View metadata, citation and similar papers at core.ac.uk brought to you by CORE
provided by Elsevier - Publisher Connector Journal of Saudi Chemical Society (2016) 20, S352–S356
King Saud University Journal of Saudi Chemical Society
www.ksu.edu.sa www.sciencedirect.com
ORIGINAL ARTICLE A simple spot test quantification method to determine formaldehyde in aqueous samples
Amara Dar a,*, Umer Shafique a, Jamil Anwar b, Waheed-uz-Zaman b, Arooj Naseer a
a Center for Undergraduate Studies, University of the Punjab, New Campus, Lahore-54590, Pakistan b Institute of Chemistry, University of the Punjab, New Campus, Lahore-54590, Pakistan
Received 26 April 2012; accepted 5 December 2012 Available online 20 December 2012
KEYWORDS Abstract A simple, accurate, and economical method has been proposed to measure formaldehyde in Chromotropic acid; aqueous samples. The method is based on quantification of classical chromotropic acid – formalde- Densitometry; hyde violet spots developed on TLC. Different parameters such as concentration of chromotropic Formaldehyde; and sulfuric acid, time of heating and order of application of reagents have been studied to find out Micro-analysis; the optimum working procedure. Spots have been quantified by scanning the spotted TLC and ana- Quantification of spot test; lyzing the image in computer with Visual Basic 6.0 based graphic application. The study consisted of Spot test analysis developing an appropriate calibration line, analyzing artificial and real samples, and comparing the new method with a standard spectrophotometric method (NIOSH Method-3500, 1994). It has been concluded that the present method had the capability for measuring formaldehyde in aqueous samples at trace level with high precision and accuracy, particularly when dealing with turbid and small volume samples where the standard method failed. ª 2012 Production and hosting by Elsevier B.V. on behalf of King Saud University. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
1. Introduction Formaldehyde is a gas at room temperature, but readily con- verts into a variety of other gaseous derivatives like trioxane. Formaldehyde or methanal (HCHO) is the simplest aldehyde Formaldehyde occurs in the environment up to 0.03 ppm parts that is an important precursor to many chemical compounds. of air. Materials having formaldehyde, such as urea–formalde- Yearly production of formaldehyde around the globe has hyde foam insulation (UFFI) can release it in the form of gas or exceeded 23 million tons during last few years (Lide, 2004). vapor. Pressed-wood products are the major source of formal- dehyde indoor pollution. Cigarette smoke, gas stoves, wood- burning stoves, and kerosene heaters can also release formalde- * Corresponding author. Tel.: +92 4299232068; fax: +92 4299232057. hyde. It is highly toxic to both plants and animals, and is par- E-mail address: [email protected] (A. Dar). ticularly dangerous for the human eyes (Swenberg et al., 1980). Peer review under responsibility of King Saud University. When present in the air at levels exceeding 0.1 ppm, it causes severe irritation in the eyes, nose, and throat and usually results in watery eyes with burning sensations, coughing, wheezing, Production and hosting by Elsevier nausea, and skin irritation (Sakai et al., 2002). There is some
http://dx.doi.org/10.1016/j.jscs.2012.12.002 1319-6103 ª 2012 Production and hosting by Elsevier B.V. on behalf of King Saud University. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Determination of formaldehyde in water S353 evidence that formaldehyde become carcinogen when inhaled proportional to the concentration of formaldehyde, a simple (Leonard, 1999). Epidemiological investigations of the fatality analytical method has been developed for precise quantification of factory workers following prolonged occupational exposure of formaldehyde in aqueous samples at trace levels. The color to formaldehyde showed a slight excess of lung cancer (Ache- densities of the spots were measured with simple software by son et al., 1984a, b). Safe exposure limit to formaldehyde for taking the image of spotty TLC in computer subsequent to 30 min is 100 lgmÀ3 (0.08 ppm) (Sawada, 2006). scanning. There are other techniques to quantify chromotropic Formaldehyde in water comes mainly from oxidation of or- acid – formaldehyde spots (Boyd and Logan, 1942; Boos, 1948); ganic matter during ozonation (Glaze et al., 1989) and chlori- the present method is only one of its kind in terms of measuring, nation (Becher et al., 1992). In drinking water, it can arise from quantity of sample, simplicity and accuracy. The most appro- leaching from plastic fittings and water treatment processes. In priate application of the method is on-field analysis of trace the dissolved form, formaldehyde converts into a diol amounts of formaldehyde in aqueous samples. CH2(OH)2. An aqueous solution of formaldehyde is called for- malin. Pure formalin is a saturated solution of formaldehyde in 2. Materials and methods water (nearly 37% by weight) (Turoski, 1985; Keilson and