Microfluidic colorimetric analysis system for sodium benzoate detection in foods

Highlights

• A MCA system for SBA concentration detection in food samples is developed.

• Modified Janovsky reaction with two layer reagents are coated in paper-microchip.

• SBA concentrations of 15 commercial food samples are measured.

• The measurement results deviate by no more than 6.0% versus official method.

Abstract

Sodium benzoate (SBA) is a widely-used additive for preventing food spoilage and deterioration and extending the shelf life. However, the concentration of SBA must be controlled under safe regulations to avoid damaging human health. Accordingly, this study proposes a microfluidic colorimetric analysis (MCA) system composing of a wax-printed paper-microchip and a self-made smart analysis equipment for the concentration detection of SBA in common foods and beverages. In the presented method, the distilled SBA sample is mixed with NaOH to obtain a nitro compound and the compound is then dripped onto the reaction area of the paper-microchip, which is embedded with two layers of reagents (namely acetophenone and acetone). The paper-microchip is heated at 120 °C for 20 min to cause a colorimetric reaction and the reaction image is then obtained through a CMOS (complementary metal oxide semiconductor) device and transmitted to a cell-phone over a WiFi connection. Finally, use the self-developed RGB analysis software installed on the cell-phone to obtain the SBA concentration. A calibration curve is constructed using SBA samples with known concentrations ranging from 50 ppm (0.35 mM) to 5000 ppm (35 mM). It is shown that the R + G + B value (Y) of the reaction image and SBA concentration (X) are related via Y = −0.034 X +737.40, with a determination coefficient of R² = 0.9970. By measuring the SBA concentration of 15 commercially available food and beverage products, the actual feasibility of the current MCA system can be demonstrated. The results show that the difference from the measurement results obtained using the macroscale HPLC method does not exceed 6.0%. Overall, the current system provides a reliable and low-cost technique for quantifying the SBA concentration in food and drink products.

Introduction

Preservatives are widely used in the food, beverage, drug and cosmetics industries to reduce microbial spoilage, maintain freshness, enhance the nutritional quality and prolong the shelf life (Piper, & Piper, 2017). Among the many preservatives available nowadays, sodium benzoate (SBA, with the chemical symbol NaC₇H₅O₂ or C₆H₅CO₂Na) is one of the most commonly employed. When dissolved in water, SBA exists in the form of sodium salt of benzoic acid, but it will be converted into active benzoic acid under acidic conditions, thus providing the same preservation mechanism as benzoic acid. However, SBA has much higher water solubility than benzoic acid and is more stable in the atmosphere. Consequently, it is much more commonly used (Hamzah, Yusof, Salleh, & Bakar, 2011). SBA is a particularly effective preservative since it has strong lipophilicity, and hence easily penetrates the cell membrane and enters the cell. It then interferes with the permeability of microbial cell membranes, such as those of mold and bacteria, thereby inhibiting the absorption of the amino acid by the cell membrane and providing a strong antiseptic effect (del Olmo, Calzada, & Nuñez, 2017). Although SBA has a strong toxicity to microorganisms, the toxicity of its sodium salt is very low. Nonetheless, the concentration of SBA in foods must be carefully controlled to avoid damaging human health. Thus, the International Programme on Chemical Safety (IPCS) of the World Health Organization (WHO) has specified an acceptable daily intake (ADI) of just 5 mg/kg body weight (WHO, 2000).

Many analytical methods are available for detecting SBA or benzoic acid, including high performance liquid chromatography (HPLC) (Le, Ngo, Le, & Huong, 2019), liquid chromatography (LC) (Özgür & Kasapoğlu 2019), capillary electrochromatography (CEC) (Tang et al., 2017), surface-enhanced Raman scattering (SERS) (Xue et al., 2020), spectrophotometry (Fujiyoshi et al., 2018, Wang et al., 2020), ion chromatography (IC) (Wang et al., 2013), paper spray mass spectrometry (PS-MS) (Yu et al., 2018), and immunoassays (Li et al., 2018). (For the comparison of the properties of several analytical methods for SBA determination, please refer to Supplementary Materials Table S1). However, these methods are expensive and slow, and often require additional extraction or sample preparation steps, which further increase the analysis time. In addition, they usually require the services of trained operators. Therefore, the use of these methods is limited to well-equipped laboratories manned by professional staff. In other words, low-cost, reliable and straightforward methods for the rapid screening of SBA in food and beverage samples are still lacking.

Microfluidic device analysis systems, also known as Lab-on-a-Chip (LOC) or Micro-Total Analysis Systems (µTAS), are a rapidly growing field (Yang et al., 2018, Chen et al., 2018, Lee and Fu, 2018, Ravanfar et al., 2018, Tseng et al., 2018, Loo et al., 2019, Nguyen et al., 2019a, Hsiao et al., 2020) with many advantages over traditional benchtop systems, including reduced sample and reagent volumes ranging from a few microns to several millimeters, a lower cost, a simpler operation, a faster response time, and better portability. Microfluidic devices have found widespread use in many different fields recently, including biochemical analysis, food processing, food diagnostics, chemical processing, pharmaceutical research and development, chemical synthesis, and environmental testing (Liu et al., 2017, Liu et al., 2018a, Yang et al., 2018a, Dayao et al., 2019, Iranifam and Al Lawati, 2019, Nguyen et al., 2019b, Kamankesh et al., 2020). Microfluidic-based analysis equipment based on rapid detection, convenience, and low cost is generally considered to be superior to traditional food safety and quality analysis methods. Accordingly, the development of microfluidic devices and platforms for food testing purposes has attracted considerable attention in the recent literature. For example, Wu et al. (Wu et al., 2020) presented a rapid microfluidic detection platform using a micro-spectrometer detection device for the concentration measurement of preservatives in food. The feasibility of the device was demonstrated by comparing the concentration measuring results acquired for 15 commercial food samples with those obtained using conventional macroscale methods. It was shown that the detection error of the proposed method was less than 7.5%. The general versatility of the platform was further demonstrated by performing the concentration measurement of sulfur dioxide, sorbic acid, formaldehyde, and benzoic acid samples. The corresponding determination coefficients were found to be R² = 0.9933, 0.9956, 0.9968 and 0.9961, respectively.

Microfluidic paper-based analytical devices (μPADs) (Guzman et al., 2018, Ma et al., 2018, Liu et al., 2018b, Yang et al., 2018b, Fu et al., 2019, Trofimchuk et al., 2020) provide a particularly attractive solution for analytical applications due to their low cost, ease of fabrication, simple operation and absence of external driving systems (Fu and Wang, 2018, Kung et al., 2019, Zhu et al., 2019). Hence, the current study presents a MCA system for SBA concentration measurement, which consists of a paper-microchip and a self-made smart analysis device comprising a micro-temperature controller, a power source, and a detection box integrated with a CMOS device, a WiFi transmission system, and a cell-phone installed with self-developed RGB analysis software. In the presented method, the sample of interest is distilled and is then reacted with NaOH solution and calcinated to form a nitro compound. The compound is dripped onto the reaction area of the paper microchip and undergoes a colorimetric reaction with two embedded regents (acetophenone and acetone) under the effects of heating at 120 °C for 20 min. The reaction image is captured by the CMOS device and transmitted to a smartphone over a WiFi connection. Finally, use the self-developed RGB software installed on the cell-phone to obtain the SBA concentration. The feasibility of the current system is demonstrated by comparing the measurement results acquired for the SBA concentrations of 15 commercially available food and beverage samples with those obtained using two conventional macroscale methods.