®, for the assessment of odour annoyance near a compost facility. The study proves the system to be sufficiently efficient to predict in real time possible

odour annoyance in the surroundings of the plant, even though the approach suffers from various uncertainties, from the sensors to the final determination of the distance of downwind annoyance.

One of the first examples in the literature in which the electronic nose is proposed as a methodology to quantitatively determine an odour impact was described by Sironi et al. in 2007 [52]. In this case, two electronic noses, each equipped with six MOS sensors, were used for monitoring odours from a composting plant. After training, one instrument was installed at the plant fence-line, while the second instrument was installed at a receptor located at about 4.3 km from the plant under investigation. The electronic noses analysed the air every 12 min for a 4-day period, then the sensor responses were analysed with the aim of classifying the analysed air into the different olfactory classes considered for training. This way, odour exposure could be assessed in terms of relative recognition frequency of odours from the monitored plant. This study also provides a sort of instrument performance evaluation by comparison of the outputs of the electronic nose installed at the receptor with the recordings of odour episodes of the residents. These data were presented in a confusion matrix and an accuracy index for classification was evaluated, which was equal to 72%. This result was considered as satisfactory, despite the reported influence of atmospheric humidity and sensor drift.

Another study in which the electronic nose responses were evaluated in combination with other sensorial observations (e.g., odour complaints reports and odour observations of experts) was reported by Milan et al. in 2012 [53]. This work describes a huge monitoring program aiming to map the odour impact in the Port of Rotterdam by using 40 fixed and four mobile electronic noses for a 3-year period. The objectives of investigating the electronic nose potential as an odour management tool for reducing odour exposure, as well as a safety management tool for the fast recognition of accidental gases resulting in incidents was considered as promising, although the validation procedure involving the comparison of instrumental and sensorial odour observations was not detailed in this work.

A different interesting application of electronic noses in this field was described by Chirmata et al. in 2015 [54], where electronic noses were used in order to implement a system for the continuous monitoring and tracking of odours in the city of Agadir. In this case, meteorological data were used in order to follow instantly the odour level in the study area and to identify the emission sources.

Finally, a very recent example of electronic nose implementation as an environmental odour monitoring tool in Italy is given by Licen et al. [55], who describes a 4-month survey close to a steel plant in Trieste. In that case, self-organizing maps proved to be a useful tool for visualizing the dynamic evolution of the system with time, thus allowing the identification of possible sources of malodour and evaluate frequency and duration of odour episodes.

Besides these examples, which describe the application of electronic noses as air quality monitoring tools for odour impact assessment, it is worth mentioning here that there are some cases in which the use of electronic noses for odour monitoring is foreseen on a regulatory level.

One first significant example is the French regulation about rendering plants [41], which, in article 46, foresees a reduction of the periodic odour measurement campaigns if a representative and permanent measurement is carried out by means of electronic noses.

Another very interesting example was recently presented by Cangialosi et al. in 2018 [42]. In this case an electronic nose was used in combination with an H2S continuous analyser in order to provide a continuous measurement of the odour concentration at the fence-line of a sanitary landfill. An automatic alert to the local authority was set when the odour concentration measured by the electronic nose exceeded 500 ouE/m3 for more than 5 min or when two consecutive H2S concentration measurements at 5 min intervals exceeded 20 ppb. Despite the very interesting application, the results of the study showed that the odour concentration values estimated by the electronic nose were poorly correlated with the H2S concentration measurements.

It is worth highlighting that such prescriptions involving the installation of an electronic nose for continuous odour measurement around some plants (and specifically landfills) are now becoming a common trend in the Region of Puglia, in Southern Italy.

#### **3. History of Standardization Attempts in the Field of Electronic Olfaction**

This section gives a brief historical overview of the standardization attempts that have been made in Europe in the field of electronic olfaction. These do not necessarily refer only to electronic noses for environmental applications.

#### *3.1. First Standardization Attempts: The Second Network on Artificial Olfactory Sensing (NOSE II)*

As reported in a recent and very interesting opinion paper by T. Nagle and S. Schiffman [43], the first attempt for standardization in the field of electronic olfaction was carried out in 2001, under the sponsorship of the European Commission. The Second Network on Artificial Olfactory Sensing (NOSE II) [56] was constituted and its main task was to work out its own recommendations for standards and to foster their use in the sensor and e-nose community. The work was organized in three working groups (WGs), dealing with the following topics:

However, in the end, no specific standards were completed. The reasons given were: (i) the large number of available sensor types and electronic nose technologies; (ii) the WG goals were too broad; (iii) failure by the industry to establish a generic electronic nose technology; (iv) no broad industry support for a common data format; and (v) fragmented markets with different application requirements [43].

#### *3.2. The NTA 9055.2012*

After that, in 2012, the national standardization body of the The Netherlands (NEN) was the first one who succeeded in releasing a technical agreement document (NTA 9055:2012 [57]) regarding the specific use of electronic noses for the monitoring of odour emissions that may cause odour nuisance or safety risks.

As stated in the scope of this document: "the purpose of NTA 9055 is to draw up a list of requirements for using an electronic nose (e-nose) to detect changes in the composition of the ambient air".

The scope defines the following fields of application:

Then, after a brief general description of the electronic nose technology given in Section 1 (and normative references, definitions, and abbreviations reported in Sections 2–4), Section 5 aims to describe the methodology for using an electronic nose. First, the principles of electronic nose training are generically reported. Training involves the recording of the electronic nose signals when exposed to air with a given composition and the correlation of these data with the data acquired by sensory or instrumental analysis of the same air, so that "relevant sensory or instrumental perceptions can be reproduced if the same electronic nose data is recorded". It is specified that training can be carried out at the measurement site or in a laboratory. In both cases the procedure involves collecting the electronic nose data together with information provided by other means of detection and then correlating the electronic nose data with the other information.

Then, Section 5 of this document gives a general description of electronic nose networks (Section 5.2) and of mobile electronic noses (Section 5.3). Finally, Section 6 very briefly gives some indications about sampling, thereby referring to other existing norms.

Although this technical document has the undeniable value of having been the first technical document published by a standardization body regarding electronic noses, it has the drawback of being extremely synthetic and too generic to achieve the goal of standardization of the procedures for the application of these instruments in the environmental and safety sectors. Despite the statement of the scope of the document, no requirements for the use of electronic noses are defined, except for a generic description of the instrument training. What is totally missing for a standard is a description of the procedures for the verification of the electronic nose functionality nor the validation of the instrument outputs.

After the publication of this first technical document, with all its limitations, the need for standardization on this topic at a European level became evident. For this reason, a few years later, in 2014, the European Committee on Standardization (CEN) promoted the constitution of a working group dedicated to the draft of a standard on instrumental odour monitoring (WG41), whose activity will be detailed in the next section.

#### *3.3. The CEN TC*/*264 WG41 "Instrumental Odour Monitoring"*

As previously mentioned, after the publication of the NTA 9055:2012, a new European working group (WG41) was established within CEN TC/264 on air quality to draft a standard related to instrumental odour monitoring systems. The WG41 was composed by experts nominated by national standardization bodies from European countries including Belgium, France, Germany, Italy, The Netherlands, Spain, and the UK [58].

The scope of this standard is to specify requirements for instrumental odour monitoring systems (IOMS) for the monitoring of odour in ambient air and in emissions to ambient air. Indoor air was excluded from the scope of the WG. The primary application is to generate odour metrics that are relevant indicators for the presence and attributes of odour as would be perceived by human observers. A benefit of instrumental odour monitoring systems is that they can be used for continuous measurement [58].

The working item intentionally refers to IOMS and not to electronic noses, in order to include any generic "instrument" that can be applied to the monitoring of environmental odours, independently from its functioning principle or sensing technology. Odour is here considered as a whole, thereby referring to "odour" as the "sensation perceived by means of the olfactory organ in sniffing certain volatile substances", and not to single odorants. Moreover, given the wide range of different devices for instrumental odour monitoring available on the market, based on a variety of different functional principles for gas sensing, the technical design of such systems, their calibration, training and the mathematical model that is used to convert sensor signals into output metrics are not part of this standard, which considers the system as a "black box".

According to this approach, the work of the WG was focusing mainly on the validation regimes that can be used to prove performance claims. This included defining specific objectives and limitations, thus establishing procedures aiming to verify the instrument suitability for a specific application and its utilization, within defined boundary conditions.