Newell, 1990). It does not persist in water for long time be- 2.1. Stock solution and standards cause it is broken down within few hours by sunlight or by bacteria. However, when ingested to a level above 2.6 mg lÀ1 Stock solution of formaldehyde (1000 mg lÀ1) was prepared by (NOEL) via contaminated food or water, formaldehyde has diluting analytical grade 10% formalin solution (Sigma–Al- been shown to cause vomiting, abdominal pain, dizziness, drich, Inc.) with proper quantity of double distilled water. and in extreme cases can cause even death. Standard solutions of desired concentrations (2–10 mg lÀ1) Li et al., 2007 developed a fluorophotometric method for were prepared by appropriate dilution of the stock solution. formaldehyde determination in environmental waters, which was founded on the reaction of formaldehyde with acetoace- tanilide and ammonia. The method was simple, rapid, eco- 2.2. Chemicals nomical, and highly sensitive. Ro´zyo_ et al., 2002 determined the levels of formaldehyde in human saliva. Formaldehyde Technical grade chromotropic acid disodium salt dihydrate, was determined as the dimedone adduct (formaldemethone) (HO)2C10H4(SO3Na)2Æ2H2O and reagent grade concentrated using OPLC. Tomkins et al., 1989 determined total formalde- sulfuric acid (Sigma–Aldrich, Inc.), 95–98%, 1.840 g mlÀ1 at hyde in drinking water samples. Formaldehyde present in 1 li- 25 C (lit.) were the chemicals that were used as it is or as dou- ter water was derivatized with 2,4-dinitrophenylhydrazine in bled distilled water diluted solutions in order to evaluate the an acidic media and subsequently extracted with chloroform. effect of these on color intensity of spots. Desired solutions After separation by solvent extraction, the product was quan- of chromotropic acid were prepared in 50% sulfuric acid. tified using reverse-phase liquid chromatography. Keyvanfard, 2010 proposed a simple and rapid catalytic kinetic method 2.3. Procedure (based on the catalytic effect of formaldehyde on the oxidation of cresyl violet by bromate in the presence of sulfuric acid) for TLC plate (Merck, Aluminum sheet, Silica gel 60F , the determination of trace amount of formaldehyde. The meth- 254 3 · 5 cm) was used to develop spots since paper could not be od was exercised for the measurement of formaldehyde in employed (burnt because of concentrated sulfuric acid). One water samples. micro-liter drop of each, chromotropic acid, sulfuric acid It is obligatory to develop simple and precise analytical and aldehyde solution was employed with micropipette (Pip- methods to determine formaldehyde because of its widespread ettman) one over the other. The order of application of re- use, toxicity, and volatility. There are several methods available agents and aldehyde solution was altered to check the effect for the detection of formaldehyde in air and water. The most on color density of the spot. Later, the TLC was placed in widely used methods for air samples are based on spectropho- an oven for 2–20 min at 60 C to notice the effect of time. Vio- tometry, while for aqueous samples high-performance liquid let color spots intensifying on cooling were obtained. chromatography (HPLC) is a preferred choice. Other methods include colorimetry, fluorimetry, polarography, gas chroma- tography (GC), infrared detection, flow injection analysis and 2.4. Quantification of spots and analysis of samples gas detector tubes (Cogliano et al., 2004). HPLC or GC coupled with the mass spectrometer (MS) are most sensitive techniques. The spotty plate was scanned on a flatbed scanner (HP 3670 However, these techniques are very expensive. Besides, difficul- CCD Reflective Flatbed Scanner) and the image was imported ties arise with the monitoring of turbid samples that can block in Visual Basic 6.0 based graphic application to measure the columns or membranes or precipitations can demand consider- color density of spots. Detail of graphic application to read able maintenance effort (Papaefstathiou et al., 1996). color densities of the spots is given in (Anwar et al., 2010). This paper describes a simple, rapid, sensitive, selective, and A calibration line was plotted taking concentration of formal- reproducible analytical method of formaldehyde analysis based dehyde’s standards as abscissa against the corresponding color on the quantitative spot test technique. The violet–red spots of density of spots (ordinate). Slope, intercept, standard error of formaldehyde with chromotropic acid were developed on TLC. estimates and correlation coefficient were measured. Finally, Chromotropic acid has been widely used as an analytical synthetic as well as true aqueous samples of formaldehyde reagent in organic as well as in biological chemistry after the dis- were analyzed using the same set of conditions by measuring covery of violet spots with formaldehyde in 1937 (Eegriwe, their color densities and interpolating the corresponding con- 1937). In view of the fact that the color density of spot is directly centrations from standards’ calibration line. S354 A. Dar et al.
