Author’s Accepted Manuscript Flow injection analysis of organic peroxide Explosives using acid degradation and chemiluminescent detection of released Hydrogen peroxide Parvez Mahbub, Philip Zakaria, Rosanne Guijt, Mirek Macka, Greg Dicinoski, Michael Breadmore, Pavel N. Nesterenko
PII: DOI: Reference:
www.elsevier.com/locate/talanta
S0039-9140(15)30007-2 http://dx.doi.org/10.1016/j.talanta.2015.05.052 TAL15644
To appear in: Talanta Received date: 8 April 2015 Revised date: 12 May 2015 Accepted date: 22 May 2015 Cite this article as: Parvez Mahbub, Philip Zakaria, Rosanne Guijt, Mirek Macka, Greg Dicinoski, Michael Breadmore and Pavel N. Nesterenko, Flow injection analysis of organic peroxide Explosives using acid degradation and chemiluminescent detection of released Hydrogen peroxide, Talanta, http://dx.doi.org/10.1016/j.talanta.2015.05.052 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting galley proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
FLOW INJECTION ANALYSIS OF ORGANIC PEROXIDE EXPLOSIVES USING ACID DEGRADATION AND CHEMILUMINESCENT DETECTION OF RELEASED HYDROGEN PEROXIDE Parvez Mahbuba, Philip Zakariaa, Rosanne Guijtb, Mirek Mackaa, Greg Dicinoskia, Michael Breadmorea, Pavel N. Nesterenkoa* a
Australian Centre for Research on Separation Science, School of Physical Sciences,
University of Tasmania b
Pharmacy School of Medicine, Australian Centre for Research on Separation Science,
University of Tasmania * * - Corresponding author: Professor Pavel N. Nesterenko Australian Centre for Research on Separation Science, School of Chemistry, University of Tasmania, Private Bag 75, Hobart 7001, Australia Tel: +61 (03) 6226 2165 Fax +61 (03) 6226 2858 Email: [email protected]
Keywords: organic peroxides, explosives, flow injection analysis, acid degradation, chemiluminescence
Abstract The applicability of acid degradation of organic peroxides into hydrogen peroxide in a pneumatically driven flow injection system with chemiluminescence reaction with luminol and Cu2+ as a catalyst (FIA-CL) was investigated for the fast and sensitive detection of organic peroxide explosives (OPEs). The target OPEs included hexamethylene triperoxide diamine (HMTD), triacetone triperoxide (TATP) and methylethyl ketone peroxide (MEKP). Under optimised conditions maximum degradations of 70% and 54% for TATP and HMTD, respectively were achieved at 162 µLǜmin-1, and 9% degradation for MEKP at 180 µLǜmin-1. Flow rates were precisely controlled in this single source pneumatic pressure driven multichannel FIA system by model experiments on mixing of easily detectable component solutions. The linear range for detection of TATP, HMTD and H2O2 was 1-200 µM (r2= 0.98~0.99) at both flow rates, while that for MEKP was 20-200 µM (r2=0.97) at 180 µLǜmin1
. The detection limits (LODs) obtained were 0.5 µM for TATP, HMTD and H2O2 and 10
µM for MEKP. The detection times varied from 1.5 to 3 min in this FIA-CL system. Whilst the LOD for H2O2 was comparable with those reported by other investigators, the LODs and analysis times for TATP and HMTD were superior, and significantly, this is the first time the detection of MEKP has been reported by FIA-CL.
