Penentuan Kadar Fe Serum 2021

Summary

This video provides a detailed guide on the chemistry practical for determining the vitamin V (Vitamin A) level in blood serum. It covers the step-by-step process, starting from sample preparation with various chemical solutions, including HCl, trichloroacetic acid, and ortho-phenanthroline. After completing the chemical reactions and centrifugation, absorbance readings are taken using a spectrophotometer to analyze the results. The video emphasizes the importance of precise measurement and the calculation of serum concentration using a standard curve and equation. It's an informative tutorial for students in biochemistry or laboratory techniques.

Takeaways

• 😀 The practical session focuses on determining the 'V' concentration in serum using a chemical reaction.
• 😀 In an acidic environment, 'V' is released from its complex bond with globulin.
• 😀 'V' is reduced by hydrogen on compounds, and the reduction product reacts with ortho-phenanthroline to form a red complex.
• 😀 The red complex formed is measured using a spectrophotometer at a wavelength of 506 nm.
• 😀 The materials used in the experiment include HCl, trichloroacetic acid, potassium acetate, hydroquinone, ortho-phenanthroline, and standard SV400 solution.
• 😀 Reaction steps involve mixing specific volumes of serum, standard solution, HCl, and other reagents in labeled test tubes.
• 😀 The mixture is allowed to incubate at room temperature for an hour before adding trichloroacetic acid (BCA) solution.
• 😀 After incubation, centrifugation is performed at 2000 rpm for 15 minutes to separate the supernatant.
• 😀 The supernatant is transferred to a new tube and further reagents are added, including potassium acetate, hydroquinone, and ortho-phenanthroline.
• 😀 Absorbance is measured with a spectrophotometer at 500 nm, and a standard curve is generated for further calculations of 'V' concentration.

Q & A

What is the main topic of the practical experiment discussed in the video?

The main topic is the determination of V in serum using an abrasive reaction.

How is V released from the complex in the experiment?

V is released from the complex due to the acidic environment, where it is freed from the binding effect of globulin.

What is the role of hydrogen on in the experiment?

Hydrogen on reduces FB, which then reacts with orthophenanthroline to form a red complex.

What instrument is used to measure the absorbance of the complex?

A spectrophotometer is used to measure the absorbance of the red complex at a wavelength of 506 nm.

What is the purpose of adding hydrochloric acid (HCl) in the experiment?

The hydrochloric acid (0.35 normal) is added to help release V from the complex during the acidic stage of the process.

How long is the reaction allowed to sit at room temperature after adding HCl?

The reaction is allowed to sit at room temperature for one hour after adding the HCl solution.

What steps are involved after adding the BCA solution to the reaction mixture?

After adding BCA, the solution is vortexed, allowed to stand at room temperature for 15 minutes, then centrifuged at 2000 rpm for 15 minutes.

Why is the supernatant carefully transferred after centrifugation?

The supernatant is carefully transferred to avoid disturbing the precipitate and to ensure accurate results.

What is the significance of the standard curve created with the standard solutions?

The standard curve is used to correlate the absorbance values to the concentration of V in the serum, and the coefficient of correlation should be close to 1 for accurate results.

How is the final concentration of V in the serum determined?

The concentration of V in the serum is determined by substituting the absorbance value of the sample into the equation derived from the standard curve.