Caballero B. (ed.) Encyclopaedia of Food Science, Food Technology and Nutrition. Ten-Volume Set

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outbreaks have been attributed. Its concurrent recov-

ery in vomitus of patients and in feces of subjects who

had eaten the implicated food, even if asymptomatic

may be important for a better understanding of the

outbreaks. (See Food Poisoning: Tracing Origins and

Testing.)

0008 A factor that may cause ambiguous interpretations

is that samples of the suspected food are generally

contaminated after they were eaten, and the rapidly

growing cultures can outgrow and completely mask

the causative organism.

0009 Information about the diffusion of these Entero-

bacteriaceae in nature may be found in the preceding

article.

0010 Members of genera Citrobacter, Edwardsiella,

Enterobacter, Hafnia, Moellerella, Proteus, and Kleb-

siella have been associated with food poisoning.

Clinical Features and Characteristics

Citrobacter

0011 In a few food-poisoning epidemics, Citrobacter has

been isolated from symptomatic patients with diar-

rhea, from food handlers, and, occasionally, from

implicated food.

0012 In the cases where the suspected organism has been

identified, the clinical picture of the disease varied

from mild to intense diarrhea, with abdominal

cramps, nausea, vomiting, fever, chills, headache,

tenesmus, dizziness, and fainting.

0013 Citrobacter has been identified as a suspected agent

of outbreaks in milk, meat, spaghetti, vegetables, and

liver sausage.

Edwardsiella

0014 Edwardsiella tarda has been isolated from the feces

of both asymptomatic and symptomatic individuals

with diarrhea. This organism has also been identified

from the feces of food-industry workers, but its role in

foodborne disease outbreaks is unknown.

Enterobacter

and

Hafnia

0015 In 1923 and 1932, two food-poisoning epidemics

were associated with organisms of the cloacae–

aerogenes group, but the symptoms and incubation

periods were consistent with staphylococcal intoxi-

cations.

0016 Hafnia alvei, previously known as Enterobacter

alvei, has been associated with an outbreak of mild

gastroenteritis where milk was suspected as the

vehicle. Other contemporary cases have been reported

in the same city where the hospital was located, and a

common source that served the implicated food has

been identified. In this investigation, the analysis of the

serotype showed that the strains isolated from the

hospital and urban cases were identical.

0017A fatal case of food poisoning has been reported in

which a man became ill after eating a stew. The

incubation period of illness was 4 days. Hafnia alvei

was isolated from the patient but not from the

suspected food, which was discarded.

0018The role of the organisms in the Enterobacter and

Hafnia genera as potential intestinal pathogens is sup-

ported by, besides clinical and epidemiological

reports, the observation that Enterobacter cloacae

has given a positive response in the isolated rabbit

ileal-loop test. The toxin implicated has been purified,

and the analysis has reported its similarity to the heat-

stable enterotoxin (ST) produced by Escherichia coli.

Moellerella

0019Moellerella wisconsensis, from the newly described

genus Moellerella, has been frequently isolated from

the feces of symptomatic individuals. Most of the

isolates have been from children with diarrhea. Its

role in foodborne disease outbreaks is unknown, but

it seems to be an intestinal potential pathogen for

humans.

Proteus

0020The presence of strains belonging to the Proteus

genus is normal in human and animal intestinal

tracts. In the past, many cases of food poisoning, in

which Proteus was suspected as potential causative

agent, have been reported before the role of staphylo-

cocci and Clostridium perfringens in determining

acute gastroenteritis was known. Nevertheless, food-

borne disease outbreaks have been reported where

P. mirabilis has been isolated from feces of subjects

with diarrhea in the absence of Salmonella spp. and

staphylococci.

0021The clinical features included nausea, vomiting,

diarrhea, abdominal cramps, collapse, cyanosis, and

sleepiness. Fever was reported in only a few cases.

The incubation period was 3–5 h after eating the

implicated food. In one of these epidemics, P. vulgaris

has been isolated from inadequately cooked brawn

prepared from brine-pickled meat. Another reported

outbreak involved a suspected P. mirabilis strain that

was isolated in large numbers from the vomitus and

feces of symptomatic subjects and from a suspected

baked ham. The clinical features and incubation

periods suggested the hypothesis of staphylococcal

intoxication, but the suspected food was not analyzed

for staphyloenterotoxin.

0022Histamine fish poisoning was responsible for 182

among 2745 (6.6 %) foodborne disease outbreaks

ESCHERICHIA COLI

/Food Poisoning by Species other than

Escherichia coli

2167

of known etiology reported to the Centers for Disease

Control (USA) from 1972 to 1986. (See Histamine.)

0023 In one outbreak, a strain of Klebsiella pneumoniae,

capable of producing large quantities of histamine,

was isolated from tuna. In general, Proteus spp. are

responsible for histamine food poisoning.

0024 The clinical features of the illness are characterized

by symptoms resembling those of a histamine reac-

tion within 5–60 min of eating the implicated fish.

Burning of the mouth and throat, headache, dizzi-

ness, and flushing are common. Other symptoms in

descending order of frequency are abdominal cramps,

nausea, vomiting and diarrhea. Urticaria and bronch-

ospasm may also occur. Symptoms of intoxication

are self-limiting and usually resolve in a few hours;

in general, no therapy is required. Histamine fish

poisoning is reported all the year round primarily

from coastal areas.