1. Introduction The simplicity of in-house preparation of the organic peroxide explosives (OPEs), such as hexamethylene triperoxide diamine (HMTD), triacetone triperoxide (TATP) and methylethyl ketone peroxide (MEKP), from readily available materials was the main reason of their use in some recent terrorists attacks [1]. So, there is a strong demand for the development of fast, simple and sensitive methods of their identification and quantitative determination [2]. This
task is not trivial because of high volatility and absence of chromophoric groups in the molecules of OPEs. For these reasons the use of common analytical methods such as GC or HPLC with UV/visible detection is not readily suitable for their direct determination, so the application of more complex hyphenated techniques, typically involving mass spectrometry (MS), is required. Schulte-Ladbeck et al. [3] proposed RP HPLC with on-line Fourier transfom infrared (FTIR) detection for direct determination of TATP and HMTD. De Tata et al. [4] reported the application of RP HPLC with quadrupole time-of-flight mass spectrometry (HPLC-QToF-MS) for direct determination of various OPEs. However, micellar electrokinetic chromatography with UV detection was employed by Johns et al. [5] recently for separation of OPEs including HMTD and TATP in post blast scenario without any marked improvement compared to the hyphenated HPLC methods in terms of sensitivity. Alternatively, the determination of OPEs can be based on electrochemical [6], fluorescent [7] and chemiluminescent [8] detection of hydrogen peroxide as the main degradation product of OPEs. It should be noted that decomposition of one OPE molecule can result in more than one molecule of H2O2, so in case of 100% degradation a magnified analytical response can be expected. Because of its robustness, sensitivity and simplicity of integration with FIA, chemiluminescent detection (FIA-CL) is one of the most popular techniques for the on-line detection of hydrogen peroxide. The application of FIA-CL has been reported for the determination of H2O2 in natural waters [9], rainwater [10] and seawater [11]. The use of chemiluminescent detection of H2O2 after decomposition of TATP and HMTD was reported for determination of these explosives by a number of research groups [3, 6 and 12]. The analysis times reported varied from 5 to 12 min with the sensitivity ranging from 2.5 µM for TATP [6] to 500 µM for HMTD [3]. Obviously, the crucial parameters in the flow-through methods of analysis of explosives via detection of hydrogen peroxide are the velocity and conversion degree of OPEs into H2O2.
According to the literature data, the degradation of peroxide explosives can be accomplished enzymatically [12, 13], photochemically [14] or by using mineral acids [15]. The latter option appears more simple and robust, but it takes almost 12 hours for complete decomposition of TATP and HMTD which is not suitable for rapid screening. Additionally, there has been no report of the decomposition of methylethyl ketone peroxide (MEKP) - listed as a priority explosive by several government agencies including US National Counterterrorism Centre and the Australian Army [16]. It should be noted that 100% degradation of OPEs does not mean the maximum possible concentration of hydrogen peroxide suitable for detection, as hydrolysis of H2O2 occurs together with organic peroxides. On this reason, acid degradation conditions of OPEs should be carefully attuned to provide fast and sensitive response in FIA system. This study was a part of a larger project directed on construction of portable automated system for the fast screening of samples on presence of traces of homemade explosives including organic peroxides and inorganic explosives. According to the proposed design the whole system composed of three separate units, namely, sample extraction unit, FIA-CL detection unit for organic peroxides and capillary zone electrophoresis unit for parallel profiling of inorganic anions contents. The high degree of automation, minimal consumption of reagents and robust operation for extended period of time were considered as important requirements for this system. The system should provide either positive or negative answer in a short time on the presence of explosives at low concentration level. Typically, multicomponent FIA analysers use multi-channel peristaltic pumps for delivery of sample and reagents solutions. According to Dasgupta to get very low flow rates at µL/min level, as required in this study, peristaltic pumps are of little help [17]. Piezoelectric pumps, pneumatic pressure or gravity driven systems are often preferred choice in this case [18]. Pressure driven FIA instruments have advantages over systems using peristaltic pumps in
terms of lower signal/noise ratio, better reproducibility of timing and expanded possibilities for the use of aggressive solvents and carriers [18]. According to Valcarcel and De Castro, the main drawback of pneumatic driven instruments is connected with difficulty in accurate control of the flow rates in multiple channel FIA systems due to complex changes in hydraulic resistances resulting from different channel geometry and reagent viscosity [19]. In this study, a special attention was paid to the development of a control mechanism of flow rates in separate lines of multichannel FIA system driven by pneumatic pressure from a single source. The objective of this study is to optimise conditions for acid degradation of organic peroxides and subsequent chemiluminescent luminol based detection of hydrogen peroxide within pressure driven FIA-CL analytical unit for the purpose of fast qualitative determination of HMTD, TATP and MEKP. The target total analysis time was less than 2 min for rapid screening and high-throughput analysis, imposing a significant challenge to the optimisation of the flow conditions.