2.5. Comparison with standard method Chromotropic acid is the most widely used reagent for formaldehyde analysis because it is highly selective. Despite The present method was compared with the standard method the advent of sophisticated methods such as HPLC and GC, of formaldehyde analysis recommended by the National Insti- spectrophotometric method based on chromotropic acid is still tute for Occupational Safety and Health (NIOSH). It is NIOSH an international reference method of formaldehyde analysis Method-3500, published in NIOSH Manual of Analytical because of simplicity and sensitivity. The classical method uses Methods in 1994. The detailed procedure of the standard spec- concentrated sulfuric acid but modifications have been made trophotometric method using chromotropic acid and concen- to replace the aforementioned hazardous and corrosive acid trated sulfuric acid is given in Georghiou and Ho (1989) and with some less dangerous acids like glacial acetic acid, hydro- Fagnani et al. (2003). The method employs chromotropic acid chloric acid, and phosphoric acid. However, it has been found that the sensitivity reduces to a significant extent in the absence (1%), sulfuric acid (H2SO4, 96%), formalin solution (37%), and deionized water. To the solution of formaldehyde, small of sulfuric acid (Fagnani et al., 2003). Therefore, the present amount of chromotropic acid is added followed by addition method was developed with sulfuric acid. The drawback of of sulfuric acid in excess. Solution is heated and cooled and sub- using concentrated sulfuric acid was that it burnt the paper jected to spectrometric analysis. The absorbances were re- and corroded the TLC. For that reason, a test was performed corded on a double beam spectrophotometer (Labomed, to find out an optimum concentration of sulfuric acid that can UVD-3500) at 580 nm against the reagent blank. Real and arti- give maximum color to spot and would not corrode the TLC. ficial formaldehyde-contaminated water samples were tested by Effect of sulfuric acid’s strength on color density of spot was À1 both methods, and the results were compared. studied working with 10 mg l solution of formaldehyde and 1% solution of chromotropic acid in 50% sulfuric acid. It was found that the color density of spot increased with an 3. Results and discussion increase in the strength of acid. However, after 50%, the TLC started to corrode. Therefore, the optimum strength of Spot tests are simple chemical procedures in which an essential acid chosen was 50%. feature is to employ a drop of the analyte on the reagent/s onto The chemistry of the reaction of chromotropic acid with paper or TLC that will produce a spot of specific color. These formaldehyde in strongly acidic media is very complex and is detection methods are responsive and highly selective. The not known completely. The often-quoted reaction consists of detections are micro-analytical and are used to identify a wide two steps that are shown in the Scheme 1. Sulfuric acid acts range of organic and inorganic compounds. These procedures as a dehydrant in the first step and as an oxidant in the next. are elementary and reasonably fast (Feigl and Anger, 1998). The p-quinoidal product is assumed to absorb light in the Spot test was originally developed for qualitative analysis range of 570–580 nm. However, this product has never been but in a near past, efforts have been made to quantify the pro- confirmed experimentally by any valid mean. Conversely, cess (Anwar et al., 2010). Working on the same principle, the NMR analysis suggested that the final product responsible classical chromotropic acid spot test method of formaldehyde for violet color is not p-quinoidal but mono-cationic dibenzo- detection has been quantified to find out the formaldehyde xanthylium (Scheme 2)(Georghiou and Ho, 1989). However, content of water at trace level with no trouble.