Klebsiella

0025 Klebsiellae are well-established opportunistic patho-

gens and are not normally considered as causative

agents of intestinal disease. Nevertheless, these organ-

isms have been occasionally reported as potential

pathogens in acute gastroenteritis. Klebsiella pneu-

moniae, like Enterobacter cloacae, causes a positive

rabbit ileal-loop response, and a heat-stable entero-

toxin similar to that produced by E. coli has been

purified and characterized.

0026 A possible outbreak of foodborne Klebsiella infec-

tion has been reported in which 190 people who had

eaten at a boarding school developed gastroenteritis.

The suspected food was eaten during lunch. All

the subjects developed watery diarrhea with violent

abdominal pain 10–15 h after eating the implicated

meal. Fever was not reported, and all the subjects

recovered within 24 h without antibiotic therapy.

Klebsiellae were identified from the soup and beef

cooked in rice. Klebsiella was the predominant

organism in stool culture and has been also identi-

fied in high numbers from the suspected food. How-

ever, these observations cannot establish these

organisms as primary causative agents of the out-

break because the food was improperly stored prior

to sampling.

0027 Klebsiellae are very important agents of opportun-

istic infections in hospitalized patients. Food pre-

pared and supplied by hospital kitchens is at risk of

colonization by Enterobacteriaceae. Klebsiellae were

identified in high numbers from 12 foods in a hospital

kitchen and have been isolated from vegetables, seeds,

sugar, and milk. The more probable origins of these

contaminants are the incoming foods, but spread of

these microorganisms in the hospital environment

may also occur by means of the colonized subjects.

Other Enterobacteriaceae, such as Enterobacter, Ser-

ratia, Citrobacter,andProteus, are also commonly

found as contaminants in food prepared in hospital

kitchens. Hospitalized patients may be colonized by

these organisms and may have a higher risk of de-

veloping opportunistic infections.

Detection

0028The detection and identification of Enterobacteriaceae

in suspected specimens are not difficult. Appropriate

specimens for laboratory diagnosis include feces,

vomitus, gastric contents, and suspected and nonsus-

pected food. Cultures of the food preparation environ-

ment and food handlers may also be useful.

0029The laboratory should be alerted to investigate also

the organisms considered part of the normal flora, as

well as the common agents of food poisoning. The

quantity of the isolate per gram from the incriminated

food item is important, but the specimen analyzed

may be derived from properly stored food after it

was served. The count of the organism in feces of

the patients is not significant; it is more important

to demonstrate the common source of origin of the

isolates. Serotyping, susceptibility to a series of anti-

biotics, and bacteriocin analysis (as well as other

methods) are utilized in confirming that isolates are

derived from a common source. Plasmid analysis may

also be useful.

0030Detailed information about isolation and identifi-

cation procedures is given in the preceding article.

(See Escherichia coli: Occurrence and Epidemiology

of Species other than Escherichia coli.)

Treatment

0031The food poisoning associated with these strains of

Enterobacteriaceae is generally self-limiting. Support-

ive measures are the mainstay of therapy in these

cases. Fluid replacement is the major consideration

in all of these illnesses.

0032Antibiotics play no role, either in therapy or in

prophylaxis. Antiperistaltic agents are of little benefit

in controlling diarrhea and are contraindicated in pa-

tients with symptoms that suggest the responsibility of

an invasive pathogen (fever, or leucocytes in feces).

0033The syndrome associated with histamine fish

poisoning may be relieved by antihistamines.

Prevention and Control

0034The mainstay of food-poisoning prevention is the

proper selection, preparation, and storage of food.

Inadequate refrigeration is the single most frequent

factor responsible for foodborne disease outbreaks.

2168

ESCHERICHIA COLI

/Food Poisoning by Species other than

Escherichia coli

0035 Enterobacteriaceae grow very well in food at tem-

peratures ranging from 4 to 60



C. Growth may be

prevented if cold food is properly refrigerated and if

hot food is held at temperatures above 60



C before

serving.

0036 Poor personal hygiene by food handlers frequently

contributes to contamination. Education of the public,

nurses, physicians, and those working in eating estab-

lishments is crucial in preventing food poisoning

by these Enterobacteriaceae, as well as by all the

other pathogens implicated in intestinal disease. (See

Catering: Nutritional Implications.)

0037 Reporting all the suspected episodes to public

health authorities is essential both in investigating

an outbreak and in preventing additional cases.

See also: Catering: Nutritional Implications; Escherichia

coli: Occurrence and Epidemiology of Species other than

Escherichia coli; Food Poisoning: Tracing Origins and

Testing; Histamine; Shigella; Yersinia enterocolitica:

Properties and Occurrence

Further Reading

Balows A (ed.) (1991) Manual of Clinical Microbiology,

5th edn. Washington, DC: American Society for Micro-

biology.

Drasar BS and Barrow PA (1985) Intestinal Microbiology.

Washington, DC: American Society for Microbiology.

Gorbach SL (ed.) (1986) Infectious Diarrhea. Boston, MA:

Blackwell Scientific Publications.

Mandell GL, Douglas RG and Bennett JE (eds) (1990)

Principles and Practice of Infectious Diseases, 3rd edn.

New York: Churchill Livingstone.

Rienman H and Bryan FL (eds) Food-borne Infections and

Intoxications, 2nd edn. New York: Academic Press.

