Quantification of Total Sulfite in Shrimps by BIOFISH 300 SUL Method: First Action 2021.09.

BACKGROUND: Sulfite is the oldest and most widely used additive in our food supply with antioxidant and preservative properties. Due to its allergenic-like reactions and other adverse health effects, its use is regulated by international regulatory bodies. Therefore, food industries as well as regulatory laboratories must ensure that the maximum concentration of sulfite permitted is not exceeded. The AOAC INTERNATIONAL-approved official method for the quantification of sulfites is the Optimized Monier-Williams Method (AOAC Official Method 990.28), which consists of a time-consuming titration. OBJECTIVE: The present study aims to demonstrate the reliability of the BIOFISH 300 SUL, a simple, fast, and accurate method, as an alternative, for the quantification of total sulfites in shrimp, with a lower LOQ than that of the OMA 990.28, set at 10 mg/Kg. METHODS: The BIOFISH 300 SUL method is a highly specific biosensor based on its proprietary enzyme-based electrode, for the rapid quantification of total sulfite. The test kit consists of an electrochemical reader (biosensor BIOFISH 300) and disposable electrodes (Biotest) that are capable of providing an electrical signal proportional to the amount of sulfite in the sample analyzed. The method mainly consists of the extraction of sulfite from the solid matrix in an aqueous solution, and its subsequent quantification by the device in less than 3 min. RESULTS: Comparative studies between BIOFISH 300 SUL and OMA 990.28 were conducted for naturally contaminated and spiked samples of raw and boiled shrimp with sulfite levels covering the 7-150 mg/kg range in order to determine linearity, recovery, repeatability, intermediate reproducibility, and accuracy. CONCLUSION: The BIOFISH 300 SUL method demonstrated high accuracy and precision for the whole range of quantification (7-150 mg/kg). Its ease of use and fast response make it the ideal technology to be implemented by the industry. HIGHLIGHTS: BIOFISH 300 SUL was adopted as a First Action Official MethodSM by the AOAC Expert Review Panel for Sulfites in Seafood Methods in February 2021 after rigorous review.

Quantification of Lactose in Lactose-Free and Low-Lactose Milk and Milk Products by BIOMILK 300/3000 Lac, Collaborative Study: Final Action 2020.09.

BACKGROUND: In April 2020, the BIOMILK 300 LAC method for the quantification of lactose in lactose-free and low-lactose dairy products was adopted as First Action Official Method of AnalysisSM2020.09 by AOAC INTERNATIONAL. To test the reproducibility of the method, as the last step toward the Final Action status, a collaborative study was organized by BIOLAN Microbiosensores. OBJECTIVE: Fifteen collaborators within the European Union took part in this study, where nine different samples were sent as duplicate blind test portions for lactose determination. The data obtained were used to determine method repeatability and reproducibility and also to validate the new version of the biosensor. METHOD: The test method is based on the direct enzymatic recognition-electrochemical detection of trace levels of lactose over a wide range of dairy samples by means of the BIOMILK 300 biosensor, in less than 5 min, and without intricate sample pretreatments. RESULTS: All samples resulted in a repeatability relative standard deviation (RSDr) of <8.1% and a reproducibility relative standard deviation (RSDR) of 14% maximum, meeting requirements from Standard Method Performance Requirement (SMPR®) 2018.09. CONCLUSIONS: The method has proved to give reproducible results for the quantification of lactose in lactose-free and low-lactose dairy products. HIGHLIGHTS: On the basis of these results, the enzymatic amperometric biosensor method developed by BIOLAN Microbiosensores was adopted as a Final Action Official Method in July 2022.

Modification of BIOFISH 300 HIS for Determination of Histamine in Fish and Fish Products: AOAC Performance Tested MethodSM 051604.

BACKGROUND: The BIOFISH 300 HIS method for the quantification of histamine in fish and fish products was validated by the AOAC Research Institute and granted Performance Tested MethodSM certification in 2016. The method is based on the use of an electrochemical reader (BIOFISH 300 device) together with disposable gold electrodes (Biotest) modified with a specific enzyme that oxidizes the histamine molecules and produces a detectable and quantifiable electric current signal. It was validated for raw fish (tuna, mackerel, sardine, and anchovy), cooked fish (tuna), canned fish (tuna in water, tuna in oil, mackerel in tomato sauce, and pickled sardine), and salted fish (canned salted anchovy). The validated ranges were 30-150 mg/kg histamine for canned salted anchovy and 10-200 mg/kg histamine for all other matrices. OBJECTIVE: The objective of the present report is to validate some method modifications, namely the use of a new reader (BIOFISH 3000), the inclusion of new quantification ranges, new matrixes in the certified claims, a histamine reference solution (Verifying Solution) that will enable the user to perform a system verification, and also the inclusion of the kit ASC1-10 for the elimination of ascorbic acid from sample extract. METHODS: New quantification ranges can be obtained by diluting the sample appropriately into the measurement cuvette, thereby adjusting the concentration to the internal calibration range of the device. New matrixes such as fish meal, preserved salted anchovy, and fish with added ascorbic acid were validated. The new digitalized reader enables new functionalities such as the use of an intuitive app and cloud storage of the results, with no changes in the data acquisition and processing. As the use of the new reader does not imply any change at the electrochemical readout level, all assays were performed using the BIOFISH 3000 device. RESULTS: The method was shown to be linear in all the quantitation ranges configured. Accuracy and recovery studies over a wide range of matrixes showed that the method yields comparable results to the reference method used. Stability and product consistency testing of kit components demonstrated that the system produces accurate results when expiration dates from the manufacturer are met. CONCLUSIONS: All modifications made to the BIOFISH 300 HIS method, PTM 051604, have been validated satisfactorily, under criteria of the AOAC Research Institute. HIGHLIGHTS: The use of the BIOFISH 300/3000 HIS method is ideal as a routine histamine control method for the fish industry. After a simple aqueous extraction of the sample, results are obtained in less than 3 minutes. All data generated by the new BIO3000 biosensor are sent to a web platform, allowing new functionalities to the user. All these technical capabilities represent a competitive advantage over other existing technologies.