The validation process consists of comparing the IOMS output metrics with odour assessment metrics obtained with suitable reference methods. The reference methods for odour measurement involve the use of human assessors.

For the task of the identification odour presence and odour classification the reference method is represented by field inspections using panel members according to EN 16841:2016 (part 1 or part 2) [59,60]. The EN 16841:2016 is a European Norm that standardizes the field inspection method for odour assessment in the field by means of qualified panel members. In more detail, part 1 of the standard describes a method ("grid method") for determining the level of odour exposure within a defined assessment area over a

sufficiently long period of time to be representative for the meteorological conditions of that location. Part 2 describes a method ("plume method") for the determination of the extent of recognizable odours from a specific source using direct observation under specific meteorological conditions.

The reference method for validating the IOMS capability to quantify odours by providing an odour stimulus indicator value is dynamic olfactometry, as described by the EN 13725:2003 [34]. This standard defines the reference method for the determination of the odour concentration of an odorous gas sample using dynamic olfactometry with human assessors, thus providing a common basis for evaluation of odour emissions.

As already mentioned, up to now, the activity of the WG has been focusing more on the instrument final validation than its quality check. The main difficulties related to the development of a European Standard on this topic are associated with the fact that the currently existing technologies and applications in the different European countries, and therefore the expectations of the national representatives in the WG, are sometimes very different from each other.

However, since the activity of the WG is still ongoing and under discussion, it will be not described further in this paper.

#### *3.4. The VDI 3518-3:2018*

Very recently, in December 2018, the German VDI (Association of German Engineers) published a guideline relating specifically to odour measurements with electronic noses, i.e., the VDI 3518 Part 3 "Multigas sensors—Odour-related measurements with electronic noses and their testing" [61].

This guideline represents an important step forward towards standardization of electronic noses, although it does not refer specifically to the environmental monitoring of odours, but also to other fields of odour measurements. In more detail, the following application categories were identified:

One interesting aspect of the guideline is that it defines three different measurement tasks (functionalities) for electronic noses:

As stated in this guideline, since electronic noses can be used for a range of different odour-related measurements, the prerequisite is general suitability for the planned application.

For this reason, the VDI guideline describes a procedure for the instrument verification, which is based on three steps.

First, minimum equipment specifications for electronic noses have to be defined. For the formal testing of the equipment's suitability, data on usage, on the construction, on operating, and on the basic functionality have to be submitted.

As a second step, metrological functionality of the electronic nose has to be tested. Although demonstrating the metrological functionality of electronic noses is not a definitive proof of their suitability for the specific odour-related application, it has to be considered as a prerequisite for reliable operation of the instrument. If the metrological functionality testing is not passed, then it could hardly be expected that the instrument will be suitable for odour measurement in the desired final application. The test shall prove that, when exposed to a test gas, mechanical effects, electric interference, and climatic ambient factors, the electronic nose outcome shall not be affected by more than a deviation of 30% from the reference value.

Finally, once the metrological suitability of the electronic nose has been demonstrated, further testing is required in order to demonstrate the basic suitability of electronic noses for odour-related measurements. The tests can demonstrate the correlation between the electronic nose outputs and the results of an olfactory (sensorial) odour measurement carried out with human assessors.

Performance testing both for metrological functionality and for suitability for odour measurement has to be carried out with at least three standard test gases, containing odorants with known properties, to be chosen from a list provided in annex B of the guideline. Appropriate tests and odorants are to be chosen and the concrete test conditions agreed in accordance with the intended application.

The principle of the guideline is very interesting, since it tries to adopt a similar verification logic as those applied for other measurement instrumentation, but still considering the peculiarities of the electronic nose with respect to other chemical analysers. However, since the guideline is not application-specific, it does not account for the peculiarities of the single odour-related measurement. As a matter of fact, testing an electronic nose destined to environmental odour monitoring should entail the verification of different requirements as those that are needed, for instance, for medical diagnosis.

As previously mentioned, the guideline fixes minimum performances of the instruments to be guaranteed. The drawback of this approach is that the electronic nose technology is still very "young" and probably not mature enough to have fixed minimum requirements, which might, in the end, represent a limitation for further instrument development. Moreover, the minimum requirements fixed in this guideline are based on industrial certifications, thus hardly applicable to research prototypes and products of small manufacturers.

#### *3.5. The Italian UNI1605848 Project*

Very recently, in February 2019, the Italian Standardization Body UNI proposed a specific standard on the determination of odours by means of IOMS (instrumental odour monitoring systems) and their qualification. As already mentioned in Section 2, in Italy the use of electronic noses as air quality monitoring tools has grown significantly in the last few years, given that in some specific applications the electronic nose outputs have a legal value. As a consequence, this standard is the expression of an urgent need in Italy to provide the local authorities and the final users with an adequate normative text allowing for qualification of extremely different instruments that are proposed for environmental odour monitoring purposes.

The interesting and innovative thing about this standard is that it defines three quality assurance levels for electronic noses, following the principles of the EN 14181:2014 referred to automatic measuring systems (AMSs) for the continuous monitoring of emissions and ambient air, which will be described in the next section.

As a first step, electronic noses for environmental odour monitoring should undergo an initial metrological verification. This verification should be carried out by the manufacturer, in order to declare the instrument properties before its installation in the field. The parameters that shall be defined are, among others, the expected sensor responses to reference gases (whose composition is not specified and shall be chosen by the manufacturer) at zero and span, the effect of temperature and humidity on the sensor responses, and the response time T90. By definition, T90 is the time needed for a detector to measure 90% of the applied concentration level. In the case of electronic noses, T90 would be the time needed for the sensor to reach 90% of its maximum response to the applied reference gas.

It is also required that the capability of the instrument to provide correct results coherently with the type of determination (i.e., odour presence, odour class, or odour quantity) is verified in the laboratory before its installation in the field. The standard does not specify how this should be done in detail; the manufacturer shall provide all information related to the metrological verifications in a specific report that describes the methodology adopted.

The usefulness of this information is that, during its application in the field, the electronic nose functionality can be tested periodically by verifying that its properties declared in this initial metrological verification are still satisfied.

The essential verification step is carried out after installation in the field in order to verify the IOMS functionality as a whole, thereby including training, according to the reference method EN 13725:2003. The procedures for the verification depend on the type of determination requested (i.e., odour presence, odour class, or odour quantity). For any type of determination, the verification shall involve the comparison of at least 15 measurements provided by the electronic nose and 15 simultaneous measurements conducted in conformity with the EN 13725:2003, in the conditions that are considered to be most representative of the application. Measurements shall be carried out on at least four different days and be distributed in at least 6 h for each test day, in order to cover a 24-h cycle. The standard does not describe how the verification shall be carried out in detail, but it is required that a verification report is produced reporting all the relevant information about the test methods and results.

A mathematical approach is proposed for the evaluation of the deviation of the odour quantities provided by the electronic nose from the odour concentrations measured by dynamic olfactometry according to EN 13725:2003.

This standard, despite not being very detailed about the testing procedures, has the big advantage of being the only approach that is based on a multi-level verification of the electronic nose, which is fundamental in order to enable the instrument qualification in every phase of its life as an air quality monitoring tool.

#### **4. Relevant Technical Norms Related to Other Instruments for Air Quality Monitoring**

Besides the technical documents or standardization attempts described above, which are related directly to electronic noses, but not necessarily to air quality monitoring, other technical norms exist that refer to other instrumentation for air quality monitoring, which can be considered as inspiring for the development of quality protocols for electronic noses to be used in the field of environmental odour monitoring.

This is the case for the so called "automatic measuring systems" (AMSs), which are automatic analysers used for the continuous monitoring either of emissions or of ambient air.

Starting from 2000, CEN experts made a great effort to define all the aspects of AMSs. In more detail, the two documents that define the basic structure of AMSs are the standards EN 14181:2014 (first edition was released in 2005) and the EN 15267 series [44]. These technical standards, which describe the quality programs that must be followed in the realisation, validation and management of an AMS, will be briefly described in this section; then their applicability to electronic noses will be discussed in the next section, dedicated to the description of the approaches that can be adopted for the development of quality protocols for instrumental odour monitoring systems.

