Purpose
To compare the protein concentration between freshwater fish and marine fish
Objectives
To measure the protein concentration in marine fish and freshwater fish by using Lowry and Biuret method
Procedure
1. 20 g of fish flesh for freshwater fish and marine fish is weighed using analytical balance.
2. After weighing, the fish flesh is sliced into smaller size
3. Smaller size of fish is blend in the blender and Phosphate Buffer saline at 1:10 ratio is added
4. The fish is blended separately between freshwater fish and marine fish.
5. Then it is incubate for 24 hours at 4C in 100 rpm shaker
6. After 24 hours, the sample is filter to obtain the pure stock by using filter funnel and filter paper.
7. Pure stock that is obtained is put into centrifuge tube using micropipette
8. Then the centrifuge tube is centrifuge at 4C, 1400 rpm, 15 minutes
9. After centrifuge, the protein content is measured by using Lowry or Biuret method
10. As for the Biuret method that we used in this experiment, we mix 0.50 mL of protein with 2.50 mL of Biuret reagent.
11. To measure the abosrbance, we use spectrophotometer
12. Then, the stocks is put into a cuvette and we measure the absorbance at 540 nm after 10 minutes.
13. Step 12 is repeated five times for each marine fish and freshwwater fish so that we are able to obtain the average value of the absorbance we needed.
14. Protein absorbance for each sample we used is then calculated by using Biuret equation y=0.1276 In (x) + 0.0346
Results:
Protein results
Discussion:
Based on the experiment that we conducted, we have used ikan Kayu Hitam (Bullet tuna) and Ikan Tilapia Merah (Oreochromis niloticus) to be our sample. Method that we choose to obtain the protein concentration are the Biuret method. From what we had done, the results are Ikan Kayu Hitam has higher protein concentration than Ikan Tilapia Merah, which is 49.6976A for Ikan Kayu Hitam and 27.9147A for Ikan Tilapia Merah.
From what I read, usually freshwater has higher protein concentration rather than marine water. Maybe because marine have higher salt concentration and it causes the fish to lost some of its protein concentration too. But then, our group have different contradict result from this as the marine fish which is ikan kayu hitam has higher protein concentration than freshwater fish ikan talapia merah. It may due to our mistakes on handling the experiments.
What new things that I learned from this experiment ?
I learn to use new apparatus such as micropippete, spectrophotometer, and learn to use centrifuge tube and also using new method which is Biuret method. Well at first, our group was a bit awkward in handlish those apparatus and procedures, but with the help of our lecturer Prof Dr. Shakinaz, we are able to conduct the experiment until the end.
Below are some of the pictures during experiment:
To compare the protein concentration between freshwater fish and marine fish
Objectives
To measure the protein concentration in marine fish and freshwater fish by using Lowry and Biuret method
Procedure
1. 20 g of fish flesh for freshwater fish and marine fish is weighed using analytical balance.
2. After weighing, the fish flesh is sliced into smaller size
3. Smaller size of fish is blend in the blender and Phosphate Buffer saline at 1:10 ratio is added
4. The fish is blended separately between freshwater fish and marine fish.
5. Then it is incubate for 24 hours at 4C in 100 rpm shaker
6. After 24 hours, the sample is filter to obtain the pure stock by using filter funnel and filter paper.
7. Pure stock that is obtained is put into centrifuge tube using micropipette
8. Then the centrifuge tube is centrifuge at 4C, 1400 rpm, 15 minutes
9. After centrifuge, the protein content is measured by using Lowry or Biuret method
10. As for the Biuret method that we used in this experiment, we mix 0.50 mL of protein with 2.50 mL of Biuret reagent.
11. To measure the abosrbance, we use spectrophotometer
12. Then, the stocks is put into a cuvette and we measure the absorbance at 540 nm after 10 minutes.
13. Step 12 is repeated five times for each marine fish and freshwwater fish so that we are able to obtain the average value of the absorbance we needed.
14. Protein absorbance for each sample we used is then calculated by using Biuret equation y=0.1276 In (x) + 0.0346
Results:
Protein results
Discussion:
Based on the experiment that we conducted, we have used ikan Kayu Hitam (Bullet tuna) and Ikan Tilapia Merah (Oreochromis niloticus) to be our sample. Method that we choose to obtain the protein concentration are the Biuret method. From what we had done, the results are Ikan Kayu Hitam has higher protein concentration than Ikan Tilapia Merah, which is 49.6976A for Ikan Kayu Hitam and 27.9147A for Ikan Tilapia Merah.
From what I read, usually freshwater has higher protein concentration rather than marine water. Maybe because marine have higher salt concentration and it causes the fish to lost some of its protein concentration too. But then, our group have different contradict result from this as the marine fish which is ikan kayu hitam has higher protein concentration than freshwater fish ikan talapia merah. It may due to our mistakes on handling the experiments.
What new things that I learned from this experiment ?
I learn to use new apparatus such as micropippete, spectrophotometer, and learn to use centrifuge tube and also using new method which is Biuret method. Well at first, our group was a bit awkward in handlish those apparatus and procedures, but with the help of our lecturer Prof Dr. Shakinaz, we are able to conduct the experiment until the end.
Below are some of the pictures during experiment: