Question

An investigator adds palmitate uniformly labeled with tritium \(\left({ }^{3} \mathrm{H}\right)\) to a specific activity of \(2.48 \times 10^{8}\) counts per minute \((\mathrm{cpm})\) per micromole of palmitate to a mitochondrial preparation that oxidizes it to acetyl-CoA. She then isolates the acetyl-CoA and hydrolyzes it to acetate. The specific activity of the isolated acetate is \(1.00 \times 10^{7} \mathrm{cpm} / \mu \mathrm{mol}\). Is this result consistent with the \(\beta\) oxidation pathway? Explain. What is the final fate of the removed tritium? (Note: Specific activity is a measure of the degree of labeling with a radioactive tracer expressed as radioactivity per unit mass. In a uniformly labeled compound, all atoms of a given type are labeled.)

Short answer

Yes, consistent with β oxidation; tritium lost during oxidation as water or other metabolites.

Step-by-step solution

Understanding the Problem

We need to determine whether the decrease in specific activity from palmitate to acetate is consistent with the \( \beta \) oxidation pathway. We are given the specific activity of palmitate and the specific activity of the resulting acetate.

Analyzing β Oxidation Pathway

In the \( \beta \) oxidation pathway, each cycle of oxidation removes a 2-carbon acetyl CoA unit from the fatty acid, reducing the chain length. For palmitate (C16), seven cycles are needed, yielding eight acetyl CoA units.

Calculating Expected Decrease

Assuming complete oxidation of palmitate (C16) into acetyl CoA units, each acetyl CoA carries a fraction of the total specific activity of palmitate. Specifically, the total specific activity is divided among the initial eight acetyl CoA units.

Calculate Theoretical Value

Given the specific activity of palmitate as \(2.48 \times 10^8\, \mathrm{cpm/\mu mol}\), and that palmitate yields 8 acetyl CoA molecules, the specific activity should theoretically be \((2.48 \times 10^8 \; \text{cpm/\mu mol})/8 = 3.10 \times 10^7 \; \mathrm{cpm/\mu mol}\).

Comparing Experimental and Theoretical Values

The experimental specific activity of acetate is \(1.00 \times 10^7 \mathrm{cpm/\mu mol}\), which is lower than the theoretically calculated \(3.10 \times 10^7 \mathrm{cpm/\mu mol}\).

Conclusion and Explanation

The specific activity decrease is consistent with \( \beta \) oxidation, as tritium is likely lost as \(\text{water or} \; \text{other metabolites due to the oxidation }\) \text{process itself or the dilution factor of unlabeled acetyl CoA.}

Key concepts

Fatty Acid Metabolism

Fatty acid metabolism is a major biochemical process in the human body, mainly responsible for breaking down fats into energy. This process occurs primarily in the mitochondria of cells. The fats are stored as triglycerides, which are then hydrolyzed into free fatty acids and glycerol. Once in the mitochondria, fatty acids undergo a series of reactions known as the β oxidation pathway.

In the β oxidation process, fatty acids such as palmitate—a common C16 fatty acid—are broken down into smaller two-carbon fragments called acetyl-CoA. This process is essential for energy production, especially during periods of low blood glucose levels, as the acetyl-CoA units can enter the citric acid cycle to generate ATP.

Key steps in fatty acid metabolism include:

• Activation: Fatty acids are converted into fatty acyl-CoA molecules.
• Transport: These molecules are transported into the mitochondria via a special carnitine shuttle system.
• Oxidation: Fatty acyl-CoA undergoes a series of oxidation reactions, releasing energy and shortening the fatty acid chain.

Understanding fatty acid metabolism is crucial, as disruptions in this process are linked to various metabolic disorders.

Radioactive Tracing in Biochemistry

Radioactive tracing is a powerful tool in biochemistry that helps scientists study metabolic pathways and enzymatic activity. By incorporating radioactive isotopes like tritium ( ^{3} H) into molecules, researchers can track where the isotopes go and how they are transformed in the body.

In the context of fatty acid metabolism, tritium-labeled palmitate provides insights into the efficiency and mechanisms of the β oxidation pathway. By measuring the specific activity, or the radioactivity per unit mass, scientists can observe how tritium from palmitate is distributed as the molecule is broken down.

Benefits of using radioactive tracing include:

• Precision: Allows tracking of specific atoms within complex biochemical processes.
• Quantitative Analysis: Helps determine the extent of metabolic reactions by comparing expected and observed radioactivity.
• Mechanistic Insights: Reveals the fate of atoms during metabolic transformations.

Despite its efficacy, it's important to handle radioactive materials with care due to their inherent hazards.

Palmitate Oxidation

Palmitate oxidation is a specific example of fatty acid metabolism focusing on the breakdown of the C16 fatty acid, palmitate. It's pivotal in the study of β oxidation pathways, as palmitate is a common substrate used in research.

The β oxidation of palmitate occurs in several steps, each shortening the fatty acid by two carbon atoms, resulting in the formation of acetyl-CoA. For a single palmitate molecule:

• Multiple cycles (specifically seven) are necessary to completely oxidize it.
• Each cycle generates one acetyl-CoA molecule until a total of eight acetyl-CoA molecules are produced.

As the acetyl-CoA units form, they enter the citric acid cycle, highlighting the critical role of palmitate oxidation in energy production. This process is particularly important in organs such as the heart and liver, where fatty acids are a primary energy source.

Acetyl-CoA Production

Acetyl-CoA is a central metabolite in energy metabolism, serving as a substrate for the citric acid cycle—a key component in the production of ATP, the body's energy currency.

During β oxidation, each cycle removes a 2-carbon segment from fatty acids, converting them into acetyl-CoA. For example, the complete oxidation of palmitate (a 16-carbon fatty acid) results in eight acetyl-CoA molecules.

Functions of acetyl-CoA include:

• Energy Production: Feeds into the citric acid cycle, which generates NADH and FADH2, subsequently used in oxidative phosphorylation to produce ATP.
• Biosynthesis: Acts as a building block for synthesizing various fatty acids, cholesterol, and ketone bodies.

Acetyl-CoA production is pivotal for maintaining energy homeostasis, particularly in fasting states when carbohydrates are sparse. It's a vital junction between catabolic (breakdown) and anabolic (biosynthesis) processes within the cell.