#### *4.1. The EN 14181:2014*

The Standard EN 14181:2014 [62] specifies the procedures for establishing quality assurance levels (QALs) for AMS installed on stationary sources for the determination of the flue gas components and other parameters. The following levels are defined [44].

standard reference method (SRM). For every pollutant or chemical compound of interest, the relevant SRM represents the only "legally binding" method for the limit verification of that compound.

#### *4.2. The EN 15267-1*/*2*/*3:2009*

EN 15267-1/2/3:2009 [63–65] specifies the general principles, including common procedures and requirements, for the product certification of AMSs for monitoring ambient air quality and emissions from stationary sources. This product certification consists of the following sequential stages [44]:

In more detail, the scope of EN 15267 Standards series is:

This European Standard applies to the certification of all AMS for monitoring ambient air quality and emissions from stationary sources for which performance criteria and test procedures are available as European Standard.

#### **5. Approaches for the Development of Quality Protocols for Instrumental Odour Monitoring Systems**

For the specific application of electronic noses to environmental odour monitoring, different approaches could possibly be adopted for the development of procedures for the instrument qualification. These approaches are discussed in this section.

#### *5.1. Approaches for the Qualification of Electronic Noses Described in the Scientific Literature*

Other more or less similar approaches for e-nose performance testing have been proposed in the scientific literature. An example of approach that can be used for qualification of electronic noses to be used as environmental odour monitoring systems is given in References [24,66].

In more detail, the first example [24] proposes a testing procedure for performance evaluation aimed at the definition of minimum performance requirements referred to electronic noses to be used for environmental odour monitoring at receptors.

In this study, the following aspects were deemed important for qualifying an electronic nose for environmental odour monitoring:

• the capability of giving repeatable and reliable responses under variable atmospheric conditions; indeed, variations of temperature and humidity are particularly critical for e-nose environmental outdoor use ("invariability of responses to variable atmospheric conditions");

Then the study describes as example of detailed procedure adopted to test a commercial electronic nose towards those aspects. According to the proposed procedure, the abovementioned aspects shall be tested using standard test gases. The use of standard test gases is preferred over "real" environmental samples, because of the intention of guaranteeing repeatability and reproducibility of the procedure, in order to make it possible to compare the performances of different instruments tested in different labs or at different times and conditions.

The second example [66] proposes a procedure for electronic nose performance testing based on a similar approach. In this case, the instrument performances are evaluated through the definition of an experimental protocol, which is structured into different levels.

The paper focuses on the first two levels of testing, which involve specific tests with standard test gases, including for instance chemical compounds that are representative of common environmental odours. These specific tests allow for the evaluation of the electronic nose limit of detection, lower detection limit towards those target compounds and repeatability of responses to a given stimulus. This type of testing is not related to specific applications and therefore raw signals are considered instead of instrument output for performance evaluations. On the other hand, the third level of testing is related to the specific application, and thus the performance of the trained system is evaluated in the field in terms of classification accuracy. This third level of testing was not described in the paper [66].

The two abovementioned papers have in common that the proposed approach is based on the idea of verifying some fundamental aspects (e.g., lower detection limit, response repeatability, capability of compensating humidity and temperature variations, and capability to classify different odour types correctly) by performing performance testing using standard test gases.

It is important to highlight that since standard test gases are never fully representative of what happens in "real" environmental conditions, these tests shall not be considered as a sufficient condition to prove the electronic nose suitability for the specific application. What is described here is a sort of pre-qualification, which always needs to be followed by a validation in the field.

This is not very dissimilar from the approach proposed in the VDI 3518-3:2018; the main difference is that here no minimum performance requirements are fixed.

#### *5.2. Applicability of EN 14181:2014 and EN 15267:2009 to Electronic Noses*

#### 5.2.1. Electronic Nose as AMS?

As described in the previous section, the Technical Standards EN 14181:2014 and EN 15267:2009 define the quality protocols that must be applied to AMS, which in facts are systems for the continuous sampling, measurement and control of pollutants in emissions and ambient air.

In a very recent paper by Cipriano [44], the author proposed the possibility of implementing the features of the technical standards for AMSs to electronic noses, thereby focusing on qualification and maintenance, and on the uncertainty aspects.

Although electronic noses are not AMSs, and thus this proposal may sound as a provocation, the implementation of electronic noses for emission and ambient air monitoring purposes, as described in the previous section, the question arises of the opportunity to partially assimilate them to AMSs. A possible integration of such instruments into the universe of AMSs could be advantageous in order to define the uncertainties associated with their use as air quality monitoring tools.

Indeed, an AMS device is intended to be used for continuous legal use, so it must ensure reliability, integrity and data security. It shall allow the calculation of both the uncertainty on the measured values vs. a SRM, and an independent verification of its metrological capabilities.

The possibility to apply the principles of the standards for AMSs to the verification of these same aspects is discussed here, as proposed in Reference [44].

5.2.2. Reliability, Integrity and Data Security

In order to allow QAL1 and QAL2 evaluation, electronic noses shall declare the performances of the entire system (sensors, data acquisition, processing, interfaces, etc.) and permit their verification. Also, in order to allow QAL3 procedures, it is necessary to have specific hardware and software solutions to implement periodic checks of the sensors array (Figure 1).

**Figure 1.** Electronic nose with quality assurance (QA)/QC control of the sensor array.

One of the biggest advantages of electronic noses is their flexibility due to possible modification of the software, by using different algorithms and calibration data. Such flexibility can be very useful for obtaining a good QAL2, so that the instrument can be trained to give results very close to the calibration values. However, this is also one of the biggest problems during ongoing verifications, as every modification done after QAL2 process implies the formal invalidation of its performances. E-nose structure shall ensure that, when the calibration process is finished, all the relevant data and configurations are locked and encrypted in order to prevent performances changes and unauthorized modifications.

#### 5.2.3. Uncertainty

Nowadays all emission and ambient air monitors shall declare their "uncertainty budget" evaluated following principles of the ISO IEC Guide 98-3:2008 "Guide to expression of uncertainty in measurement" [67]. In this guide, the output of the measuring process is described by Equation (1) [44]:

$$y = f(\mathbf{x}\_1, \mathbf{x}\_2, \dots, \mathbf{x}\_n). \tag{1}$$

The estimation of the total uncertainty *u*(*y*) can be obtained by the propagation of the single uncertainty terms for each *xi*, *u(xi)*, by the means of Equation (2):

$$
\mu(y) = \sqrt{\sum \mathcal{C}\_i^2 \mu^2(x\_i)},
\tag{2}
$$

where *Ci* is the sensitivity coefficient of the single *xi* term, and *u(xi)* its uncertainty.

Function *f* shall cover all the measuring process, from the single sensor acquisition up to the final output, including calibrations, interferences, nonlinearities and software-related errors. For that, the use of a new CEN standard, the FprCEN/TS 17198:2017 "Stationary source emissions—Predictive Emission Monitoring Systems (PEMS)—Applicability, execution and quality assurance" could be useful. Such a standard is designed for emission prediction models, but could be easily adapted to electronic noses. Furthermore, its scope is to achieve conformity to EN 14181 and EN 15267 of software predictive systems and furnishes a simplified formula for uncertainty evaluation [44]:

$$u(y) = \sqrt{\left(\mu\_{model}^2 + \mu\_{input}^2 + \mu\_{other}^2\right)}\tag{3}$$

where *umodel* is the uncertainty of the mathematical model, *uinput* the uncertainty from sensors array, *uother* the uncertainty due to parameters not included in the model, evaluated by confrontation with independent odour measurements used to calibrate the electronic nose.

#### 5.2.4. Independent Verifications

An e-nose shall make it possible for the final user to verify its metrological capabilities, in order to validate its operational status and guarantee its calibrations are still aligned with QAL2 results. This means that, for instance, a specific gas matrix should be realised and used for periodic system validation.

#### **6. Discussion and Conclusions**

As stated in the introduction, this paper had the primary aim of discussing the need for the definition of quality protocols for electronic noses that are intended as air quality monitoring tools for the detection and the measurement of environmental odours. As a part of this discussion, this work also described some of the possible approaches that can be adopted for a standardization and qualification of such instruments, thereby reviewing the relevant documents—technical standards and scientific publications—on the issue.

In order to allow a better evaluation of these qualification approaches in terms of practical applicability to e-noses for environmental use, as well as the possible advantages and drawbacks related to their application, Table 1 schematizes the most relevant aspects relevant to the different approaches, together with their pros and cons.