2. Experimental Section 2.1. Reagents and Chemical Standards Hydrated copper sulfate (CuSO4ǜ5H2O), 32% concentrated hydrochloric acid (HCl), 30% (w/w) reagent grade H2O2, isopropanol and sodium hydroxide (NaOH) pellets were purchased from Sigma-Aldrich (Sydney, Australia). Luminol was purchased from Fluka (Sydney, Australia). Element free deionised water was used to prepare all stock and working solutions. The TATP standard (10,000 mg L-1, 99.9% single component) and HMTD standard (5,000 mg L-1, 98.4% single component) were procured from Accustandard, USA. The MEKP standard (10,000 mg L-1) was supplied by the Australian Defence Science and Technology Organisation (DSTO).
2.2. Preparation of precise assay of the standard solution for H2O2 A stock solution of approximately 1000 mg L-1 of the 30% H2O2 was prepared through serial dilution. Then 50 mL of stock solution was transferred into a 500 mL conical flask, diluted with 200 mL of deionised water, and then 30 mL of 25% sulfuric acid was added. The solution was titrated with a standard 0.02 M potassium permanganate solution until the colour changed to pink. The working solutions were prepared by further dilution of stock solution in DIW.
2.3. Instrumentation A FIA-Cl system consisting of a low pressure Cheminert 6 port 2 position injector valve (C22-3186EH-FL, VICI, Houston USA), five SMC precision pressure regulators (IR 100001, SMC, Japan) and a Hamamatsu photomultiplier (10493-001, Hamamatsu, Japan) were used. A schematic of the instrument used for acid degradation study of TATP, HMTD and MEKP is illustrated in Figure 1. The system comprised SMC pressure regulators connected through a manifold to an external compressed air supply. Each regulator was used to control the reagent solution flow from a 500 mL glass bottle (Schott AG, Sigma-Aldrich, Australia) by applying pressures ranging from 0.01-0.2 MPa (1.5 psi to 29 psi). The pneumatic lines consisted of polyurethane tubing (2.5 mm I.D., SMC, Japan) and the hydraulic lines consisted of FEP tubing (0.203 mm I.D., Upchurch, USA). The 100 µL sample plug was carried into the Cheminert mixer (CM1XKF, VICI, Houston, USA) in 50:50 v/v deionised water - isopropanol where it was mixed with 32% HCl. Isopropanol was used to ensure complete dissolution of OPEs from the collected and extracted samples (the exact procedure is not included in this paper). The OPE containing
samples were degraded in acidic mixture in a 1 m PTFE knitted tubing coil reactor (0.25 mm I.D., 49.1 µL internal volume, Biotech AB, Onsala, Sweden) resulting in the release of H2O2 molecules. The excess HCl was neutralised by the addition of 18% NaOH and the resulting solution was then mixed with the luminol – Cu2+ reagent in a tee mixer (P-712, 2.9 µL swept volume, Upchurch, Oak Harbor, USA). The subsequent chemiluminescence reaction was detected with the photomultiplier tube. The chemiluminescence flow cell was fabricated inhouse using FEP tubing (0.508 mm I.D., 1548L, Upchurch, USA) with the total volume of the flow cell being 85.32 µL. The chemiluminescence signal was acquired by a Powerchrom data acquisition system (ER280, EDaq, Sydney, Australia) with proprietary software version 8.1. The pH of the effluent was monitored by an in-line pH monitoring flowcell (ColeParmer, Australia, vertical flow glass electrode 50 L internal volume) as shown in Figure 1. Insert Figure 1 3. Results and Discussions 3.1.Optimisation of reagent concentrations The crucial part of FIA-CL under development is acid degradation of OPEs in isopropanolwater (50:50) extracts of swabs used for sample collection from different surfaces. It was shown that such mixture provides the best extraction of various types of inorganic and organic explosives [20]. However, as a result of elevated viscosity of this mixture the FIA system in this study need to operate at pressures higher than those normally provided by peristaltic pumps. Also, initial experiments using peristaltic pumps demonstrated poor stability of the silicone tubing when in contact with solutions of isopropanol, hydrochloric acid and sodium hydroxide requiring the frequent replacement of the tubing and leading to poor reproducibility of data. For this reason a pneumatic FIA-CL system consisting of five
pressure lines equipped with precision low-pressure regulators (maximum operating pressure 0.2 MPa) was constructed as shown in Fig. 1. The acid fumes and the heat generated from the reaction between concentrated HCl and NaOH was a practical challenge in selecting the acid and base concentrations in this study. Additionally, the solubility of NaCl, formed as a product of the neutralisation reaction of HCl and NaOH, in isopropanol is very low (0.013 gǜg-1 at 23.5 C). To avoid frequent regulator malfunction and system blockage due to NaCl precipitation, 50:50 v/v deionised water isopropanol mixture was used to carry the 100 µL sample plug into the cheminert mixing chamber. The system operated optimally without blockage from NaCl precipitation and system malfunction from acid fumes and excessive heat when 32% HCl (concentrated) and 