ESSENTIAL FATTY ACIDS

R Shireman, University of Florida, Gainesville,

FL, USA

Essentiality of Polyunsaturated Fatty

Acids

0001 In the late 1920s, the polyunsaturated fatty acid

(PUFA) linoleic acid (18:2n-6) was first described as

necessary in the diet of rats to promote normal health.

Soon, it was apparent that both 18:2n-6 (a-linoleic)

and 18:2n-3 (a-linolenic) are essential in the diet be-

cause animals require these for growth and function

of all tissues but lack the capacity for inserting double

bonds in the n-6 and n-3 positions. In the 1930s,

G. O. Burr and M. N. Burr described nutritional

essential fatty acid deficiency (EFAD) in a variety of

animals, and A. E. Hansen demonstrated its existence

in children. In ensuing years, work in the laboratories

of R. T. Holman and S. B. Johnson established normal

levels of total serum fatty acids (FA) in humans of

different ages, as well as the distributions in various

serum lipid classes (e.g., triacylglycerols, phospho-

lipids). These data allowed comparisons of serum

fatty acids of the n-9, n-6, and n-3 series in normal

and disease states. The single best index of essential

fatty acid status was reported first to be the total

serum n-6 concentration. Later, it was realized that

in deficient states, endogenous synthesis of the

20:3n-9 from 18:1n-9 occurs; thus, the serum level

of 20:3n-9 provides another index of status. Essential

fatty acids (EFAs) may be classified in two ways: (1)

those essential in the diet because animals cannot

synthesize n-6 and n-3 FA de novo and (2) PUFA

with essential functions. As pointed out by A. A.

Spector, linoleic acid appears to be essential not so

much because of an immediate cellular function, but

because it is the precursor of 20:4n-6, which has

numerous essential functions. Likewise, the import-

ance of dietary 18:3n-3 is that it is the precursor of

22:6n-3, which is found in membrane phosphatidy-

lethanolamine (PE) and phosphatidylserine (PS) in

large proportions in nervous tissue. Animal cells are

able to elongate and desaturate dietary 18:2n-6 and

18:2n-3 to form two long-chain fatty acids, arachi-

donic (AA, 20:4n-6) and docosahexaenoic (DHA,

22:6n-3). Since these are essential for cellular growth

and function, they are usually listed as EFA. How-

ever, 10:5n-3 (eicosapentaenoic, EPA) has many

effects in cells, but it has no proven essential func-

tions. DHA (22:6n-3) has been recognized as espe-

cially important for premature and newborn infants.

Some experts suggest using the terms ‘indispensable’

and ‘conditionally dispensable’; for example, if

20:4n-6 is present in the diet, there is an absence of

deficiency symptoms in healthy adults who have little

or no dietary 18:2n-6. Thus, the ‘true’ linoleate

requirement depends greatly upon the intake of

other FA.

ESSENTIAL FATTY ACIDS 2169

0002 Essentiality is based on biochemical evidence and

functional measures associated with deficiency. Nu-

merous experiments with nonhuman primates and

other animals have shown that functional develop-

ment of the retina and occipital cortex in the fetus and

neonate is abnormal in a-linolenic acid (18:3n-3)

deficiency because of a subsequent lack of 22:6n-3

production. Such studies are difficult to perform

because two or more generations of animals usually

have to continue on the deficient diet before definitive

effects are seen. Biochemical interactions and various

conversions of dietary PUFA make it difficult to state

an exact requirement for the EFAs. PUFA intakes

among adults are highly variable, but the average

estimated current intake is shown in Table 1. Major

sources of individual PUFAs are shown in Table 2.

Most diets contain much more n-6 than n-3, unless

supplemented with products such as fish oil or

flaxseed oil.

Essential Fatty Acid Deficiency

0003 Human EFAD of dietary origin is not common in

Western countries, and most EFAD research has

been conducted on rats. It may occur in patients on

long-term total parenteral nutrition (TPN) when

EFAs are not included in the TPN formulation. In

countries where protein malnutrition occurs, EFAD

may occur because of either very low dietary EFA or

lack of enzyme synthesis. Originally, it was thought

that 18:3n-3 could substitute for 18:2n-6, but this

proved not to be so because the products of the me-

tabolism of these PUFA differ in function. Dietary

18:3n-3 does not alleviate the dermatitis of EFAD

nor promote growth to the same extent as 18:2n-6.

The estimated minimal 18:3n-3 dietary requirement

for children is 0.54% of calories, whereas that for

18:2n-6 is about 1.1%. The estimated minimal re-

quirement of 18:3n-6 for adults is 1–2 energy percent,

with an optimum of 3–6% (approximately 1 table-

spoon of oil); the optimum for 18:2n-3 is about 2%.

The estimated ideal intake range of n-3/n-6 is 1:2 or

1:3. In both children and adults, the ratio of the two

should be considered, because an overabundance of

either suppresses some of the desaturation/elongation

reactions for the other.