Based on the information reported in Table 1, it is possible to appreciate the pros and cons of the different approaches. Leaving out the Dutch NTA 9055:2012, which is so generic that no real standardization/qualification approach is proposed, and the CEN TC/264 WG41 activity, which is still in progress and at a very preliminary stage, it is possible to make some more specific considerations about the other technical documents reviewed here.

The qualification approach based on the combination of the EN 14181:2014 and the EN 15267:2009 has the enormous advantage, compared to the others, that it is the only one validated. However, despite being a consolidated and effective approach, it is only valid (and validated) for monitoring systems that are not e-noses, and thus would need to be deeply re-adapted in order to make it applicable to odour measurement systems.

On the other hand, the other approaches, which are specific for e-noses and for odour measurement, to the best of our knowledge, are not validated yet. Therefore, up to now, there is no available data comparing how different instruments would perform by application of any of these testing protocols. The only data available in literature concern testing procedures that are not part of a standard [66,67].

**Table 1.**

*Cont.*

One aspect that emerges from the information reported in Table 1, is that two different concepts for standardization/qualification can be distinguished. The first one involves the definition of minimum performance requirements. Even though the establishment of minimum requirements may result in a higher level of guarantee for the end user, this may entail the drawback of being a bit premature for a technology that is still under development. This is particularly true in consideration that e-noses for environmental use can be applied in a variety of different situations and with different purposes, for which different instrumental performance characteristics may be required. For this reason, it might be better to have a more flexible approach involving instrument testing and performance declaration without the necessity to achieve a minimum performance level.

Thus, a possible alternative approach for a quality protocol regarding electronic noses to be used for environmental odour monitoring would be a combination of the concept of performance testing with the requirements of the quality standards for AMS. This would entail a method for performance testing and ongoing quality procedures (QAL3), based on the principle that the instrument manufacturer shall specify the electronic nose characteristics, metrological functionality, and design limits, without fixing minimum performances. Thus, it is the final user who evaluates the instrument suitability and makes the choice of the most appropriate electronic nose features according to his specific needs, or, for instance, to the current regulatory requirements.

This is basically the approach followed by the Italian UNI1605848 project, which, despite its lack of details regarding the testing procedure, is the only multi-level verification approach allowing for the characterization of the different aspects of the electronic nose functioning in every phase of the instrument's life and application.

This approach seems to guarantee higher flexibility, thus making it applicable also to prototypal solutions, which is an important aspect for a technology that is still at an initial stage as proper air quality monitoring tool and in continuous development.

As a conclusion, application of specific quality programs to electronic noses for air quality monitoring is a delicate and complicated process, not yet deeply explored, and where there are continuous implementations of both the instruments and the relevant technical standardisation. However, the current status of the technology seems to be sufficiently mature to undergo such a process, which is necessary in order to make electronic noses a widespread and effective environmental odour impact assessment tool.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

## *Review* **Use of Electronic Noses for Diagnosis of Digestive and Respiratory Diseases through the Breath**

#### **Carlos Sánchez 1,2, J. Pedro Santos 1 and Jesús Lozano 3,\***

Received: 12 January 2019; Accepted: 21 February 2019; Published: 28 February 2019

**Abstract:** The increased occurrence of chronic diseases related to lifestyle or environmental conditions may have a detrimental effect on long-term health if not diagnosed and controlled in time. For this reason, it is important to develop new noninvasive early diagnosis equipment that allows improvement of the current diagnostic methods. This, in turn, has led to an exponential development of technology applied to the medical sector, such as the electronic nose. In addition, the appearance of this type of technology has allowed the possibility of studying diseases from another point of view, such as through breath analysis. This paper presents a bibliographic review of past and recent studies, selecting those investigations in which a patient population was studied with electronic nose technology, in order to identify potential applications of this technology in the detection of respiratory and digestive diseases through the analysis of volatile organic compounds present in the breath.

**Keywords:** electronic nose; gas sensors; biomarkers; diseases; digestive system; respiratory system; volatile organic compounds; breath

#### **1. Introduction**

The relationship between aromas present in the breath and disease has been known by doctors for a several hundred years. The use of aromas to identify diseases dates back to the fourth century, when the doctor, based on experience, could determine what disease a patient was suffering. For example, a fruity aroma in the breath was identified as a sign of ketoacidosis associated with diabetes, and the smell of ammonia can be indicative of kidney failure. This method was not accurate, as it was necessary for the disease to be in an advanced stage to be detected by human olfaction. Based on this, electronic systems have been developed to diagnose diseases through the breath and to provide information about the state of the human body [1]. Human breath has a considerable amount of volatile organic compounds (VOCs) that are a product of metabolic activity. These VOCs can differ according to genetic or environmental factors such as age, weight, sex, lifestyle, or eating habits, and can influence the chemical composition of a person's breath, depending on the amount and concentrations of these compounds. Diseases can also cause an alteration of VOCs in the breath [2].

Currently, there has been an exponential development of technology in medical applications, in both the prevention and diagnosis or treatment of disorders. The growth in areas such as chemical sensors, microelectronic designs, material sciences, and artificial intelligence is contributing to the development of medical technology, and to improved health and increased life expectancy of the population. According to the World Health Organization (WHO), the average life expectancy in the world increased by five years in the period 2000–2016 [3]. The problem is that quality of life has not improved as much as life expectancy. The average increase in life years does not mean that this corresponds to a good state of health. People can suffer diseases and problems that cause a loss of

quality of life without endangering their lives. On the other hand, health inequalities persist among different countries and people at different income levels [4].

One of the biggest challenges of modern medicine is to develop equipment that allows early diagnosis in a noninvasive way. This could avoid disease exacerbation, permitting control of the evolution of chronic patients in the medium and long term and maintaining their quality of life. It also would contribute to a reduction in hospital costs, as it could reduce the number of hospital/health center visits and the amount of medicine used. The principal limitation of the early detection of disease is that it is impossible to analyze healthy persons continuously, because this would be economically infeasible. In addition, the health system does not have the resources or the medical staff for this purpose. For this reason, the devices should be cheap and portable, so they can be available to more hospitals and health centers. On the other hand, this would open the possibility for patients to have devices at home to perform regular analysis.

These arguments and necessities have motivated the development of new technologies such as the electronic nose. The first electronic nose was developed in 1964 by Wilkens et al. [5], but it was not until 1982 when the electronic nose as a system using chemical sensors to classify odors was described by Persaud et al. [6]. In the 1980s, there was an evolution of this technology, which led to the creation of a technological sector that tried to imitate the olfactory systems of mammals [7]. Since then, the development of electronic noses has gone hand-in-hand with the technology being used in various sectors and different applications [8]. One is the medical sector, particularly the diagnosis and control of respiratory and digestive diseases. This technology could offer the possibility of diagnosing or evaluating disease states in a noninvasive and quick way with low-cost instruments.

#### **2. The Olfactory Organ and Electronic Nose**

Living organisms receive information about the surrounding environment through different sensory organs. Nowadays, the scientific community is focusing on the development and generation of systems and devices that can mimic the sensory organs: first, to replace the function of one of these organs in case of malfunction due to atrophy, pathology, or accident; and second, to be used in a wide variety of industrial applications in fields such as medicine, agriculture, food, environment, etc. Given that a deficiency in the ability to smell does not limit a person's normal life, except for the detection of gas leaks or fires, this technology becomes increasingly important in other applications [9,10].

The electronic nose is capable of detecting, discriminating, and identifying different types of chemicals present in the headspace of a sample as VOCs. The device's response to smell is produced by the interaction between volatile compounds and sensors, whereas this function is done in the biological nose by the olfactory epithelium, which works as a transductor, as it generates electrical signals from chemical stimuli. These signals are preprocessed in the olfactory bulb, then transmitted to the brain, where they are stored. Finally, the data are used to identify odors in the learning stage. Analogously, in an electronic nose, an analogical–digital conversion is produced to preprocess the signals, with a microcontroller employed for this purpose. In this case, the data are stored in the database of a pattern recognition machine to identify the aromas that are learned. Figure 1 shows a comparison between the electronic nose and biological nose [10,11].

Due to the wide variety of applications that have been generated recently in several sectors, sensors based on different detection principles have been developed to satisfy the needs that have arisen. Sensors undergo a physical or chemical change by interacting at the molecular level when exposed to a gas. This process is reversible and allows the sensors to be used again in other tests [8].

