## 2 Material and Methods
### Physicochemical Properties of Fish Hamburger  
All physicochemical tests were done during the first month after production. A pH meter (Metrohm-744 pH meter, Switzerland) was calibrated using standard buffer solutions and was used to measure the pH of microalgae-fortified and unfortified fish burgers. The water activity (a\({}_{w}\)) of burger samples was determined using SPRINT Novasina Thermoconstanter (TH-500, Switzerland) equipment at 25 \({}^{\circ}\)C. The equipment was previously calibrated according to the calibration procedure of the equipment manufacturer, using the following salts: MgCl\({}_{2}\), NaCl, BaCl\({}_{2}\), and K\({}_{2}\)Cr\({}_{2}\)O\({}_{7}\) (Sigma-Aldrich, St. Louis, MO, USA). All measurements were carried out in triplicate. The infrared absorption spectra of all burger formulations were obtained using the Cary 630 FTIR spectrometer (Agilent Technologies., Santa Clara, CA, USA). The transmission spectra were obtained in the range of 600-4000 cm\({}^{-1}\) at a resolution of 4 cm\({}^{-1}\). An average of 10 scans was carried out for each sample to obtain a spectrum. All of the samples were tested in duplicate with plain KBr pellets as the blank. The acquisition and processing of spectra were carried out using the Agilent MicroLab PC software v.5.3 (Agilent Technologies Inc.). Total solid (TS) content was determined after drying the samples in an air oven (Thermoline Scientific Equipment, Wetherill Park, Australia) at a temperature of 105 \({}^{\circ}\)C to a point where the weight no longer changed. Dried burgers were fired in an electric furnace (Labec Laboratory Pty Ltd., Marrickville, NSW, Australia) at a high temperature of 550 \({}^{\circ}\)C to burn all organic matter and determine the mineral content. Total lipids were extracted using the modified procedure of [PERSON] and Dyer [12] and measured as described by [PERSON] et al. [13]. Briefly, lipids were obtained by using a hand-held homogenizer and extracted with a solvent mixture (chloroform-methanol-water; 2:1:1 _v/v_) at a sample-to-solvent ratio of 1:3 (_w/v_). The lipid contents were determined gravimetrically after oven-drying (80 \({}^{\circ}\)C) the extract overnight. The protein content was fixed according to the classical Kjeldhal procedure with a nitrogen conversion factor of 6.25. Approximately 1 g of raw material was hydrolyzed with 15 mL concentrated sulfuric acid (H\({}_{2}\)SO\({}_{4}\)) containing two copper catalyst tablets (Sigma-Aldrich) in a heat block (Kjeltec system 2020 digestor, Tecator Inc., Herndon, VA, USA) for 2 h. After cooling, H\({}_{2}\)O was added to the hydrolysates before neutralization and titration. The determination of total dietary fiber in burger samples was done using the non-enzymatic gravimetric method (AOAC Official Method 993.21.) as described by [PERSON] et al. [6]. Burger samples (0.5 g) were suspended in 5 mL of distilled water and incubated at 37 \({}^{\circ}\)C for 90 min to solubilize sugars and other water-soluble components. Water-soluble fibers were then precipitated with 20 mL of 95% ethanol. Residue was washed sequentially with 4 mL of 78% ethanol, 2 mL of 95% ethanol, and 2 mL of acetone and then dried at 105 \({}^{\circ}\)C. The total dietary fiber content was calculated as the weight of residue minus the weights of protein and ash. \[\begin{array}{l}\text{Total dietary fiber (\%) = 100\times((weight ${}_{Residue}$-[(weight ${}_{Proteins}$+ weight ${}_{\text{Ash}}$)/100]\times weight ${}_{\text{Residue}}$)/weight ${}_{\text{Sample}}$)}\\ \text{Total dietary fiber (\%) = 100\times((weight ${}_{Residue}$-[(weight ${}_{Proteins}$+ weight ${}_{\text{Ash}}$)/100]\times weight ${}_{\text{Residue}}$)/weight ${  
For the determination of chlorophylls and carotenoids, burger samples (0.2 g) were sonicated in 2 mL of 95% ethanol at 65 \({}^{\circ}\)C for 30 min using an ultrasonic bath (Bandelin Electronic, Berlin, Germany).