Ketone bodies

Formation and fate of ketone bodies

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CONTENTS



Topic Page no.
 Introduction …………………………………………………………... .. 1

 Ketogenesis …………………………………………………………… 1-3

 Utilisation of ketone bodies …………………………………………... . 3-4

 Regulation of ketogenesis …………………………………………… . 4

 General overview of ketone bodies ………………………………….. 5

 Overproduction in diabetes mellitus and starvation………………….. 5 -6

 Jamaican vomiting sickness …………………………………………. 6-7

 Rothera’s test …………………………………… …………………… 7

 References …………………………………………………………… 8

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Ketone bodies- Introduction
The compounds namely acetone, acetoacetate and β-hydroxybutyrate (or 3-Hydroxybutyrate)
are known as ketone bodies. Only the first two are true ketone* bodies while β-hydroxybutyrate
does not possess a keto (C=O) group. Ketone bodies are water soluble and energy yielding.
Acetone, however, is an exception, since it cannot be metabolized.

Fig.1 Ketone bodies.
In humans and most other mammals, acetyl-CoA formed in the liver during
oxidation of fatty acids can either enter the citric acid cycle or undergo conversion to the “ketone
bodies,” acetone, acetoacetate, and β-hydroxybutyrate, for export to other tissues. (The term
“bodies” is a historical artifact; the term is occasionally applied to insoluble particles, but these
compounds are quite soluble in blood and urine.) Acetone, produced in smaller quantities than the
other ketone bodies, is exhaled. Acetoacetate and β-hydroxybutyrate are transported by the blood
to tissues other than the liver (extrahepatic tissues), where they are converted to acetyl-CoA and
oxidized in the citric acid cycle, providing much of the energy required by tissues such as skeletal
and heart muscle and the renal cortex. The brain, which preferentially uses glucose as fuel, can
adapt to the use of acetoacetate or β-hydroxybutyrate under starvation conditions, when glucose is
unavailable. The production and export of ketone bodies from the liver to extrahepatic tissues
allows continued oxidation of fatty acids in the liver when acetyl-CoA is not being oxidized in the
citric acid cycle.

Ketogenesis
The synthesis of ketone bodies is termed as ‘ketogenesis’. Ketogenesis occurs only in the
mitochondria of liver cells. It occurs when there is a high rate of fatty acid oxidation in the liver.
The enzymes for ketone body synthesis are located in the mitochondrial
matrix. Acetyl CoA, formed by oxidation of fatty acids, pyruvate or some amino acids, is the
precursor for ketone bodies. Ketogenesis occurs through the following reactions:
i. Two moles of acetyl CoA condense to form acetoacetyl CoA. This reaction is catalyzed by
thiolase, an enzyme involved in the final step of β-oxidation. Hence, acetoacetate synthesis
is appropriately regarded as the reversal of thiolase reaction of fatty acid oxidation.
ii. Acetoacetyl CoA combines with another molecule of acetyl CoA to produce β-hydroxy β-
methyl glutaryl CoA (HMG CoA). HMG CoA synthase, catalysing this reaction, regulates
the synthesis of ketone bodies.
iii. HMG CoA lyase cleaves HMG CoA to produce acetoacetate and acetyl CoA.

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v. Acetoacetate can undergo spontaneous decarboxylation to form acetone.
*The term ‘ketones’ should not be used because 3-hydroxybutyrate is not a ketone and
there are ketones in blood that are not ketone bodies e.g. pyruvate, fructose.

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iv. Acetoacetate can undergo spontaneous decarboxylation to form acetone.
vi. Acetoacetate can be reduced by a dehydrogenease to β-hydroxybutyrate.



Fig. 2 Ketogenesis.
β-Hydroxy-β-methylglutaryl CoA (HMG-CoA) is an intermediate in the pathway
of ketogenesis. Enzymes responsible for ketone body formation are associated mainly with the
mitochondria. Two acetyl-CoA moleccules formed in β-oxidation condense with another to form
acetoacetyl-CoA by a reversal of the thiolase reaction. Acetoacetyl-CoA, which is the starting
material for ketogenesis, also arises directly from the terminal four carbons of a fatty acid during
β-oxidation. Condensation of acetoacetyl-CoA with another molecule of acetyl-CoA by β-
hydroxy-β-methylglutaryl-CoA lyase then causes acetyl-CoA to split off from the HMG-CoA,

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leaving free acetoacetate. The carbon atoms split off in the acetyl-CoA, leaving free acetoacetate.
The carbon atoms split off in the acetyl-CoA molecule are derived from the original acetoacetyl-
CoA molecule. Both enzymes must be present in mitochondria for ketogenesis to take place.
This occurs solely in liver and rumen epithelium. β-Hydroxybutyrate
is quantitatively the predominant ketone body present in the blood and urine in ketosis.

Utilisation of ketone bodies (Catabolism)
The utilization of ketone bodies by the extrahepatic tissues requires the
activity of the enzyme thiolase. The catabolism of ketone bodies can be summarized by the
following steps:
i. Conversion of β-hydroxybutyrate to acetoacetate is necessary as a first step.
ii. Thiophorase then catalyzes transfer of CoA to acetoacetate to produce acetoacetyl
CoA.
a. Succinyl CoA is the donor for this transesterification reaction.
b. Acetoacetyl CoA is then split into two molecules of acetyl CoA, which can
enter the TCA cycle for fuel.
c. The liver does not contain thiophorase, so it cannot use ketone bodies as fuel.
Therefore, only those organs that express thiophorase can utilize ketone bodies
for energy.


Fig. 3 Catabolism of ketone bodies.

The acetyl CoA thus formed is oxidized in the citric acid cycle. If the blood level is
raised, oxidation of ketone bodies increases until, at a concentration of approximately
12 millimole/litre, they saturate the oxidative machinery.

The ketone bodies, being water soluble, are easily
transported from the liver to various tissues. The two ketone bodies- acetoacetate and
β-hydroxybutyrate serve as important sources of energy for the peripheral muscles such
as skeletal muscles, cardiac muscle, renal cortex, etc. The tissues which lack
mitochondria (e.g. erythrocytes) however, cannot utilize ketone bodies.

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Fig. 4 Transport of ketone bodies from the liver and pathways of utilization and oxidation in
extrahepatic tissues.

Regulation of ketogenesis
Ketogenesis is regulated at the following three crucial steps:
1. Control of free fatty acid (FFA) mobilization from adipose tissues.
2. The activity of carnitine palmitoyltransferase-I (CPT-I) in liver, which determines the
proportion of fatty acid flux that is oxidized rather than esterified.
3. Partition of acetyl-CoA between the pathway of ketogenesis and the citric acid cycle.

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Fig. 5 Regulation of ketogenesis.
General overview of ketone bodies
The general overview of ketone bodies can be better understood from the
chart given below:


Fig. 6 Formation, utilization, and excretion of ketone bodies (The main pathway is indicated by
the solid arrows).


Overproduction of ketone bodies during diabetes mellitus and starvation
In healthy people, acetone is formed in very small amounts from acetoacetate, which is easily
decarboxylated, either spontaneously or by the action of acetoacetate decarboxylase. Because
individuals with untreated diabetes produce large quantities of acetoacetate, their blood contains
significant amounts of acetone, which is toxic. Acetone is volatile and imparts a characteristic odor
to the breath, which is sometimes useful in diagnosing diabetes.
In untreated diabetes, when the insulin level is insufficient,
extrahepatic tissues cannot take up glucose efficiently from the blood, either for fuel or for
conversion to fat. Under these conditions, levels of malonyl-CoA (the starting material for fatty
acid synthesis) fall, inhibition of carnitine acyltransferase I is relieved, and fatty acids enter
mitochondria to be degraded to acetyl-CoA—which cannot pass through the citric acid cycle
because cycle intermediates have been drawn off for use as substrates in gluconeogenesis. The