18% NaOH (w/v) were used. 3.2. Optimisation of the operating pressures and calculation of individual reagent flow rates The optimisation of reagent concentrations and optimum flow rates influencing degradation degree of OPEs and maximum response of chemiluminescent detection is not trivial task in multichannel FIA system with pressure driven flows. This is due to difficulties in control of flow rate in separate lines of pressure system, when changes in backpressure/flow rate in one line may effect on flow rates in other lines. Therefore, the exact concentration of used reagents should be measured in a separate experiment. Initially the system was investigated to establish the optimum input pressure at the regulators to provide the required flow rates with the least variance. The mass flow rates of deionised water (DIW) in all five lines were determined at 0.03, 0.07, 0.1, 0.15 and 0.18 MPa by precise measuring the weight losses in the containers after 8 min operation. The experiments were repeated for 5 consecutive days. The relative standard deviations (RSD) of the flow
rates in five lines ranged from 25% to 51% for 0.4 MPa supply pressure whilst RSDs ranged from 15% to 30% for 0.6 MPa supply pressure. As 0.6 MPa supply pressure resulted in the lower range of RSDs of flow rates for a working pressure range of 0.03 MPa to 0.18 MPa in the FIA system, 0.6 MPa supply pressure was used in all further experiments. To understand the precision of reagent delivery in the FIA system, the relative standard deviations of the mass flow rates of IPA/DIW, 32% HCl, 18% NaOH, 0.5 mM Cu2+ and 0.88 mM luminol in five lines were calculated and presented in Table 1. Insert Table 1 In-depth investigations of flow rates in each line and working pressures revealed that mass flow rates were linearly varied with the working pressures in copper and luminol lines of Fig.1. The line mass flow rates remained almost constant in the NaOH line and varied nonlinearly with working pressures in acid and IPA/DIW line. The variations of mass flow rates and working pressures are plotted in Figs. S1 to S5 in supplementary information. As the FIA system was designed to work at very low Reynolds number (
PII: DOI: Reference:
www.elsevier.com/locate/talanta
S0039-9140(15)30007-2 http://dx.doi.org/10.1016/j.talanta.2015.05.052 TAL15644
To appear in: Talanta Received date: 8 April 2015 Revised date: 12 May 2015 Accepted date: 22 May 2015 Cite this article as: Parvez Mahbub, Philip Zakaria, Rosanne Guijt, Mirek Macka, Greg Dicinoski, Michael Breadmore and Pavel N. Nesterenko, Flow injection analysis of organic peroxide Explosives using acid degradation and chemiluminescent detection of released Hydrogen peroxide, Talanta, http://dx.doi.org/10.1016/j.talanta.2015.05.052 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting galley proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
FLOW INJECTION ANALYSIS OF ORGANIC PEROXIDE EXPLOSIVES USING ACID DEGRADATION AND CHEMILUMINESCENT DETECTION OF RELEASED HYDROGEN PEROXIDE Parvez Mahbuba, Philip Zakariaa, Rosanne Guijtb, Mirek Mackaa, Greg Dicinoskia, Michael Breadmorea, Pavel N. Nesterenkoa* a
Australian Centre for Research on Separation Science, School of Physical Sciences,
University of Tasmania b
Pharmacy School of Medicine, Australian Centre for Research on Separation Science,
University of Tasmania * * - Corresponding author: Professor Pavel N. Nesterenko Australian Centre for Research on Separation Science, School of Chemistry, University of Tasmania, Private Bag 75, Hobart 7001, Australia Tel: +61 (03) 6226 2165 Fax +61 (03) 6226 2858 Email: [email protected]
Keywords: organic peroxides, explosives, flow injection analysis, acid degradation, chemiluminescence
Abstract The applicability of acid degradation of organic peroxides into hydrogen peroxide in a pneumatically driven flow injection system with chemiluminescence reaction with luminol and Cu2+ as a catalyst (FIA-CL) was investigated for the fast and sensitive detection of organic peroxide explosives (OPEs). The target OPEs included hexamethylene triperoxide diamine (HMTD), triacetone triperoxide (TATP) and methylethyl ketone peroxide (MEKP). Under optimised conditions maximum degradations of 70% and 54% for TATP and HMTD, respectively were achieved at 162 µLǜmin-1, and 9% degradation for MEKP at 180 µLǜmin-1. Flow rates were precisely controlled in this single source pneumatic pressure driven multichannel FIA system by model experiments on mixing of easily detectable component solutions. The linear range for detection of TATP, HMTD and H2O2 was 1-200 µM (r2= 0.98~0.99) at both flow rates, while that for MEKP was 20-200 µM (r2=0.97) at 180 µLǜmin1
. The detection limits (LODs) obtained were 0.5 µM for TATP, HMTD and H2O2 and 10
µM for MEKP. The detection times varied from 1.5 to 3 min in this FIA-CL system. Whilst the LOD for H2O2 was comparable with those reported by other investigators, the LODs and analysis times for TATP and HMTD were superior, and significantly, this is the first time the detection of MEKP has been reported by FIA-CL.