0004Primary (dietary-related) and secondary etiologies

of EFAD in humans are listed in Table 3. There may

be other conditions that are secondary to gastrointest-

inal (GI) pathology or enzyme defects. In cases such

as chronic liver disease or hereditary enzyme deficien-

cies, the EFAD is not due to a lack of 18:2n-6 and

18:3n-3 in the diet but is due to an inability to

metabolize them further to the necessary end prod-

ucts (20:4n-6 and 22:6n-3). Some experts contend

that if 18:2n-6 is sufficient in the diet, typical symp-

toms of EFAD do not occur, but it is accepted by most

that n-3 FA are needed for normal neuronal develop-

ment and visual acuity, based on animal studies. The

importance of the long-chain 20:4n-6 and 22:6n-3 in

humans is illustrated by the effects of genetic enzyme

deficiencies such as Sjogren–Larsson syndrome and

acrodermatitis enteropathica; despite normal intakes

of 18:2n-6, there are severe symptoms of EFAD in-

cluding mental retardation, ichthyosis, and very ab-

normal prostaglandin metabolism (Table 4).

0005In the past, a serum triene/tetraene ratio of 1 was

considered a biochemical indication of serious EFAD.

An EFA-deficient diet results in insufficient 18:2n-6,

20:4n-6, and 22:6n-3 in tissues, but there is also

marked enrichment in eicosatrienoic acid (20:3n-9)

tbl0001 Table 1 Estimated average intake of PUFA in Western diets

Polyunsaturated fatty acid Intake (g perday)

18:2n-6 22.5

18:3n-3 1.2

20:4n-6 0.6

20:5n-3 0.05

22:6n-3 0.08

Total n-6/n-3 ratio 16.7

tbl0002 Table 2 PUFA major sources and functions

Fattyacid Majorsources Major functions

18:2n-6 a-Linoleic Safflower, sunflower, sesame, soybean,

and corn oils

Component of epidermal permeability barrier;

precursor of 20:4n-6

18:3n-3 a-Linolenic Linseed, canola, flax and other seed oils Precursor of EPA, DHA

20:4n-6 Arachidonic Meats, dairy products Precursor to bioactive eicosanoids; signal

transduction; gene expression

20:5n-3 Eicosapentaenoic (EPA) Marine fish oils Competitive inhibitor of 20:4n-6

22:6n-3 Docosahexaenoic (DHA) Marine fish oils, marine algae Neuronal membrane structure; gene expression;

prevention of apoptosis

18:3n-6 g-Linolenic (GLA) Evening primrose oil, borage seed oil Precursor to 20:3n-6

20:3n-6 Dihomo-g-linolenic (DHLA) Milk (small amount) Competitive inhibitor of 20:4n-6

2170 ESSENTIAL FATTY ACIDS

in blood and liver. Status is now usually measured

more accurately as the 20:3n-9/20:4n-6 ratio in

serum phospholipids. A normal value is considered

to be about 0.1–0.2. Occasionally, when abnormal-

ities are due to an enzyme deficiency rather than

to diet, some other index may be more useful. For

example, the normal conversion of 18:2n-6 to 20:4n-

6 results in a ratio of 18:2n-6/20:4n-6 of about 1.6,

but in severe dietary EFAD, it may be 1.2. In

achrodermatitis enteropathica, however, very little

arachidonic acid is synthesized, and this ratio may

be 10 or larger.

0006 Serum FA patterns and symptoms that resemble

EFAD occur in a variety of disorders. Marginal or

frank EFAD is somewhat common in young cystic

fibrosis (CF) patients, but whether this is a conse-

quence of decreased fat absorption or a defect in FA

metabolism, or both, is not known. While all serum

FA levels are decreased in some CF patients, the fact

that fewer desaturation products are present has led

to the speculation that there is an enzymatic defect in

FA metabolism. Oxidative stress disorders tend to

show FA patterns characteristic of EFAD. The reason

is unknown. Perhaps reactive oxygen species or prod-

ucts of oxidative damage act to decrease the activity

of desaturases. Whereas high amounts of PUFA in the

diet are said to increase the requirement for vitamin

E, in studies on concomitant EFAD and vitamin E

deficiency, measures of autooxidative susceptibilty

in rat red blood cells indicated that EFAD actually

potentiated the vitamin E deficiency. Patients with

malabsorption syndromes or long-term depressed

oral intake or TPN may exhaust stores of EFA and

exhibit clinical signs of EFAD, especially dermatitis.

Also, TPN formulations with very high ratios of n-6

to n-3 have resulted in symptoms of n-3 deficiency,

including neuropathy and immune impairment in

animals.

0007Nervous tissues in primates and other animals are

rich in n-3 PUFAs and their products. There have been

numerous reports of learning deficits and other effects

on nervous tissue in animals deprived of all n-3

PUFAs. If 22:6n-3 is not added to the media of retinal

photoreceptor cells in culture, they degenerate and

die, presumably because of activation of an apoptotic

pathway. But reports of effects in humans have been

less convincing, largely because long-term controlled

dietary experiments cannot be conducted. Episodic

numbness, weakness, pain in the legs, and blurring

of vision occurred in children on TPN formulas that

did not contain n-3 PUFA. These effects were reversed

after inclusion of n-3 FA in the formula. Researchers

have noted that some of the physical symptoms of

attention-deficit hyperactivity disorder are similar

to those of EFAD and that many of the diagnosed

children have decreased serum 20:4n-6 and 22:6n-3

levels. Conversion of 18:2n-6 and 18:3n-3 requires

tbl0003 Table 3 Etiology and characteristics of primary and secondary EFAD

Etiology Result Effect

Chronic malnutrition, especially in children Lack of dietary fat and protein Very abnormal serum FA patterns