Quantification of Lactose in Lactose-Free and Low-Lactose Milk and Milk Products by BIOMILK 300 Lac Biosensor: First Action 2020.09.

BACKGROUND: In 2018, the AOAC Stakeholder Panel on Strategic Food Analytical Methods approved Standard Method Performance Requirement (SMPR®) 2018.009, for lactose in low-lactose or lactose-free milk, milk products, and products containing dairy ingredients, establishing the minimum recommended performance characteristics to be addressed during the evaluation of methods. Subsequently, AOAC INTERNATIONAL opened a call for methods under the Official Method of AnalysisSM program with the aim of finding a candidate method for confirming compliance with regulatory standards and dispute resolution. OBJECTIVE: A biosensor-based analytical method, BIOMILK 300 LAC, was developed by BIOLAN Microbiosensores S.L. (www.biolanmb.com) to rapidly, easily, and accurately quantify lactose in free or low-lactose dairy products. In response to the AOAC call for methods, BIOLAN performed a single laboratory validation of this method against SMPR 2018.009. Several different matrixes were tested, including: milk, sugary plain yogurt, fruit plain yogurt, flavored liquid yogurt, Greek yogurt, cream, soft cheese, infant formula, café latte, chocolate milk, and high-protein milk shake. Evaluated method parameters included: linearity, selectivity, matrix effect, recovery, accuracy, repeatability, intermediate reproducibility, robustness, reagent lot-to-lot consistency, and stability. METHODS: The method is based on the use of the Biotest gold electrode together with the BIOMILK 300 biosensor reader, for the quantification of residual levels of lactose in dairy samples via an enzymatic recognition/electrochemical transduction system. RESULTS: Assay linearity, applicability to different matrixes, recovery, and precision demonstrated that the method is fit for purpose. The method proved to be robust, consistent, and stable, under conditions detailed in the Instructions For Use guide. CONCLUSION: The overall results were within requirements stated by SMPR 2018.009 for low-lactose and lactose-free milk, milk products and products containing dairy ingredients. HIGHLIGHTS: The BIOMILK 300 LAC method enables the quantification of reduced levels of lactose in less than 5 min, without the requirement for expert technicians, toxic solvents or intricate procedures, maintaining a high degree of precision and accuracy in the results. BIOMILK 300 LAC was adopted as a First Action Official MethodSM by the Expert Review Panel of Low-lactose Methods in April 2020 after rigorous review.

Validation of the Biofish-300 HIS Enzymatic Biosensor for the Detection of Histamine in Fishery Products.

The Biofish-300 HIS method is a simple, reliable, and specific enzymatic biosensor for the detection of histamine. This technology is highly specific and selective and allows quantification of histamine in fishery products (fresh/frozen and processed) in a short time frame (2-3 min). Histamine in raw tuna, raw mackerel, raw sardine, raw anchovy, boiled tuna, canned tuna in water, canned tuna in oil, canned mackerel in tomato sauce, canned pickled sardine, and canned salted anchovy was analyzed using a water-based extract. Matrix-specific assay procedures and calibration curves were used to enable analyses to be carried out across multiple sample types. The performance of this assay was examined using samples that were naturally contaminated (reference materials and interlaboratory studies) and spiked with histamine. All data were judged against previously established acceptance criteria. Performance measures were evaluated for linearity, selectivity, matrix, lot consistency, and robustness. Results produced in all performance measures, except robustness, were within acceptable ranges. Out-of-range robustness results reflected deviation in sample volume compared to the standard assay procedures. Positive interferences from the presence of agmatine were shown.

On the arrangements of R6G molecules in organophilic C12TMA/lap clay films for low dye loadings.

Absorption and fluorescence spectroscopies with linearly polarized light are applied to characterize the adsorbed species of rhodamine 6G (R6G) laser dye in ordered organophilic laponite (Lap) clay films for low dye loadings. The organophilic character of the clay was controlled by the number of organic surfactant dodecyl-trimethylammonium (C12TMA) cations intercalated into the interlayer space of the clay. Experimental results suggest that for moderate to high surfactant contents (>70% of the total cation exchange capacity, CEC, of the clay) the accessibility of the interlayer space for R6G molecules is reduced. In these cases, the first stage in the adsorption of R6G molecules is at the external surface, where dye molecules can self-associate even for very low dye loading (around 0.1%CEC), probably for the limitation of the external surface area. The presence of a nonfluorescent H-type aggregate with a short-displaced coplanar structure and a fluorescent oblique head-to-tail J aggregate is reported. The inclined structure of this J aggregate is stabilized by the surfactant molecules at the external surface.

Application of fluorescence with polarized light to evaluate the orientation of dyes adsorbed in layered materials.

A new method of fluorescence polarization is applied to evaluate the angle of the preferential orientation of Rhodamine 6G (R6G) dye adsorbed in supported thin films of Laponite (Lap) clay. The method is based in the determination of the fluorescence dichroic ratio, obtained from the recorded fluorescence spectra with the detection polarizer horizontally and vertically oriented, as a function of the twisted angle of the film around its vertical axis, keeping the excitation polarizer in a fixed direction. The validity of the method is checked by comparing the experimental results obtained with both vertically and horizontally polarized excitations to that previously provided by absorption spectroscopy with linearly polarized light. A preferential orientation angle with respect to the normal to the clay layer of 62 degrees is derived for R6G monomers adsorbed in Lap films.