- SO HO S 3 SO3H 3
HO HO OH - H2O 2 + HCHO HO HO OH
- SO 3 SO3H HO3S
-H2O +O
HO S SO3H 3
HO O
HO OH
SO3H HO3S Violet-red
Scheme 1 Seses. Determination of formaldehyde in water S355
- - propanal, butanal, isovaleric aldehyde, chloral hydrate, gly- O3 S OHHO SO3 oxal, aromatic aldehydes, formic acid, phenoxyacetic acid + O and isopropyl ester, only the last two produced violet spots at optimum conditions (2% chromotropic acid, 50% sulfuric acid, and heating at 60 C for 14 min), while furfural, fructose and sucrose produced yellow spots. The order of application of - - reagents and aldehyde solution was also changed and it was SO3 SO3 noted that the correct order was to apply formaldehyde solu- Scheme 2 Asasa. tion foremost, then sulfuric acid followed by chromotropic acid. When chromotropic acid was applied foremost, the final color of spot was brownish rather than violet. It may be be- the nature of this chromogen and its formation has never been cause of silica or some impurity on TLC that interfered with conclusively proven. chromotropic acid and did not allow it to produce violet chro- The effect of concentration of chromotropic acid on color mogen (Scheme 2) on contact with formaldehyde. intensity of the spot was studied. It was found that the color TLC containing violet spots was imported to specially de- intensity had increased with an increase in the concentration signed Visual Basic 6.0 based graphical application using flat- of chromotropic acid up to 2%. However, afterward there bed scanner at 300dpi, 24-bit RGB mode. Each spot was was no prominent increase in color density with further in- selected inside a selection marquee and the color density was crease in concentration. In the same way, time to heat TLC measured. The software measured the red, green, and blue com- at 60 C was changed from 2 to 20 min and it was found that ponents of each pixel inside the margin and sums up the values 14 min were optimum to obtain maximum intensity of the spot. to give a final color density. Calibration lines (Fig. 1) were plot- To check the selectivity of the method, different compounds ted for formaldehyde standards in the range of 2–10 mg lÀ1 by were tested separately and with formaldehyde to study the the present method (color density against concentration) and interference if any. It was noted that among acetaldehyde, the standard NIOSH method (absorbance against concentra-
Figure 1 Calibration lines of the present and standard method using Microsoft Excel 2007.
Table 1 Comparison of calibration lines of the present and standard method using Microsoft Excel 2007. Parameter Standard method* Present method Slope 0.049 ± 0.002 18.700 ± 0.627 Y-intercept when X = 0.0 0.086 ± 0.015 38.000 ± 4.160 X-intercept when Y = 0.0 À1.778 À2.032 1/slope 20.580 0.053 95% Confidence intervals Slope 0.041 to 0.056 16.700 to 20.700 Y-intercept when X = 0.0 0.038 to 0.135 24.760 to 51.240 X-intercept when Y = 0.0 À3.219 to À0.692 À3.043 to À1.206 Goodness of fit r2 ** 0.993 0.997 Sy.x *** 0.014 3.967 Result for 5 mg lÀ1 solution 4.745 5.044 Percentage error 5.100% 0.880% * Spectrophotometric method (NIOSH Method-3500 (1994), NIOSH Manual of Analytical Methods (NMAM), ALS Columbia). ** Coefficient of determination. *** Standard error of estimates. S356 A. Dar et al. tion, Method-3500 published in 1994). The calibration data are new analytical technique for micro samples. Anal. Lett. 43, 367– compared in Table 1. Coefficient of determination for the pres- 371. ent method (0.997) was better than the standard method Becher, G., Ovrum, N.M., Christman, R.F., 1992. Novel chlorination (0.993). The coefficient of determination is useful as it gives by-products of aquatic humic substances. Sci. Total Environ. 117 (118), 509–520. the amount of the fluctuation of one variable that is predictable Boos, R.N., 1948. Quantitative colorimetric microdetermination of from the other variable. In addition to the standard statistical À1 methanol with chromotropic acid reagent. Anal. Chem. 20, 964– analysis of calibration, an artificial sample (5 mg l ) was tested 965. by both methods. The output of the present method was Boyd, M.J., Logan, M.A., 1942. Colorimetric determination of serine. À1 5.044 mg l (0.88% €) whereas the standard method found it J. Biol. Chem. 146, 279–287. to be 4.745 mg lÀ1 (5.10% €). The present method used only Cogliano, V., Grosse, Y., Baan, R., Straif, K., Secretan, B., El 1 ll of the sample while the standard method used 5 ml. More- Ghissassi, F., 2004. Advice on formaldehyde and glycol ethers. over, present method was applicable to turbid samples but the Lancet Oncol. 