Decreased enzyme synthesis Low total serum PUFA

Long-term fat-free TPN Lack of dietary EFA Abnormal serum 20:3n-9/20n-6

Various fat malabsorption conditions Secondary to pancreatic insufficiency,

bowel cancer, or other serious GI

diseases

Low total serum PUFA

Sjogren–Larsson syndrome Genetic enzyme defect in desaturation

and elongation

Low serum levels of C20 and 22 PUFA

Acrodermatitis enteropathica Genetic defect in e5 and e6 desaturase

activity

Very low levels of 22:6n-3

End-stage liver disease Impairment in desaturase and elongase

activities

Very low serum levels of LCPUFA

tbl0004 Table 4 Characteristics of EFAD in humans

Known clinical manifestations

Dermatitis; dry, scaly skin; impetigo; eczema; generalized

erythema

Coarse, sparse hair

Increased frequency of stools

Decreased growth rate

Cellular hyperproliferation in skin, alimentary tract, and

urinary tract

Possible immune impairment

Slow wound repair

Suggested manifestations associated primarily with 22:6n- 3

deficiency

Slower early learning behavior (based on animal studies)

Visual perturbations (based on animal studies)

Mental retardation in human genetic defects involving

desaturases and elongases

Known biochemical aberrations

Serum 20:3n-9/20:4n-6 greater than 0.4

Increased serum 16:1n-7, 18:1n-9, and 20:3n-9; decreased

18:2n-6, 20:3n-6, and 20:4n-6

ESSENTIAL FATTY ACIDS 2171

desaturase and elongase enzymes, which may not be

fully active in the preterm infant and neonate. Long-

chain PUFAs (LCPUFAs), especially 22:6n-3, are

particularly important for brain and nerve cell devel-

opment in utero, but the fetus does not synthesize the

long-chain EFA to any extent. Thus, maternal intake

may be more important than has generally been rec-

ognized. Many investigators support the inclusion of

n-3 PUFA, especially 22:6, in formulas for both pre-

mature and term infants. There remains some contro-

versy over whether the inclusion is needed, since

several studies have shown no detectable benefits for

breast-fed infants whose mothers took supplemental

dietary 22:6n-3. While there is some evidence of the

efficacy of n-3 long-chain PUFA in increasing early

visual maturation in preterm infants, there is no

demonstrated long-term benefit. Six randomized

trials with supplementation of infant formulas with

LCPUFA were reviewed by Simmer, who concluded

that there is little evidence to support the hypothesis

that supplementation with 20:5n-3 and 22:6n-3 con-

ferred any benefit on visual or cognitive development

or influenced the growth of term infants.

Metabolism of PUFAs

0008 In general, cells try to maintain a balance of FA of

different chain lengths and degrees of desaturation to

control membrane fluidity. PUFAs can be oxidized for

energy, but the functional and structural roles of EFA

depend largely on their metabolic products. All 20

carbon FA are referred to as eicosanoids, and

20:3n-6, 20:4n-6, and 20:5n-3 can all be used for

prostaglandin synthesis. The metabolism of the n-9,

n-6, and n-3 families can occur along the same path-

ways but normally does not, at least to the same

extent (Figure 1). Furthermore, major products of

the pathways may differ among cells, depending on

enzyme activities. For example, the end product of

20:4n-6 in endothelial cells is prostacyclin (PGI

whereas the end product in platelets is thromboxane

(TXA

). The balance of LCPUFAs in the cell is

regulated largely by induction or suppression of the

relevant desaturases and elongases. In EFAD, there is

increased desaturation of both 18:0 and 18:1. Negli-

gible amounts of oleic acid (18:1n-9) are normally

converted via the pathway in Figure 1, but in EFAD,

cells try to produce long-chain PUFA needed for

structure and functions by conversion of 18:1n-9

(oleic acid) to 20:3n-9. The ratio of 20:3n-9 to

20:4n-6 can be used as an index of deficiency.

0009In the presence of excess PUFA, the D9, D6, and D5

desaturases are suppressed, in part at the level of gene

expression. Experiments with cultured endothelial

cells have shown that added n-3 FA will be mostly

elongated to C22 products, thus effectively removing

competitors of 20:4n-6. But such experiments are

lacking in many other types of cells. A. A. Spector

has pointed out that even though cells can produce

22:5n-6, very little is normally produced possibly

because most of the precursor, 20:4n-6, leaves the

pathway for later prostaglandin synthesis. In n-3

PUFA deficiency, more 22:6n-6 is produced, but

it does not fully replace the function of 22:6n-3.

EFA desaturation and elongation occurs mainly in

the liver and is then delivered to other cells. DHA

(22:6n-3) is not synthesized in neurons, despite its

high concentration there; however, it can be synthe-

sized from 18:3n-3 and 20:5n-3 in the surrounding

astrocytes. It is hypothesized that astrocytes synthe-

size DHA and release it as a trophic factor for

neurons. EFA retroconversion, or shortening of the

chain, can occur but its metabolic significance, if any,

is not known.

∆6 Desaturase

∆5 Desaturase

∆4 Desaturase

Elongase

Oleic (18:1n-9)

18:2n -9

20:2n -9

20:3n -9

22:3n -9

Linoleic (18:2n-6)

18:3n -6

20:3n -6

20:4n -6

22:4n -6

22:5n -6

Linolenic (18:3n-3)

18:4n -3

20:4n -3

20:5n -3

22:5n -3

22:6n -3

fig0001 Figure 1 Metabolic pathways for fatty acid elongation and desaturation. Bold type designates major end products of the pathway.

