Analysis of Furan and Alkylfurans in Food Samples (Part 1): Choosing GC-MS Conditions and a Sample Preparation Technique

Cooking, or the process of heating up foodstuffs, leads to diverse chemical reactions that permit the development of aroma and a richer food taste, facilitate food preservation, and in many cases, allow for an easier absorption of nutrients and improved digestibility.  During such process, compounds like furan and alkylfurans are generated as a result of the degradation of carbohydrates, ascorbic acid (vitamin C), and derivatives, as well as some lipids[1–4]. The International Agency for Research of Cancer classified furan as an animal carcinogen and as a possible human carcinogenic compound, so there is a general concern about the possible health risks associated with the occurrence of furan and alkylfurans in food [5]. Indeed, the elevated concentrations of these process contaminants in food commodities such as coffee, some baby foods, baked goods, and canned foodstuff, make them a concerning source of exposure to humans. International food organizations such as the European Food Safety Authority (EFSA) and the US Food and Drug Administration (US FDA) initially started to collect information related to furan analytical methods, concentration in various foods, and exposure through consumption [6,7]. Then, in 2019, the EFSA published a report calling for data to assess furan and alkylfurans exposure through food consumption [8].   

Due to the high volatility of furan and alkylfurans, the two main analytical methods reported for the analysis of these compounds include headspace (HS) and HS solid phase microextraction (HS-SPME), both coupled to gas chromatography-mass spectrometry (GC-MS). The former has been broadly employed in several manuscripts and some official methods, and it is particularly useful in the quantitation of samples with high levels of furan and alkylfurans (e.g., coffee) [9,10]. HS-SPME, on the other hand, has demonstrated improved sensitivity for the quantitation of furan and alkylfurans at lower concentration levels in samples such as baby food [3,11,12]. Unfortunately, issues related to the fragility of traditional SPME extraction devices are always a matter of concern. For this reason, we decided to evaluate the performance of our sturdy SPME Arrow in furan and alkylfurans analysis.

For this work, furan, 2-methylfuran, 3-methylfuran, 2-ethylfuran, 2,5-dimethylfuran and 2-pentylfuran were selected as target analytes (Figure 1), and furan-d4, 2-methylfuran-d6, 2-ethylfuran-d5 and 2-pentylfuran-d11 were chosen as internal standards. First, we optimized our GC-MS method to be able to detect and resolve all target analytes and internal standards (Table 1). To achieve improved sensitivity, selected ions were monitored in four different time windows, as shown in Table 2.

Figure 1. Structure of target analytes

Table 1. GC-MS conditions for the analysis of furan and alkylfurans.

Instrument	Agilent 7890B GC & 5977B MSD
Column	Rxi-624Sil MS, 30 m, 0.25 mm ID, 1.40 µm (cat.# 13868)
Injection Mode	Split (1:10 and 1:100)
Liner	Topaz 1.8 mm ID SPME/straight liner (cat# 23280)
Inj. Temp.	280°C
Split Flow	14.0 mL/min (10:1) and 140 mL/min (100:1)
Purge Flow	5 mL/min
Oven	35°C (hold 3 min) to 75°C by 8°C/min, then to 200°C (hold 1 min)  by 25°C/min
Carrier Gas	He, constant flow
Flow Rate	1.40 mL/min
Analyzer	MS (quadrupole)
Acquisition Type	SIM
Ionization Mode	EI (70 eV)
Transfer Line Temp.	280 °C
Source Temp.	325 °C
Quadrupole Temp.	200 °C
Solvent delay	2.2 min

Table 2. MS parameters for the analysis of furan, alkylfurans, and their internal standards (SIM mode).

Segment start time, min	Compound (Rt, min)	Ions	Dwell time, ms
2.2	Furan (2.447)	39	50
		68*
	Furan-d4 (2.428)	42
		72*
4.2	2-Methylfuran (4.536)	53	30
		81
		82*
	3-Methylfuran (4.846)	53	30
		81
		82*
	2-Methylfuran-d6 (4.464)	58	30
		88*
6.6	2-Ethylfuran (7.100)	53	30
		81*
		96
	2-Ethylfuran-d5 (7.001)	55	30
		101*
	2,5-Dimethylfuran (7.243)	67	30
		95*
	2,5-Dimethylfuran-d3 (7.179)	84	30
		99*
10.6	2-Pentylfuran (11.570)	81	30
		138*	30
	2-Pentylfuran-d11 (11.501)	83	30
		149*	30

*Quantifier ions

The selected GC-MS conditions provided good retention and separation of all analytes and internal standards in less than 14 min. As shown in Figure 2, separation between 2,5-dimethylfuran and 2-ethylfuran (isomeric compounds), and between 2,5-dimethylfuran-d3 and 2-ethylfuran, which shared multiple ions, was successfully attained.