**Figure 1.** Schematics of electronic nose and biological olfactory system [13].

This offers a wide range of possibilities, so it is possible to use one type of sensor or another according to the application [8,12]. For example, sensors with good sensitivity are MOS, MOSFET, and AGS. SAW, FL, and OF-LC combine sensibility with specificity. However, the principal disadvantages of the gas sensor are slow recovery (MOS), drift in the response (MOS, CP, SAW), low noise immunity (PID), and lack of reproducibility between sensors of different sets (CP, MOSFET, QCM, SAW) and in the response of the same sensor in the long term. Because of this, it is normal to use a matrix of sensors of different types to avoid the disadvantages presented by each one separately and maximize their advantages [12,14].

#### **3. Biomarkers and Diseases**

In 1899, Thomson, who was interested in measuring the mass/charge ratio of an electron, created the first instrument similar to a mass spectrometer (MC), improving on the work previously done by Wien. In 1941, Martin and Synge published a paper describing the liquid–liquid chromatographic partition. However, it was not until 1952 when chromatography took on its gas–solid version (James and Martin). In later years, equipment was developed that used both techniques, which eliminated the disadvantages that each one presented separately [15,16]. One of the main problems encountered in the analysis of VOCs was the impossibility of capturing these compounds, as they are very volatile. This inconvenience was solved with the emergence of solid-phase microextraction (SPME) [17]. Since then, new techniques have been developed, such as gas chromatography–mass spectrometry (GC-MS), proton transfer reaction–mass spectrometry (PTR-MS), selected ion flow tube–mass spectrometry (SIFT-MS), field asymmetric ion mobility spectrometry (FAIMS), time-of-flight mass spectrometry (TOF-MS), ion mobility spectrometry (IMS), liquid chromatography–mass spectrometry (LC-MS), and high-performance liquid chromatography (HPLC).

With these techniques, it is possible to extract information about the respiratory or digestive system through the breath. However, the existence of VOCs in respiration does not necessarily imply that these volatile molecules are produced by the human body; rather, they can be produced in an exogenous process. For example, acetonitrile is commonly found in the breath of smokers, occurring exogenously. Exposure to a dangerous atmosphere, contaminants, or even certain medications can generate new compounds and also alter the concentration of other endogenous compounds [18].

Table 1 shows descriptions of different processes associated with oxidative stress and airway inflammation and the influence of these processes on the composition of VOCs in the breath. Endogenous compounds are produced by respiratory or digestive system cells, which metabolize

the molecules present in the inspired air and generate others. When the cells function abnormally, they can alter the composition of VOCs in the breath. For example, oxidative stress is an imbalance of the normal state of the human body caused by the production of free radicals, which alters the composition of the compounds present in the breath as H2O2, CO, or 8-isoprostane [19,20]. Another example is the inflammation of airways, where the immune response produces an increase of various biomarkers in the breath [19,21].

\* Cytokines are agents responsible for cellular communication.

Microorganisms that may be present in the digestive or respiratory tract, such as viruses, bacteria, or fungi, can also directly or indirectly alter the composition and concentration of VOCs in the breath. These cannot modify the concentration by themselves; otherwise, these microorganisms can affect the normal functioning of the cells of the body, causing the same effect. Therefore, the alteration of volatile compounds can have an exogenous or endogenous cause, which can be produced by a malfunction of the cells or by external biological agents [18].

#### *3.1. Endogenous Biomarkers*

The following section describes the pathologies detected by different chemical techniques of analysis that are mentioned in the bibliography consulted. Asthma and chronic obstructive pulmonary disease (COPD) are chronic diseases that cause difficulty breathing due to inflammation of the airways, which become rigid [24,25]. Inflammation of the epithelial cells causes increased concentrations of NO, pentane, isoprene, and ethane in the expired air in asthma (Lärstad et al. [26]) and ethane in the expired air in COPD (Paredi et al. [27]) (Table 2). This occurs with other diseases described in the bibliography, such as acute respiratory distress syndrome (ARDS), cystic fibrosis (CF), and lung cancer. ARDS is a pathology that prevents enough oxygen from reaching the lungs and the

bloodstream [28]. CF is a genetic and chronic disorder that affects organs such as the pancreas, liver, kidneys, and intestine, causes difficulty breathing, and generates dense mucus in the airways [29]. A study by Antuni et al. [30] used NO and CO as biomarkers for the discrimination of this disease. In the case of NO, the concentration of this biomarker decreases in patients with CF in comparison to healthy individuals. In the case of CO, the concentration increases.

Finally, in lung cancer, the process of cell division is disrupted by the creation of new cells in an uncontrolled way and when they are not necessary, as well as allowing old or damaged cells to survive [31]. Bajtarevic et al. [32] employed isoprene, acetone, methanol, and benzene as biomarkers of lung cancer. The concentrations of these compounds decreased in patients with this disease in comparison with healthy persons. Table 2 shows chemical analysis techniques and biomarkers used to detect different diseases.

**Table 2.** Concentrations of biomarkers used in the detection of different diseases. NS, not stated; ppb, parts per billion; ppmv, parts per million by volume.

**Table 2.** *Cont.*

As already mentioned, diseases of the digestive system, such as diabetes mellitus, liver fibrosis, and hypolactasia, can alter the concentration of VOCs in the same way as respiratory diseases. Diabetes is a disease in which the body does not produce insulin or does not use it correctly [42]. Liver fibrosis is a disorder that causes decreased blood supply through the liver and produces the accumulation of scar tissue [43]. Das et al. [37] and Kearney et al. [39] used acetone as the major biomarker to detect both pathologies. The concentration of this compound increased in patients with these disorders in comparison with healthy persons. Lastly, hypolactasia is a deficiency of lactase (the enzyme that metabolizes lactose) in the intestinal mucosa, which causes this molecule to not be metabolized [44]. For the detection of this disorder, Alkhouri et al. [41] used hydrogen as a biomarker.

#### *3.2. Exogenous Biomarkers*

In this section, diseases generated by external pathogens, such as pneumonia and pulmonary tuberculosis, which produce exogenous volatile organic compounds, are described. Pneumonia is a lung infection that can be caused by many pathogenic agents, such as bacteria, viruses, or fungi [45], while tuberculosis is a bacterial infection (Mycobacterium tuberculosis) that mainly attacks the lungs or other parts of the organism [46]. However, it should be noted that there are diseases (e.g., cystic fibrosis and COPD) that predispose the patient to infections such as pneumonia [29]. Therefore, in this pathology it is common to find endogenous and exogenous VOCs in the breath. In order to determine whether a person is ill, it is necessary to know the aromatic profile of a healthy person. The concentrations of existing major compounds in the breath of healthy persons can be seen in Table 3.

**Table 3.** Major volatile organic compounds present in the breath of healthy individuals [47].

#### **4. Traditional Methods of Diagnosis**

Traditionally, chemical analysis techniques have not been used to diagnose respiratory or digestive diseases, as these are very expensive. Breath analysis may mitigate some of the disadvantages of conventional diagnostic tests. Additionally, it could complement conventional methods as a screening tool. The main disadvantage of traditional diagnostic tests is that there is usually a long wait to conduct the test. In addition, some are invasive tests, which may require irradiation of the patient or a surgical procedure. In other cases, the test result is not obtained immediately, as in the case of cultures.

Both invasive and noninvasive diagnostic techniques are currently used. For example, spirometry and fractional exhaled nitric oxide (FeNO) are noninvasive techniques used to determine the patient's lung capacity, mainly used to diagnose diseases such as asthma and COPD. The sweat test and sputum cultures are also employed as noninvasive techniques for the diagnosis of cystic fibrosis (described by Gibson and Cooke [48] in 1959) and respiratory infections, respectively.

#### **5. Recent Developments in Electronic Noses for the Diagnosis of Respiratory Diseases**

One of the major objectives in medicine today is to develop equipment and techniques to achieve early diagnosis of diseases. For this, it is necessary to develop devices that are portable, economical, and, above all, noninvasive, in order to reduce the risk to patients. In addition, this would contribute to decentralizing medical resources and facilitate access to these devices, reducing waiting lists. Until now, noninvasive tests that have been performed were based only on the chemical analysis of body fluids (feces, urine, and blood), but the possibility of analyzing the volatile compounds generated by the samples (as well as exhaled air) has not been explored.