2172 ESSENTIAL FATTY ACIDS

0010 Arachidonic acid undergoes metabolism to the

series-2 prostaglandins (i.e., two double bonds) and

to the leukotrienes of series-4 (four double bonds), as

simplified in Figure 2. The oxidation of eicosaenoic

acids (20:4n-6, 20:3n-6, and 20:5n-3) is catalyzed by

the enzyme prostaglandin endoperoxide synthetase

(PES), which has two separate activities usually re-

ferred to as cyclooxygenase (CO) and peroxidase.

Figure 2 shows the difference in the products in the

pathways that involve the same enzymes. Cyclooxy-

genase converts 20:3n-6 to PGG

, 20:4n-6 to PGG

and 20:5n-3 to PGG

. Subsequent conversion to

hydroxy analogs occurs next via peroxidase activity.

Metabolism of these endoperoxides to characteristic

products depends largely on the cell type. For

example, in arterial endothelial cells, PGG

is largely

converted to PGI

, prostacyclin, which causes vessel

vasodilation and also inhibits platelet aggregation. In

platelets, however, PGG

is converted to TXA

which

is important to normal blood clotting, since it causes

vasoconstriction and platelet aggregation. These

metabolites must be continuously synthesized as

they have a very short half-life. The various

products tend to act in a ‘balanced antagonistic’

way; while TXA

is the most potent proaggregatory

agent known, it is normally counteracted by prosta-

cyclin. When the balance is altered, excess thrombox-

ane leads to abnormal platelet adhesion and

aggregation and therefore it may be implicated in

coronary thrombosis. Although 20:3n-6 and 20:5n-

3 can be used to form similar products that have one

or three double bonds, respectively, the series-1 and

series-3 prostaglandins do not have the same bio-

logical activity as the series-2. In addition, the precur-

sors (18:3n-6 and 20:5n-3) compete with 20:4n-6 for

cyclooxygenase action. This is why EPA (20:5n-3) in

fish oil is said to be beneficial in preventing throm-

bosis.

0011 Lipoxygenase (LO) catalysis of 20:4n-6 forms

hydroxy fatty acids known as leukotrienes (e.g.,

LTA

,LTB

) These are compounds that have four

double bonds, at least three of which are conjugated

double bonds. Other eicosanoids can compete with

20:4n-6 for LO activity and thereby reduce the

amounts of series-4 leukotrienes formed.

Functions of EFAs and their Metabolites

0012There are numerous known and some purported

functions of PUFA. The major function of both

a-linoleic acid (18:2n-6) and a-linolenic acid (18:3n-

3) is as a precursor to other essential long-chain fatty

acids, although 18:2n-6 itself may be important in

membrane and lipoprotein structure and may have

some cellular functions. The functions of the metab-

olites are very different, except that both 20:4n-6 and

22:2n-3 are components of membrane phospholipids

and are involved in signal transduction. Also, both

are known to modulate or regulate gene expression

for several enzymes. All LCPUFAs suppress transcrip-

tion of genes encoding several lipogenic enzymes

including fatty acid synthetase and S14 protein and

induce certain genes encoding peroxisomal and cyto-

chrome P

450

enzymes. The net result is that at a high

PUFA, suppression of lipogenesis and triacylglycerol

synthesis occurs. They regulate the expression and/or

activity of the acyl desaturases and perhaps some of

the enzymes of the glycolytic pathway. Otherwise, the

effects of 20:4n-6 and 22:2n-3 are quite different. The

various products of 20:4n-6 metabolism also have

very diverse, sometimes antagonistic, functions. In

addition, the competition of the other 20Cn-6 and

n-3 FA for the same enzymes makes it difficult to

assign cause-and-effect functions. General functions

are shown in Table 5.

0013Among the lesser known effects of EFA are the

effects on lipoprotein metabolism. Normal EFA

status is necessary for overall lipoprotein metabolism

and especially for efficient high-density lipoprotein

processing by the liver. In dietary deficiency with

normal liver function, there is decreased incorpor-

ation of phosphatidylcholine into bile and decreased

chylomicron production. Altered partitioning of

available EFA has been noted in animals and in-

creased hepatic conversion of linoleic to arachidonic

acid occurs. Also, hepatic D6andD5 desaturases are

upregulated. More eicosanoid precursors are distrib-

uted into hepatic phosphatidylethanolamine than

tbl0005Table 5 Some general functions of arachidonic acid,

docosahexaenoic acid, or their metabolites

Membrane structure Blood clotting

Signal transduction Immune responses

Vasoconstriction Vasodilation

Cell proliferation Apoptosis

Modulation of gene expression Lipoprotein synthesis and

metabolism

20:4n-6

20:5n-3

22:6n-3

LO PES PES

PGE

PGI

TXA

PGE

TXA

Leukotrienes

elongase

desaturase

fig0002 Figure 2 Summary of arachidonic and eicosapentaenoic acid

metabolism.

ESSENTIAL FATTY ACIDS 2173

usual. The n-3 PUFA decrease serum triacylglycerol

through reduced very low density lipoprotein produc-

tion. The precise biochemical nature of these effects

has not been established. Another less known effect of

EFAs is that they enhance the effects of vitamin D in

increasing calcium absorption and help decrease urin-

ary excretion of calcium. EFA-deficient animals de-

velop severe osteoporosis and ectopic calcification.