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resulting accumulation of acetyl-CoA accelerates the formation of ketone bodies beyond the
capacity of extrahepatic tissues to oxidize them. The increased blood levels of acetoacetate and β-
hydroxybutyrate lower the blood pH, causing the condition known as acidosis. Extreme acidosis
can lead to coma and in some cases death. Ketone bodies in the blood and urine of individuals with
untreated diabetes can reach extraordinary levels—a blood concentration of 90 mg/100 mL
(compared with a normal level of <3 mg/100 mL) and urinary excretion of 5,000 mg/24 hr
(compared with a normal rate of ≤125 mg/24 hr). This condition is called ketosis. Ketosis is mild
in starvation but severe in diabetes mellitus.
Individuals on very low-calorie diets, using the fats stored in adipose tissue as their
major energy source, also have increased levels of ketone bodies in their blood and urine. These
levels must be monitored to avoid the dangers of acidosis and ketosis (ketoacidosis).
These effects lead to major clinical manifestations, including nausea, vomiting,
dehydration, electrolyte imbalance, loss of consciousness and, potentially, coma and death.
A characteristic sign of this condition is a fruity odor on the breath due to expiration of large
amounts of acetone.

Jamaican vomiting sickness
Jamaican vomiting sickness is an acute illness caused by the toxin hypoglycin A,
which is present in unripened fruit of the ackee tree. Hypogylcin A is present in the unripe aril
(external covering of seeds that develops after fertilization as an outgrowth from the ovule stalk)
at levels of over 1000ppm, which falls to less than 0.1ppm in the fully ripened aril.
When ingested, hypoglycin A is metabolized to produce
methylenecyclopropylacetic acid (MCPA). MCPA acts to inhibit the beta-oxidation of fatty acids
in two ways. First, it interferes with the transport of long-chain fatty acids into mitochondria. Also,
it inhibits acyl-CoA dehydrogenase, so that only unsaturated fatty acids can be fully oxidized.
Fatty acids accumulate in the liver. In absence of fatty acid metabolism, the body becomes
dependent on glucose and glycogen for energy.
Once the liver glycogen stores are depleted, the body cannot synthesize
glucose and severe hypoglycemia occurs.
Abdominal discomfort begins two to six hours after eating unripe ackee fruit,
followed by sudden onset vomiting. In severe cases, profound dehydration, seizures, coma and
death may occur. Children and those who are malnourished are more susceptible to the disease.

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Fig. 7 Fruit of Blighia sapida (Ackee fruit)
The ackee fruit (Blighia sapida) is native to West Africa. Although native to West Africa, the use
of ackee in food is especially prominent in Jamaican cuisine. Ackee is the national fruit of
Jamaica. Ackee pods should be allowed to ripen on the tree before picking. Prior to cooking, the
ackee arils are cleaned and washed. The arils are then boiled for approximately 5 minutes and the
water discarded.

Rothera’s test
It is a test for ketone bodies. 5ml of fresh urine is saturated with solid ammonium
sulphate and mixed with 10 drops of freshly prepared 2% sodium nitroprusside solution, which is
then mixed with 10 drops of concentrated ammonia water and allowed to stand for 15 minutes; the
presence of acetoacetic acid or of larger concentrations of acetone is indicated by the development
of a purple-blue color.

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Fig. 8 Rothera’s test.











REFERENCES:

 Satyanarayana, U & Chakrapani, U (2013). Biochemistry 4
th
Edition. New Delhi, India:
Reed Elsevier India Private Limited. 293-295.

 Nelson, DL & Cox, MM (2008). Lehninger Principles of Biochemistry 5
th
Edition. New
York: W.H. Freeman and Company. 666-668.

 Murray, RK, Granner, DK, Mayes, PK & Rodwell, VK (2003). Harper’s Illustrated
Biochemistry 26
th
Edition. USA: The McGraw-Hill Companies. 183-189.

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 MacDonald, RG & Chaney, WG (2007). USML Road Map Biochemistry. USA: The
McGraw-Hill Companies. 113-115.