Traditionally, chemical analysis techniques such as GC-MS and, to a lesser extent, the techniques described in Section 3 have been used to detect VOCs and the chemical composition of gaseous samples. With the evolution of gas sensors and the electronic nose in recent decades, they are beginning to be used as diagnostic methods. It is true that currently gas sensors are not as selective as chemical analysis equipment, but the electronic nose allows for much smaller and compact equipment, which offers a lot of possibilities.

This paper includes a summary of the specialized bibliography on the detection of respiratory and digestive diseases through the breath using an electronic nose (Table 4). It should be noted that the scientific research carried out in recent years is included here. Therefore, only the electronic noses used in such studies are described, with the intention that this paper may help to determine the current

state of research in the field and be a starting point for future studies. In the bibliography, it is possible to find some authors who have used commercial electronic noses and others who have developed their own equipment adapted to specific applications. It should be noted that the diagnostic tests employed are used to corroborate the results of the studies, but they do not have to be standard tests. Most of the authors described in this paper used a commercial electronic nose, the Cyranose 320, which has a matrix of 32 sensors of carbon black polymer. Only a few used a self-developed electronic nose optimized for a specific application.

Asthma is one of the respiratory diseases that affects the largest number of people in the world, based on more papers being found on this disease. The population studied by Dragonieri et al. [49,50] was 40 adult subjects, and they employed the Cyranose 320. In data processing, they used principal component analysis (PCA) as an alternative diagnostic test to corroborate the results obtained by using spirometry and FeNO. The other authors studying asthma used similar population sizes, and also used the Cyranose 320. They used spirometry or FeNO as a diagnostic test. While not all authors used the same techniques in the processing phase, they employed PCA and other techniques such as receiver operating characteristic (ROC) curve and neural network algorithms (ANNs). On the other hand, Brinkman et al. [51] studied a smaller population of subjects and performed the measurements using three electronic noses, two commercial (Cyranose 320, carbon black polymer sensors) and Tor Vergata Electronic Nose (QCM sensors; developed by the Sensors Group at the University of Rome Tor Vergata, along with the Departments of Electronic Engineering and Chemical Science and Technology), and one home-developed incorporating metal oxide semiconductor sensors.

In the case of COPD, a variety of scientific articles were found that used the same two commercial electronic noses as in the previous case: Cyranose 320 and Tor Vergata Electronic Nose. Dymerski et al. [52] used an electronic nose formed by commercial sensors from Figaro. In this case, the samples were generated in the laboratory, unlike in the other studies. To distinguish between patients and healthy individuals, Paredi et al. [35] employed three biomarkers: ethanol, CO, and NO, the concentrations of which in the exhaled air of patients with COPD increased compared to the healthy people. The literature found for lung cancer varies in terms of the type of cancer described; each author focused on the detection of one, studying populations of between 50 and 100 adult patients, using the commercial electronic noses that were used by other authors.

In the detection of pulmonary tuberculosis, a wider variety of authors were found to use different electronic noses with different types of sensors: commercial ones such as Cyranose 320, Aeonose, and Bloodhound BH1114, and home-developed equipment, such as Zelota et al. [53] using QCM, Bruins et al. [54] using metal oxide semiconductor sensors, and Fend et al. [55] using conducting polymer sensors.

For the rest of the pathologies shown in the table, fewer bibliographical references were found. Saasa et al. [56] discriminated between samples of patients with diabetes mellitus by detecting acetone. On the other hand, Schnabel et al. [57] stand out, discriminating between patients with pneumonia and healthy people. They studied a remarkable sample of 125 adult patients using a commercial electronic nose called DiagNose, which uses metal oxide sensors.

Biomedicalapplicationsdevelopedusingcommercialandexperimentalelectronic

**Table4.***Cont.*

#### **6. Future and Challenges**

Commercial electronic noses, used by most researchers, are designed for general use and are not optimized to detect the biomarkers of the diseases of interest. For medical applications, it is necessary to design specific sensors and optimized electronic noses, since it is an extremely complex sector, and it is a critical factor for the diagnosis and control of pathologies to be effective. In the medical sector, it is necessary to develop equipment that allows early diagnosis of diseases, enables simple and effective control, and is noninvasive.

On the one hand, this would allow health centers and small hospitals to have access to this type of equipment for the diagnosis and control of diseases (due to its lower cost) and thus minimize waiting lists. On the other hand, this would provide an opportunity for patients to have devices at home. In addition, in conjunction with the evolution of mobile technologies and Internet communication, it would allow patients to diagnose and control diseases from home by themselves or with remote medical supervision. It could even permit these types of sensors to be connected to phones either by incorporating them or through a gadget. The tendency of medicine is to advance along this line so that analyses and explorations can be performed at a distance. It is hospitals and health insurers themselves that could provide these types of devices to patients. In this way, it would be possible to achieve a unified protocol of measurement from the breath that is simple for patients to perform.

It is a promising technology for the diagnosis of respiratory and digestive diseases in a noninvasive way, since it allows the development of portable and cost-effective equipment. This is a key factor for this technology to succeed. However, current sensors still present complications that limit their operation. Therefore, one of the challenges from a technological point of view is to improve the selectivity of these sensors and their sensitivity to lower concentrations. To this end, it is important to use and develop new nanomaterials that can significantly improve selectivity and reduce the size and consumption, essential issues for miniaturization. It is therefore perhaps more important to focus efforts on developing technology that would enable these devices to be used in this application, and then study which is the ideal methodology to take samples in patients that is easy for a person without medical education to do.

#### **7. Conclusions**

Electronic nose technology has evolved remarkably in the last decade. Many researchers have focused their efforts on developing this type of equipment in a multitude of sectors, such as food, medicine, environment, and detection of hazardous materials.

This study presents a bibliographic review of research done on the use of the electronic nose for the detection of respiratory and digestive diseases through the breath. This technology bases its design and operation on the human sensory organ. The e-nose technology permits the study of diseases from a new approach, since until now analytical tests have focused exclusively on blood, urine, or feces samples, which allowed additional information to be obtained and studied. Although this technology is still in development and currently has some limitations, it has several advantages over other diagnostic methods. The ability to take measurements through the breath makes it possible to conduct analyses noninvasively, eliminating risks to the patient.

In addition, it would allow the production of portable equipment at a lower price than other equipment available in the market, giving health centers and hospitals greater access to this equipment. With the evolution in recent years and current development (improvement and miniaturization of gas sensors), the electronic nose is presented as a promising technology that should contribute to improving the quality of life of patients with chronic pathologies as well as early diagnosis of different diseases, contributing to the reduction of direct and indirect costs of the health system.

The purpose of this paper is to demonstrate the viability of this technology through a large number of studies done in this regard. From now on, it will be necessary to improve and solve the problems presented by the e-nose, fine-tuning this technology for the different applications in which it is intended to be used. This will require a great effort on the part of the research community, but once these problems have been solved, this technology should result in a great advance in the control and diagnosis of diseases, and satisfy many other needs that currently exist in the medical sector.

**Funding:** This research was funded by Department of Education and Research of the Madrid Autonomous Community (Spain) grant number IND2017/TIC7714.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **Abbreviations**

#### **References**

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

## *Review* **Use of Electronic Noses in Seawater Quality Monitoring: A Systematic Review**

#### **Alessandro Tonacci \*, Francesco Sansone, Raffaele Conte and Claudio Domenici**

National Research Council of Italy, Institute of Clinical Physiology (CNR-IFC), Via Moruzzi 1, 56124 Pisa, Italy<EMAIL_ADDRESS>(F.S<EMAIL_ADDRESS>(R.C<EMAIL_ADDRESS>(C.D.)

**\*** Correspondence<EMAIL_ADDRESS>Tel.: +39-050-315-2175

Received: 29 October 2018; Accepted: 19 November 2018; Published: 23 November 2018

**Abstract:** Electronic nose (eNose) systems are particularly appreciated for their portability, usability, relative low cost, and real-time or near real-time response. Their application finds space in several domains, including environmental monitoring. Within this field, marine monitoring is of particular scientific relevance due to the fragility of this specific environment, daily threatened by human activities that can potentially bring to catastrophic and irreversible consequences on marine wildlife. Under such considerations, a systematic review, complying with the PRISMA guidelines, was conducted covering the period up to 15 October 2018, in PubMed, ScienceDirect, and Google Scholar. Despite the relatively low number of articles published on this specific topic and the heterogeneity of the technological approaches employed, the results obtained by the various groups highlight the positive contribution eNose has given and can provide in near future for the monitoring and safeguarding of this delicate environment.