5, 528. standard method could only analyze clear solutions. Eegriwe, E., 1937. Reaktionen und reagenzien zum nachweis organ- Actual samples (tap and canal water from university campus, ischer verbindungen. Z. Anal. Chem. 110, 22–25. Fagnani, E., Melios, C.B., Pezza, L., Pezza, H.R., 2003. Chromotropic and textile effluent from local industry) were also tested by both acid–formaldehyde reaction in strongly acidic media. The role of methods. Tap water was used as such and no formaldehyde was dissolved oxygen and replacement of concentrated sulphuric acid. detected by both methods. Canal water and effluent were al- Talanta 60, 171–176. lowed to settle for 30 min and then filtered ahead of analysis with Feigl, F., Anger, V., 1998. Spot Tests in Inorganic Analysis. Elsevier the standard method, whereas the sample was used without fil- Science Co., Amsterdam. tration in case of the present method. The formaldehyde content Georghiou, P.E., Ho, C.K., 1989. The chemistry of the chromotropic of canal water was 2.546 and 2.429 mg lÀ1 while the concentra- acid method for the analysis of formaldehyde. Can. J. Chem. 67, tions of formaldehyde in textile effluent were 8.926 and 871–876. 8.654 mg lÀ1 by the present and standard method, in that order. Glaze, W.H., Koga, M., Cancilla, D., 1989. Ozonation by-products. 2. The present method is not only economical but also accurate, Improvement of an aqueous-phase derivatization method for the detection of formaldehyde and other carbonyl compounds formed and it can be used for on-field analysis with the help of portable by the ozonation of drinking water. Environ. Sci. Technol. 23, 838– scanners and laptops/smart phones (having color density mea- 847. suring software) since it does not involve painstaking sample Keilson, G.R., Newell, G.W., 1990. Formaldehyde-an Assessment of preparations and sophisticated instruments. Its Health Effects. National Academy of Sciences, Washington, DC. 4. Conclusion Keyvanfard, M., 2010. Catalytic spectrophotometric determination of formaldehyde based on its catalytic effect on the reaction between bromate and cresyl violet. Asian J. Chem. 22, 6708–6714. A trouble-free, precise, and inexpensive method of formalde- Leonard, B., 1999. Eighth Annual Report on Carcinogens: 1998 hyde analysis for aqueous samples has been proposed. The Summary. DIANE Publishing Co., Pennsylvania. method was based on quantification of classical chromotropic Li, Q., Oshima, M., Motomizu, S., 2007. Flow-injection spectrofluo- acid – formaldehyde violet spots developed on TLC. Optimum rometric determination of trace amounts of formaldehyde in water working conditions were 2% chromotropic acid, 50% sulfuric after derivatization with acetoacetanilide. Talanta 72, 1675–1680. acid, and 14 min heating at 60 C to get utmost color intensity Lide, D.R., 2004. CRC Handbook of Chemistry and Physics. CRC Press, Florida. spots. The correct order of reagent’s application was sample Papaefstathiou, I., Bilitewski, U., Luque de Castro, M.D., 1996. (formaldehyde), followed by sulfuric acid and then chromotro- Pervaporation: an interface between fermentors and monitoring. pic acid (1 ll drop of each). Spots were quantified by scanning Anal. Chim. Acta 330, 265–272. the spotted TLC and analyzing the image in computer with Ro´zyo,_ T.K., Zabin_ ´ska, A., Ro´zyo-Kalinowska,_ I., 2002. Use of OPLC homemade Visual Basic 6.0 based graphical application. Cali- for quantitation of HCHO, as the dimedone adduct, in human bration line of the present method was compared with that of saliva in different dental pathologies. JPC-J. Planar Chromatogr. – the standard spectrophotometric method. The present method Mod. TLC 15, 19–22. was found to have better capability for measuring formalde- Sakai, T., Tanakaa, S.-I., Teshimaa, N., Yasudab, S., Urac, N., 2002. hyde in aqueous samples at trace level with high precision Fluorimetric flow injection analysis of trace amount of formalde- and accuracy, especially when dealing with turbid and small hyde in environmental atmosphere with 5,5-dimethylcyclohexane- 1,3-dione. Talanta 58, 1271–1278. volume samples where the standard method was inapplicable. Sawada, Y., 2006. Report of mini association on chemical substances. Annual Report, Japan Industrial Safety and Health Association, References Japan. Swenberg, J.A., Kerns, W.D., Mitchell, R.I., Gralla, E.J., Pavkov, Acheson, E.D., Barnes, H.R., Gardner, M.J., Osmond, C., Pannett, B., K.L., 1980. Induction of squamous cell carcinomas of the rat nasal Taylor, C.P., 1984a. Formaldehyde process workers and lung cavity by inhalation exposure to formaldehyde vapor. Cancer Res. cancer. Lancet 323, 1066–1067. 40, 3398–3402. Acheson, E.D., Gardner, M.J., Pannett, B., Barnes, H.R., Osmond, C., Tomkins, B., McMahon, J., Caldwell, W., Wilson, D., 1989. Liquid Taylor, C.P., 1984b. Formaldehyde in the british chemical industry: chromatographic determination of total formaldehyde in drinking an occupational cohort study. Lancet 323, 611–616. water. J. Assoc. Official Anal. Chem. 72, 835–839. Waheed-uz-Zaman, Anwar J., Shafique, M.U., Salman, M., 2010. Turoski, V., 1985. Formaldehyde: Analytical Chemistry and Toxicol- Computational quantification of spot tests by image scanning-a ogy. American Chemical Society, Washington, DC.