0014 The effects of PUFAs on cell proliferation, tumors,

and cancers are highly variable, depending on the cell

type and proportions of FA. They may be modulators

of carcinogenesis with distinct tissue-specific pro- or

anticancer effects. LCPUFAs activate the peroxisomal

proliferation activator receptor, which may mediate

their purported effects on gene expression, cellular

growth and differentiation, and apoptosis. Both

18:2n-6 and 20:4n-6 are said to inhibit mitogen-

stimulated proliferation of lymphocytes in culture,

but they may have no effect or a stimulatory effect

in other cell types. In EFAD in animals, diminished

immunity occurs because 20:4n-6 metabolites are

needed for normal immune responses. However, the

n-3 PUFAs suppress immune response and may be of

benefit in inflammatory and autoimmune disorders.

As mentioned above, the prostaglandin and throm-

boxane derivatives of 20:4n-6 are bioactive molecules

that act as paracrine hormones and have many well-

known, important functions on blood clotting, smooth

muscle contractions, and immune responses. When

released from phospholipids, 20:4n-6 activates pro-

tein kinase C and therefore modulates the activation

of growth factor receptors such as EGH receptors. It

is thought to have other cell-signaling functions as it

can inhibit GTPase activated protein.

0015 The n-3 PUFAs are said to inhibit endothelial acti-

vation. This may occur in at least two ways; 20:5n-3

competes with 20:4n-6 for conversion to the prosta-

glandin, which normally stimulates this activity, and

22:6n-3 decreases expression of endothelial leukocyte

adhesion molecules. Usually, prostacyclin-mediated

cytokine stimulation of expression of adhesion mol-

ecules is a normal physiologic protective function, but

in pathological states, this leads to an undesirable

sequence of events that may result in atheroma for-

mation. These n-3 PUFAs may help confer a protect-

ive effect against atherogenesis. Also, n-3 FA promote

an antiarrhythmic action, apparently owing to a

slight hyperpolarization of heart muscle membranes

that prevents ‘easy’ induction of action potentials.

0016 Leukotriene metabolites have profound effects on

white blood cell chemotaxis and phagocytosis,

regulation of neutrophil and eosinophil function,

smooth muscle contraction and bronchoconstriction

in the pulmonary system, and changes in capillary

permeability. In neutrophils, 20:4n-6 is metabolized

primarily by the 5-LO pathway leading to the for-

mation of LTA

, which is a precursor for a potent

chemotactic factor. Thus, a decrease in 20:4n-6

concentration can result in an antiinflammatory

response. Since 20:5n-3 and 20:3n-6 compete with

20:4n-6 for the LO pathway, the result is similar

because less conversion to LTA

occurs. Although

the prostanoids and leukotrienes formed from

20:4n-6 have many extremely important physiologic

functions, overproduction of some of these com-

pounds appears to be involved in certain disease pro-

cesses or states, including thrombosis, myocardial

infarction, cardiac arrythmia, arthritis, asthma, in-

flammatory conditions, and autoimmune disorders.

Reduction of tissue levels of 20:4n-6, competition for

enzymes by other FA, or inhibition of the enzymes by

drugs may in some cases ameliorate some of the

symptoms associated with the disorder(s). For

example, aspirin, ibuprofen, and several other drugs

inhibit the activity of cyclooxygenase, and steroidal

antiinflammatory compounds such as hydrocortisone

inhibit the enzyme phospholipase A

that releases

20:4n-6 from membrane phospholipids. All of these

result in less prostaglandin synthesis. Inhibition of

phospholipase A

also reduces leukotriene synthesis.

0017DHA (22:6n-3) is the major PUFA in neuronal

membranes; it is especially enriched in PS. PS is im-

portant structurally and functions in translocation of

specific proteins, such as Raf-1 and protein kinase C,

to the plasma membrane. Also, it is hypothesized to

be involved in cellular apoptosis. Raf-1 translocation

is a first step in the transduction of growth factor

signaling. Depletion of 22:6n-3 leads to a decrease

in the synthesis of PS and lower levels of PS in brain

cell membranes. PS levels are much higher in nerve

tissues than in other tissues, and n-3 deficiency results

in a dramatic reduction of PS only in nerve cells.

Thus, n-3 deficiency seems to have profound effects

on PS-related signaling events in the nervous system.

In animals, a decrease in 22:6n-3 may be associated

with compensatory increased levels of 22:5n-6 leading

to: modification of neural membrane composition

and functional changes in enzyme activities, sub-

optimal retinal and brain development, and a poorer

performance in learning tasks. It may also be cor-

related with peroxisomal disorders associated with

neuronal deterioration. Brain concentrations of

22:6n-3 plateau with 18:3n-3 intake of about 0.7%

energy, but this is influenced by the 18:2n-6 intake.

Several studies have shown adverse effects on growth

and function of animals with high intakes of 22:6n-3.

0018The amount, type, and balance of dietary FA along

with antioxidant nutrients impact the immune system.

The amelioration of symptoms of autoimmune dis-

eases in some studies with high doses of fish oils

2174 ESSENTIAL FATTY ACIDS

was attributed to effects of 20:5n-3 suppression of T-

lymphocyte proliferation, apoptosis of autoreactive

lymphocytes, and decreased proinflammatory cyto-

kine production. Long-term effects of such treatment

on host immunity require further study. T-cell prolif-

eration was shown to be decreased after administra-

tion of 2.4 g per day 18:3n-6 (g-linolenic) in borage

seed oil, but not with 18:3n-3 or 18:2n-6. The effect

therefore appears to be a competitive effect of 20:5n-

3 and 20:3n-6 with 20:4n-6. Dietary 20:5n-3 and

18:3n-6 also appear to inhibit arachidonic induced

production of thromboxane (TBB

) by platelets.