**Keywords:** chemical sensors; eNose; environmental monitoring; seawater; sensors; volatile organic compounds

## **1. Introduction**

Anthropogenic activities produce every day large quantities of pollutants that are discharged in the environment [1,2], including oceans, and in the seawater at large. Such discharges, together with the exploitation of the sea resources nowadays represent a significant threat for the marine environment and cause a continuous, unprecedented degradation of oceans, seas and coastal areas [3–5]. The monitoring of such compounds is a pivotal action when aiming at promoting the preservation of marine and coastal areas and the sustainability of the ecosystem, at large [6].

To this extent, the approaches commonly used are based on traditional water and air quality evaluation methods, including analytical tools that, however, commonly operate at laboratory settings, and are therefore (i) unable to provide real-time or near real-time results, and (ii) possibly being associated with the occurrence of sample degradation during transportation from the sampling site to the laboratory bench.

To properly overtake such bottlenecks, portable devices can represent a smart, mostly robust, easy-to-use, low-cost alternative, capable of bringing fast, reliable, reproducible responses, without the need for sample transportation and, likely, consequent degradation.

Among such systems, electronic nose (eNose)-like tools are emerging as a popular alternative, maximizing the above mentioned advantages and keeping, at the same time, low drawbacks.

The first evidences for eNose systems dates back to the early 1980s, when Persaud and Dodd [7] first attempted mimicking the functioning of the mammalian olfactory system by means of an "electronic nose". Since then, a plethora of systems has been realized for several different applications, including biomedical and diagnostic, environmental, as well as food industry/food quality usage [8,9].

Under such considerations, here we present a systematic review to investigate the use of eNose systems within a specific field of application, represented by the monitoring of seawater pollution. To this extent, we will first outline the search strategy adopted for this work, then we will present the results obtained, critically discussing such evidences, and highlighting pros and cons of the single approaches.

#### **2. Materials and Methods**

#### *Search Strategy*

A systematic review of the literature, covering the period up to 15 October 2018, was conducted in PubMed, ScienceDirect, and Google Scholar database, according to the PRISMA guidelines [10]. The search strategy was as follows: ("eNose" or "electronic nose") and ("seawater pollution" or "seawater monitoring" or "seawater" or "pollution"). The search was limited to research articles published in English language in peer-reviewed journals or in international conferences' proceedings. After having discarded multiple hits, the results obtained were sorted by relevance and the most significant works dealing with seawater monitoring by means of eNose systems were selected (as displayed in Figure 1). Given the relatively low number of systems specifically designed for seawater monitoring, some tools designed for water monitoring in general, but adaptable to marine monitoring, were also included and critically discussed.

**Figure 1.** Flowchart for literature review.

#### **3. Results**

This systematic review retrieved a handful articles directly dealing with seawater monitoring by eNose systems. The articles taken into account are displayed in Table 1.

**Table 1.** Study selected (AUV: autonomous underwater vehicle; FGC: fast gas chromatography; GC-MS: gas chromatography—mass spectrometry; GSM: geosmin;InternetofThings;MIB:2-methylisoborneol;MOS:metaloxidesemiconductor;ORs:olfactoryreceptors;piD:photo-ionizationdetector(s);

Specifically focusing on the results reported in Table 1, one of the first studies performed in this field was conducted by Bourgeois and colleagues [11] in the UK. The work described the development and use of an eNose system for monitoring water and wastewater, potentially usable also for seawater assessment. The first prototype was based on an eNose5000 system consisting of an array of 12 conducting polymers, interfaced with a sampling vessel and with a PC for data analysis. Aside from this laboratory prototype, a second system was realized, with a ProSAT sensor array with eight conducting polymers as the sensing system, a PC for data storage and a data transfer system. This second system was tested on-field at the Cranfield University Sewage Works facilities and, through a proper conditioning in a temperature-controlling environment, has been demonstrated to be able to reduce humidity variations, at the same time improving reproducibility. The system was capable of detecting the presence of low concentrations (around 5 ppm) of 2-chlorophenol, with optimal performances at 25.5 ◦C of temperature, 170 mL/min of flow, and 0 of porosity.

As widely known, hydrocarbons are among the major pollutants of the marine environment, with oil spills occurring throughout the seas and the oceans as a consequence of marine accidents, petroleum pipeline leakages and fraudulent events. Therefore, systems capable of specifically assessing the presence of oil spills on the sea surface can be critical to reduce the environmental impact of such discharges in marine and coastal environment.

Under such consideration, Tzing and colleagues [12] developed a system, based on the eNose technology, to identify the source of spilled oil in an accident site, comparing it with a "gold standard" system composed by a gas chromatography—mass spectrometry (GC-MS) device that served to ensure the correct classification of the source site.

The system developed was composed of a Cyranose 320 eNose system, composed of an array of 32 polymer sensors, a pump for air inlet and purge, as well as a PC equipped with a dedicated software for data acquisition and analysis. Together with this system, a so-called z-Nose system was employed, constituted by an Electronic Sensor Technology FGC/SAW 7100 eNose, equipped with a pre-concentrating trap containing Tenax absorbent, a short gas chromatography (GC) column, a pneumatic control, a fast SAW detector, as well as a programmable gate array microprocessor. As for reference analysis, a Varian CP-3800 GC connected to a Saturn 2000 ion-trap mass spectrometer (MS) was used.

Spilled oil samples were collected from surface water at the site of an accident, whereas a set of known compounds, including gasoline, jet fuel, diesel, and fuel oil were also collected, properly stored and transported to the laboratory, representing the reference samples for the eNose (1 mL of oil loaded into a 30 mL vial, equilibrated at ambient condition for 5 min).

By means of a PCA, the eNose classified the unknown stimulus as "jet fuel", as further agreed by zNose and GC-MS systems, demonstrating the good classification capability of the eNose system. However, the main limitation of this promising approach resides in the fact that the test set employed was represented by only one sample that, despite the high internal consistency of the eNose response, could lead to misleading conclusions.

A couple of years later, Goschnik et al. [13] employed the semiconductor-based eNose system KAMINA to analyze water samples polluted with chloroform (chloro-organic solvents-like) or ammonia (fecal contamination-like). Despite this specific trial was mainly dealing with wastewaters, this application could have had the potentialities to be translated into a system usable for seawater monitoring, therefore, it was added to the present investigation. The trials were performed with a flow rate of 500 mL/min and an entrance temperature of 20 ◦C, subsequently increased up to 240 ◦C and 300 ◦C at the two microarray ends, respectively. The results obtained for this investigation, showing a detection limit below 1 ppm, demonstrated the usefulness of the KAMINA system for this purpose, however revealing a strong need for building up a populated, reliable reference database to support the correct classification by the eNose system. The absence of such a support would dramatically decrease the performances of the system, as frequently occurs with eNose approaches.

A very interesting approach was proposed by Lozano et al. in 2014 [14] and updated a few years later [15]. There, the authors proposed a portable eNose system equipped with an IEEE 802.11 transceiver for wireless communication in order to be ready for inclusion within a wide eNose network for distributed measurements. The system developed, also featuring an electronic pump and valve, as well as embedded electronics and rechargeable batteries, was able to work with several different resistive microsensors. By performing measurements on cycles of 60 s of adsorption and 540 s of desorption by means of an headspace sampling system, the prototype presented was demonstrated to be able to reliably discriminate between pollutants (especially when a relatively low number of compounds are taken into account) using principal component analysis (PCA) and a few different artificial neural networks (ANNs). More specifically, the compounds used to evaluate the response of the system included: blank water, acetone, toluene, ammonia, formaldehyde, hydrogen peroxide, ethanol, benzene, dichloromethane, acetic acid, xylene and dimethylacetamide. The correct discrimination occurred in more than 90% of samples in most cases, with concentrations of the various compounds of around 1 mL in a 20 mL vial at 16 ◦C constant temperature.

