# Freshness Monitoring of Packaged Vegetables
## 4 Intelligent Packaging (IP)
### Sensors
#### 4.1.2 Biosensors
SPR imaging biochips were used to measure change in refractive index, in order to reduce non-specific signals from background [85, 86]. Another approach is the grating-coupled SPR imaging that uses optical diffraction gratings for coupling and providing angle readings, and it is a low cost and wide dynamic range imaging [87, 88, 89, 90]. In [90], an SPR-based protein chip, as illustrated in Figure 9b, was used for the detection of _E. coli O157:H7_, _Salmonella_, and other pathogens. This means that there is a high potential of using such a system in packaging. By integrating this chip as part of the package, the package can be screened with the use of light source and SPR spectroscopy to check if pathogens are present. Furthermore, the response of antigens binding to the immobilized antibodies on Au-substrate was enhanced with the use of G protein [80, 90]. Other popular optical biosensors that are used for pathogens include Raman spectroscopy [91, 92], Fourier transform infrared (FT-IR) [93], and fiber optics [94]. However, these techniques need culturing to increase the biomass of the pathogen before detection. In addition, the difficulty in integrating these as part of the packaging where contact of food with nanoparticles (NP) is unavoidable, makes them suitable for a different stage of supply chain, for instance in pre-packaging. Other techniques developed for bacterial detection as a type of freshness indicator include those based on surface plasmon resonance (SPR) [95], hyperspectral imaging [79], or polymerase chain reaction [96, 97]. However, these methods require expensive equipment that make them non-commercial for use in supermarkets for monitoring packaged vegetables. Figure 9: (**a**) Working principle of surface plasmon resonance (SPR) biosensors. (**b**) Working principle of SPR biosensors chip. Electrochemical biosensors are those in which the binding elements are antibodies and the transducers are electrochemical. The binding response of antibodies to antigens will be converted into an electrical signal with the electrodes. The advantages of this biosensor category are low-cost and the possibility of coupling with other biosensing techniques. However, its sensitivity is less than optical biosensors. This category can be classified according to the output formats as amperometry, impedimetry, potentiometry, or conductivity [80]. Amperometric is when an applied voltage excites the electroactive species, leading to oxidation or reduction and current flow. The higher the concentration of the analyte, the higher is the current produced. This measurement method gives higher sensitivity compared to potentiometry. In a potentiometric biosensor, the voltage between the electrodes for near the zero current is measured and the potential will build-up as a result of the bio-recognition process. A conducitmetric biosensor relies on the use of conductive polymer to turn the analyte into electrical signal. In this type, incorporating nanoparticles (NPs) can increase the conductivity which will enhance the sensitivity of the system. The last one is the impedimetric type which depends on the metabolites of the microbes as a result of redox reactions that will decrease the impedance; that is, the conductance and capacitance increases [30; 80]. Table 2 shows a comparison between the common electrochemical biosensors for pathogens' detection of vegetables.