1. Introduction The simplicity of in-house preparation of the organic peroxide explosives (OPEs), such as hexamethylene triperoxide diamine (HMTD), triacetone triperoxide (TATP) and methylethyl ketone peroxide (MEKP), from readily available materials was the main reason of their use in some recent terrorists attacks [1]. So, there is a strong demand for the development of fast, simple and sensitive methods of their identification and quantitative determination [2]. This
task is not trivial because of high volatility and absence of chromophoric groups in the molecules of OPEs. For these reasons the use of common analytical methods such as GC or HPLC with UV/visible detection is not readily suitable for their direct determination, so the application of more complex hyphenated techniques, typically involving mass spectrometry (MS), is required. Schulte-Ladbeck et al. [3] proposed RP HPLC with on-line Fourier transfom infrared (FTIR) detection for direct determination of TATP and HMTD. De Tata et al. [4] reported the application of RP HPLC with quadrupole time-of-flight mass spectrometry (HPLC-QToF-MS) for direct determination of various OPEs. However, micellar electrokinetic chromatography with UV detection was employed by Johns et al. [5] recently for separation of OPEs including HMTD and TATP in post blast scenario without any marked improvement compared to the hyphenated HPLC methods in terms of sensitivity. Alternatively, the determination of OPEs can be based on electrochemical [6], fluorescent [7] and chemiluminescent [8] detection of hydrogen peroxide as the main degradation product of OPEs. It should be noted that decomposition of one OPE molecule can result in more than one molecule of H2O2, so in case of 100% degradation a magnified analytical response can be expected. Because of its robustness, sensitivity and simplicity of integration with FIA, chemiluminescent detection (FIA-CL) is one of the most popular techniques for the on-line detection of hydrogen peroxide. The application of FIA-CL has been reported for the determination of H2O2 in natural waters [9], rainwater [10] and seawater [11]. The use of chemiluminescent detection of H2O2 after decomposition of TATP and HMTD was reported for determination of these explosives by a number of research groups [3, 6 and 12]. The analysis times reported varied from 5 to 12 min with the sensitivity ranging from 2.5 µM for TATP [6] to 500 µM for HMTD [3]. Obviously, the crucial parameters in the flow-through methods of analysis of explosives via detection of hydrogen peroxide are the velocity and conversion degree of OPEs into H2O2.