Specifically dealing with seawater pollution, the work published by Tonacci and colleagues [16] described the implementation of an eNose system to be used within an autonomous underwater vehicle (AUV) for monitoring oil leakages in a marine protected area, including the Tuscan Archipelago and Cetacean Sanctuary, North Mediterranean. The system implemented was composed by a sensing part, featuring Photo-Ionization Detectors (piD), a system for air inlet and purge made up of an electronic pump and a valve, and a smart embedded electronics, based on an Arduino Mega 2560 board. A PC for data acquisition was also foreseen, and ANNs of the type Kohonen self-organizing map (KSOM) were implemented and trained in order to: (i) recognize the level of pollution, independently from its source (e.g., high, medium, low level), and (ii) identify the substance detected within a set of known compounds (e.g., crude oil, diesel fuel, gasoline, jet fuel). Despite the good accuracy of classifiers, especially for the first KSOM (around 74% of correct classification for the first KSOM, 67% for the second one, and a fast response (below 20 s), with pollutants' concentration of 10−3 vol/vol), the system was seen to provide not reliable response when exposed to relative humidity (RH) above 70%, thus representing a noteworthy limitation of the tool described. Two further expansions of this application were also presented by the same group. The first one adopted the same approach including the eNose system within a moored buoy in order to combine dynamic (AUV) and static (buoy) monitoring of a given marine area [19], whereas the second one scaled the payload system in order to make room for future inclusion of such system within a real-time context of distributed monitoring [17].

A novel approach to the problem was adopted by Son and colleagues in 2015 [18].  The system functioning was assessed to distinguish the presence of geosmin and 2-methylisoborneol, compounds produced by bacteria and reliable indicators of contamination in the water supply system. The approach adopted can be tailored upon the users' needs also to investigate the presence of contaminants in seawater. The system was demonstrated to be reliable in detecting the proposed stimuli at concentration as low as 10 ng/L, with the possibility to assess the presence of both compounds at the same time, thanks to the specific binding between hOR51S1 and hOR3A4 and geosmin and 2-methylisoborneol, respectively. Very importantly, the correct classification of the eNose system does not require a specific pre-treatment of water samples, allowing the use of this solution for rapid monitoring of water quality. The binding between other olfactory receptors and specific pollutants could enable applying the solution to other, customized applications and experimental settings.

Recent technological advances allowed researchers to explore new solutions in this specific field. Internet of Things (IoT), for example, represents a landmark revolution in the field of sensing and bio-sensing. This promising and extremely actual approach was employed by Climent and colleagues [20], which developed a low-cost, portable eNose, named Multisensory Odor Olfactory System (MOOSY4), for water quality assessment purposes. In that work, the MOOSY4 was employed for the quality control of bottled water, but can be also used for seawater monitoring. The prototype was composed of four metal oxide semiconductor (MOS) gas sensors, suitable for the IoT technology, with a system architecture featuring data acquisition, storage, processing and user interfacing parts. The volatiles monitored, dimethyl disulphide, dimethyl diselenide, sulphur, were correctly recognized in 86% of cases, making the system extremely suitable for the purpose it was conceived for, and potentially applicable to a wide range of volatiles monitoring.

As hydrocarbons are, as reported, the main source of marine environmental pollution, specific approaches have been recently adopted tailored at this peculiar domain. For example, Aliaño-González and colleagues [21] used an AlphaMOS eNose combined with chemometrics to identify and discriminate 444 samples composed 40 μL of different petroleum-derived products (PDPs), including gasoline, diesel and paraffin, poured on a support and subjected to a natural weathering process by evaporation for one month. Taking advantage of the use of the linear discriminant analysis allowed the scientists to perform a correct discrimination of presence/absence of PDP in all cases and of the different PDPs in 97.7% of samples.

#### **4. Discussion**

The problem of environmental pollution is central in today's society, and full attention is paid by both National and International Environmental Organisms, as well as researchers, in finding solutions that can properly respond to the pressing necessity of environmental safeguarding. The extremely fragile ecosystem represented by marine and coastal areas is continuously threatened by manmade activities (e.g., ship transits, industrial discharges, etc.) that, in the medium- to long-term, can result in serious and irreversible consequences for marine areas and their wildlife [22,23].

To this extent, several initiatives, including national and international research projects, task-forces, position papers, and regulations have been promoted in last decades, many of which highlighting the need for tailored interventions both in a more preventive sense (monitoring of oil spills, etc.) and concerning accident remediation [24]. On the edge between these two phases, a number of smart monitoring systems have been developed in many research centers, to ensure a proper monitoring of a given area and, at the same time, allowing the possibility to trigger eventual alarms when the concentration of pollutants or, more specifically, of a single pollutant, exceeds a given safety threshold. Such solutions are especially useful in marine protected areas, where even the detection of very low levels of pollutants could enable the competent authorities to undergo quick, effective countermeasures, often preventing real environmental disasters. Within this framework, the use of low-cost, sensitive tools, ensuring a fast, real-time, or near real-time response is of particular interest, and this fact paved the way for the employment of eNose systems for this purpose.

In the last years, several prototypes and products based on this technology have been developed and employed for seawater monitoring, using different technologies as time flowed.

Among the most widely used technologies, especially in the first works published, are conducting polymers [11,12], providing good results, but in some cases obtained on a small number of trials or compounds. However, it is worth noting that such sensors are largely affected by environmental conditions, that could dramatically impact on the recognition performances of the tool, overall. The experimental setting is also pivotal with this approach, since the sensors should often be carefully cleaned after each measurement, reducing their applicability in a real environmental setting for continuous measurements unless a coupled purging device is used.

Semiconductors, which are the basis of most eNose systems developed to date, were also employed several times to this extent [13,20,21] and, considering the timeline of existing publications on the argument, are probably the most actual approach to the problem. Some contrasting results are present under this technology (e.g., considering the limits highlighted by the work by Goschnik and colleagues), however, the use of such devices probably guarantees the best performances, to date. The main limitation identified under this approach is the need for a well populated reference database to train the related recognition algorithms, but with the technological developments of recent times the issue was, in most cases, successfully resolved.

Other approaches retrieved in this domain included the use of photo-ionization detectors [16,17,19], featuring very fast response times, however displaying significant issues with high humidity environments and being characterized by high costs, and resistive detectors like the ones employed by Lozano and colleagues [14] and Herrero and colleagues [15]. The last solution mentioned is characterized by the strong need to control environmental conditions for the signal stability, a common problem with most of the other technologies normally employed in this specific field.

Additionally, promising hybrid approaches including a biological part, consisting of human olfactory receptors, as well as a technological part, were successfully employed by Son and colleagues [18], possibly opening the way for new explorations to reduce the impact of one of the most known issues of the eNose systems, that is non-specificity. In fact, thanks to the selective bond between compound and receptor, this approach could enable the detection of specific compounds, even though the reversibility of this bond remains questionable. The provision of such devices for new, promising technologies, including the IoT, as already experimented by Climent et al. [20], could enable new perspectives, at large in environmental, and specifically in seawater, monitoring.

**Author Contributions:** A.T. defined the rationale of the work, performed the literature search, critically revised the selected articles, and wrote the manuscript; F.S. participated in the bias and quality assessment, discussed the results obtained, and critically revised the manuscript; R.C. participated in the bias and quality assessment, discussed the results obtained, and critically revised the manuscript; and C.D. participated in the definition of the work rationale, discussed the results obtained and critically revised the manuscript. All the authors approved the final version of the manuscript.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**

© 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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**Luigi Sirleto \* and Maria Antonietta Ferrara**

National Research Council (CNR), Institute of Applied Sciences and Intelligent Systems, Via Pietro Castellino 111, 80131 Naples, Italy<EMAIL_ADDRESS>
**\*** Correspondence<EMAIL_ADDRESS>
Received: 10 January 2020; Accepted: 24 February 2020; Published: 26 February 2020

**Abstract:** Nowadays, in fiber optic communications the growing demand in terms of transmission capacity has been fulfilling the entire spectral band of the erbium-doped fiber amplifiers (EDFAs). This dramatic increase in bandwidth rules out the use of EDFAs, leaving fiber Raman amplifiers (FRAs) as the key devices for future amplification requirements. On the other hand, in the field of high-power fiber lasers, a very attractive option is provided by fiber Raman lasers (FRLs), due to their high output power, high efficiency and broad gain bandwidth, covering almost the entire near-infrared region. This paper reviews the challenges, achievements and perspectives of both fiber Raman amplifier and fiber Raman laser. They are enabling technologies for implementation of high-capacity optical communication systems and for the realization of high power fiber lasers, respectively.

**Keywords:** stimulated raman scattering; fiber optics; amplifiers; lasers; optical communication systems


**Coatings Tribology**

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