Once the instrumental conditions for furan and alkylfurans analysis were optimized, we performed a comparison among HS, HS-SPME (traditional fibers), and HS-SPME Arrow. For the SPME devices, we selected a wide carbon range (CWR) coating because it is the best phase for the extraction of highly volatile compounds of low molecular weight. As seen in Figure 3, the SPME-based techniques significantly outperformed HS analysis, and the SPME Arrow provided the best results in terms of analyte responses in all the cases. Remarkable instrumental response differences of more than 2-fold were observed for furan, 2-methylfuran, 3-methylfuran, 2-ethylfuran, and 2,5-dimethylfuran when comparing traditional SPME vs. SPME Arrow. In the case of 2-pentylfuran, the difference in response between the two SPME-based techniques was 1.3-fold. Based on this data and taking into account the improved mechanical robustness of these devices, the SPME Arrow was chosen for further method development.

Figure 2. GC-MS chromatogram corresponding to furan and alkyl furan standards and internal standards.

Figure 3. Results corresponding to the headspace analysis of furan and alkylfurans spiked (20 ng of each analyte) in 10 mL of sodium chloride solution at 30% (n=3). SPME sampling: 2 min extraction time, 5 min incubation at 40 ⁰C, 1 min desorption at 280 C, agitation at 250 rpm. Headspace tool conditions: 1000 uL sample volume, 20 min incubation time, 120 s purge time, 80 ⁰C incubation temperature, 150 ⁰C HS tool temperature.

Now that we have selected our GC-MS conditions and the SPME Arrow as the sample prep technique for the analysis of furan and alkylfurans in food, we will be discussing relevant steps to obtain a successful SPME method in future blogs. So please stay tuned!!

References

1. Limacher, A., Kerler, J., Davidek, T., Schmalzried, F., Blank, I., Formation of Furan and Methylfuran by Maillard-Type Reactions in Model Systems and Food. n.d., DOI: 10.1021/jf800268t.
2. Crews, C., Castle, L., A review of the occurrence, formation and analysis of furan in heat-processed foods. Trends Food Sci. Technol. 2007, 18, 365–372.
3. Frank, N., Dubois, M., Huertas Pérez, J. F., Detection of Furan and five Alkylfurans, including 2-Pentylfuran, in various Food Matrices. J. Chromatogr. A 2020, 1622, 461119.
4. Märk, J., Pollien, P., Lindinger, C., Blank, I., Märk, T., Quantitation of furan and methylfuran formed in different precursor systems by proton transfer reaction mass spectrometry. J. Agric. Food Chem. 2006, 54, 2786–2793.
5. International Agency for Research on Cancer, IARC Monographs on the Evaluation of the Carcinogenic Risks to Humans. 1995.
6. Katrine Knutsen, H., Alexander, J., Barreg ard, L., Bignami, M., Br, B., Ceccatelli, S., Cottrill, B., Dinovi, M., Edler, L., Grasl-Kraupp, B., Hogstrand, C., Hoogenboom, L., Stefano Nebbia, C., Oswald, I. P., Petersen, A., Rose, M., Roudot, A.-C., Schwerdtle, T., Vleminckx, C., Vollmer,  unter, Chipman, K., De Meulenaer, B., Mennes, W., Schlatter, J., Schrenk, D., Baert, K., Dujardin, B., Wallace, H., SCIENTIFIC OPINION Risks for public health related to the presence of furan and methylfurans in food EFSA Panel on Contaminants in the Food Chain (CONTAM Panel members. EFSA J. 2017, 15, 5005.
7. FDA, Exploratory data on furan in food: individual food products, sept., https://www.fda.gov/food/chemical-contaminants-food/exploratory-data-furan-food (last time accessed: June 16, 2022).
8. (European Food Safety Authority), E., Call for continues collection of chemical occurrence data in food and feed, https://www.efsa.europa.eu/en/consultations/call/190410 (last time accessed: June 16, 2022).
9. Cao, P., Zhang, L., Yang, Y., Wang, X. dan, Liu, Z. ping, Li, J. wen, Wang, L. yuan, Chung, S., Zhou, M., Deng, K., Zhou, P. ping, Wu, P. gu, Analysis of furan and its major furan derivatives in coffee products on the Chinese market using HS-GC–MS and the estimated exposure of the Chinese population. Food Chem. 2022, 387, 132823.
10. Shen, M., Liu, Q., Jia, H., Jiang, Y., Nie, S., Xie, J., Li, C., Xie, M., Simultaneous determination of furan and 2-alkylfurans in heat-processed foods by automated static headspace gas chromatography-mass spectrometry. LWT - Food Sci. Technol. 2016, 72, 44–54.
11. Frank, N., Dubois, M., Nguyen, K. H., Fromberg, A., Delatour, T., Quantification of furan and 5 alkylfurans with complete separation of 2-ethylfuran and 2,5-dimethylfuran isomers in cereals, coffee and Infant products by GC–MS. J. Chromatogr. B Anal. Technol. Biomed. Life Sci. 2021, 1179, 122765.
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