According to the literature data, the degradation of peroxide explosives can be accomplished enzymatically [12, 13], photochemically [14] or by using mineral acids [15]. The latter option appears more simple and robust, but it takes almost 12 hours for complete decomposition of TATP and HMTD which is not suitable for rapid screening. Additionally, there has been no report of the decomposition of methylethyl ketone peroxide (MEKP) - listed as a priority explosive by several government agencies including US National Counterterrorism Centre and the Australian Army [16]. It should be noted that 100% degradation of OPEs does not mean the maximum possible concentration of hydrogen peroxide suitable for detection, as hydrolysis of H2O2 occurs together with organic peroxides. On this reason, acid degradation conditions of OPEs should be carefully attuned to provide fast and sensitive response in FIA system. This study was a part of a larger project directed on construction of portable automated system for the fast screening of samples on presence of traces of homemade explosives including organic peroxides and inorganic explosives. According to the proposed design the whole system composed of three separate units, namely, sample extraction unit, FIA-CL detection unit for organic peroxides and capillary zone electrophoresis unit for parallel profiling of inorganic anions contents. The high degree of automation, minimal consumption of reagents and robust operation for extended period of time were considered as important requirements for this system. The system should provide either positive or negative answer in a short time on the presence of explosives at low concentration level. Typically, multicomponent FIA analysers use multi-channel peristaltic pumps for delivery of sample and reagents solutions. According to Dasgupta to get very low flow rates at µL/min level, as required in this study, peristaltic pumps are of little help [17]. Piezoelectric pumps, pneumatic pressure or gravity driven systems are often preferred choice in this case [18]. Pressure driven FIA instruments have advantages over systems using peristaltic pumps in
terms of lower signal/noise ratio, better reproducibility of timing and expanded possibilities for the use of aggressive solvents and carriers [18]. According to Valcarcel and De Castro, the main drawback of pneumatic driven instruments is connected with difficulty in accurate control of the flow rates in multiple channel FIA systems due to complex changes in hydraulic resistances resulting from different channel geometry and reagent viscosity [19]. In this study, a special attention was paid to the development of a control mechanism of flow rates in separate lines of multichannel FIA system driven by pneumatic pressure from a single source. The objective of this study is to optimise conditions for acid degradation of organic peroxides and subsequent chemiluminescent luminol based detection of hydrogen peroxide within pressure driven FIA-CL analytical unit for the purpose of fast qualitative determination of HMTD, TATP and MEKP. The target total analysis time was less than 2 min for rapid screening and high-throughput analysis, imposing a significant challenge to the optimisation of the flow conditions.
2. Experimental Section 2.1. Reagents and Chemical Standards Hydrated copper sulfate (CuSO4ǜ5H2O), 32% concentrated hydrochloric acid (HCl), 30% (w/w) reagent grade H2O2, isopropanol and sodium hydroxide (NaOH) pellets were purchased from Sigma-Aldrich (Sydney, Australia). Luminol was purchased from Fluka (Sydney, Australia). Element free deionised water was used to prepare all stock and working solutions. The TATP standard (10,000 mg L-1, 99.9% single component) and HMTD standard (5,000 mg L-1, 98.4% single component) were procured from Accustandard, USA. The MEKP standard (10,000 mg L-1) was supplied by the Australian Defence Science and Technology Organisation (DSTO).
2.2. Preparation of precise assay of the standard solution for H2O2 A stock solution of approximately 1000 mg L-1 of the 30% H2O2 was prepared through serial dilution. Then 50 mL of stock solution was transferred into a 500 mL conical flask, diluted with 200 mL of deionised water, and then 30 mL of 25% sulfuric acid was added. The solution was titrated with a standard 0.02 M potassium permanganate solution until the colour changed to pink. The working solutions were prepared by further dilution of stock solution in DIW.
2.3. Instrumentation A FIA-Cl system consisting of a low pressure Cheminert 6 port 2 position injector valve (C22-3186EH-FL, VICI, Houston USA), five SMC precision pressure regulators (IR 100001, SMC, Japan) and a Hamamatsu photomultiplier (10493-001, Hamamatsu, Japan) were used. A schematic of the instrument used for acid degradation study of TATP, HMTD and MEKP is illustrated in Figure 1. The system comprised SMC pressure regulators connected through a manifold to an external compressed air supply. Each regulator was used to control the reagent solution flow from a 500 mL glass bottle (Schott AG, Sigma-Aldrich, Australia) by applying pressures ranging from 0.01-0.2 MPa (1.5 psi to 29 psi). The pneumatic lines consisted of polyurethane tubing (2.5 mm I.D., SMC, Japan) and the hydraulic lines consisted of FEP tubing (0.203 mm I.D., Upchurch, USA). The 100 µL sample plug was carried into the Cheminert mixer (CM1XKF, VICI, Houston, USA) in 50:50 v/v deionised water - isopropanol where it was mixed with 32% HCl. Isopropanol was used to ensure complete dissolution of OPEs from the collected and extracted samples (the exact procedure is not included in this paper). The OPE containing
samples were degraded in acidic mixture in a 1 m PTFE knitted tubing coil reactor (0.25 mm I.D., 49.1 µL internal volume, Biotech AB, Onsala, Sweden) resulting in the release of H2O2 molecules. The excess HCl was neutralised by the addition of 18% NaOH and the resulting solution was then mixed with the luminol – Cu2+ reagent in a tee mixer (P-712, 2.9 µL swept volume, Upchurch, Oak Harbor, USA). The subsequent chemiluminescence reaction was detected with the photomultiplier tube. The chemiluminescence flow cell was fabricated inhouse using FEP tubing (0.508 mm I.D., 1548L, Upchurch, USA) with the total volume of the flow cell being 85.32 µL. The chemiluminescence signal was acquired by a Powerchrom data acquisition system (ER280, EDaq, Sydney, Australia) with proprietary software version 8.1. The pH of the effluent was monitored by an in-line pH monitoring flowcell (ColeParmer, Australia, vertical flow glass electrode 50 L internal volume) as shown in Figure 1. Insert Figure 1 3. Results and Discussions 3.1.Optimisation of reagent concentrations The crucial part of FIA-CL under development is acid degradation of OPEs in isopropanolwater (50:50) extracts of swabs used for sample collection from different surfaces. It was shown that such mixture provides the best extraction of various types of inorganic and organic explosives [20]. However, as a result of elevated viscosity of this mixture the FIA system in this study need to operate at pressures higher than those normally provided by peristaltic pumps. Also, initial experiments using peristaltic pumps demonstrated poor stability of the silicone tubing when in contact with solutions of isopropanol, hydrochloric acid and sodium hydroxide requiring the frequent replacement of the tubing and leading to poor reproducibility of data. For this reason a pneumatic FIA-CL system consisting of five
pressure lines equipped with precision low-pressure regulators (maximum operating pressure 0.2 MPa) was constructed as shown in Fig. 1. The acid fumes and the heat generated from the reaction between concentrated HCl and NaOH was a practical challenge in selecting the acid and base concentrations in this study. Additionally, the solubility of NaCl, formed as a product of the neutralisation reaction of HCl and NaOH, in isopropanol is very low (0.013 gǜg-1 at 23.5 C). To avoid frequent regulator malfunction and system blockage due to NaCl precipitation, 50:50 v/v deionised water isopropanol mixture was used to carry the 100 µL sample plug into the cheminert mixing chamber. The system operated optimally without blockage from NaCl precipitation and system malfunction from acid fumes and excessive heat when 32% HCl (concentrated) and 18% NaOH (w/v) were used. 3.2. Optimisation of the operating pressures and calculation of individual reagent flow rates The optimisation of reagent concentrations and optimum flow rates influencing degradation degree of OPEs and maximum response of chemiluminescent detection is not trivial task in multichannel FIA system with pressure driven flows. This is due to difficulties in control of flow rate in separate lines of pressure system, when changes in backpressure/flow rate in one line may effect on flow rates in other lines. Therefore, the exact concentration of used reagents should be measured in a separate experiment. Initially the system was investigated to establish the optimum input pressure at the regulators to provide the required flow rates with the least variance. The mass flow rates of deionised water (DIW) in all five lines were determined at 0.03, 0.07, 0.1, 0.15 and 0.18 MPa by precise measuring the weight losses in the containers after 8 min operation. The experiments were repeated for 5 consecutive days. The relative standard deviations (RSD) of the flow
rates in five lines ranged from 25% to 51% for 0.4 MPa supply pressure whilst RSDs ranged from 15% to 30% for 0.6 MPa supply pressure. As 0.6 MPa supply pressure resulted in the lower range of RSDs of flow rates for a working pressure range of 0.03 MPa to 0.18 MPa in the FIA system, 0.6 MPa supply pressure was used in all further experiments. To understand the precision of reagent delivery in the FIA system, the relative standard deviations of the mass flow rates of IPA/DIW, 32% HCl, 18% NaOH, 0.5 mM Cu2+ and 0.88 mM luminol in five lines were calculated and presented in Table 1. Insert Table 1 In-depth investigations of flow rates in each line and working pressures revealed that mass flow rates were linearly varied with the working pressures in copper and luminol lines of Fig.1. The line mass flow rates remained almost constant in the NaOH line and varied nonlinearly with working pressures in acid and IPA/DIW line. The variations of mass flow rates and working pressures are plotted in Figs. S1 to S5 in supplementary information. As the FIA system was designed to work at very low Reynolds number (