Recent Advances

0019 In the past decade, much research has focused on fish

oil supplementation, with high levels of 20:5n-3 and

22:6n-3. The fatty acid content of menhaden oil is

shown in Table 6, and possible benefits and concerns

of supplementation in Table 7. Three areas have been

of special interest: effects on immune responses,

coronary heart disease, and behavior. In all of these

areas, there have been inconsistencies in the reported

results, some of which may be due to different diets

and conditions.

0020 Some researchers have suggested a role for EPA as

well as DHA in normal nerve cell membrane struc-

ture. This is based largely on reports of lower than

normal levels of EFA in some patients with schizo-

phrenia and the moderately positive effect observed

with EPA supplementation. However, since EPA can

undergo further metabolism to DHA, it is unclear

whether EPA has anything other than a precursor

role in these cells. There are trials in progress on the

use of EPA in the treatment of schizophrenia.

0021Rats with decreased brain levels of 22:6n-3 show

an impaired performance in learning tasks compared

to rats fed 22:6n-3 and increasing the amount of

22:6n-3 to a 10:1 ratio n-6/n-3 in infant formula

of primates was associated with greater accretion of

22:6n-3 in neonatal brain and retina. Patients with

moderately severe dementia of thrombotic cardio-

vascular disorder and Alzheimer’s dementia supple-

mented with fish oil were followed for one year and

showed improved scores on a psychometric test as

well as better red blood cell (RBC)-deformation

scores. It was unclear whether the improvement was

a direct effect on neuronal cells or due to improved

RBC function. Low-serum 22:6n-3, along with the

associated decrease in phosphatidylcholine, is now

considered to be a significant risk factor for Alzhei-

mer’s dementia. Of concern is the reduction in D6

desaturase activity that occurs with aging and pos-

sible lack of adequate 22:6n-3 synthesis, even with

adequate intake of 18:3n-3. This FA comprises more

tbl0006Table 6 Composition of menhaden oil

Polyunsaturated fatty acid Grams per 100 g of oil

18:2n-6 2.0

18:3n-3 1.4

20:4n-6 1.1

20:5n-3 13.0

22:5n-3 5.0

22:6n-3 8.5

tbl0007 Table 7 Possible beneficial effects vs. safety concerns related to dietary n-3 PUFA

Possible benefits of supplementation in selected individuals

Decrease in serum triacylglycerols

Decrease in myocardial infarctions in persons with diagnosed CHD

Decreased inflammation and relief of certain conditions, as rheumatoid arthritis, psoriasis, lupus and inflammatory bowel syndrome

Decreased cardiac arrhythmias (animal studies, not very definitive in human studies)

Possible safety concerns related to excess intake (based on animal or human studies)

Increased bleeding times and associated risk of hemorrhagic stroke

Increased peroxidation of fatty acids

Inhibition of immune cell function and decreased resistance to infection

No change or increase in serum LDL-cholesterol

Imbalance of PUFAs (very low n-6/n-3) with resultant behavioral abnormalities

Concerns related to interactions with drugs

Interactions with drugs that affect eicosanoid metabolism and clotting mechanisms

Interactions with drugs that affect eicosanoid metabolism and immune responses

Potentiation of cytotoxicity of some anticancer drugs

Possible cases for ‘special’ supplementation

Chronic liver disease – may require supplementation of 20:4n-6 and 22:6n-3

Cystic fibrosis – may require supplementation of 20:4n-6 and 22:6n-3 in addition to 18:2

Inherited desaturase and/or elongase deficiencies, as in some peroxisomal disorders

Subnormal 20:6n-3 serum levels associated with certain types of depression, dementia, attention-deficit disorder,

adrenoleukodystrophy, long-chain hydroxyacyl-CoA dehydrogenase deficiency, and dyslexia

ESSENTIAL FATTY ACIDS 2175

than 30% of the structural lipid of neurons and is very

enriched in retinal rod outer segments. Subnormal

levels are associated with neurological conditions,

including depression, attention deficit hyperactivity

disorder, and dementia. Dietary intervention to

increase plasma 22:6n-3 levels has been associated

with improvements in visual or neurological deficits

in adrenoleukodystrophy, long-chain hydroxyacyl-

CoA dehydrogenase deficiency, dyslexia, and Alzhei-

mer’s dementia. Intervention trials are now underway

to determine the effects of dietary 22:6n-3 from algal

sources on development of Alzheimer’s dementia.

This source, which contains much less 20:5n-3, will

be evaluated for safety and efficacy in hopes of redu-

cing the side-effects of high levels of 20:5n-3, includ-

ing immune suppression and bleeding tendencies.

0022 Diets rich in n-3 PUFAs have been associated with

suppression of cell-mediated immune response.

Although this accounts for the reported beneficial

effects of fish oil in arthritis, suppression of immune

response is of great concern in some individuals. Fish-

oil diets suppress lymphocyte proliferation, monocyte

and neutrophil chemotaxis, and tumor necrosis

factor, interleukin-2, and interleukin-6 production,

and also reduce expression of adhesion molecules

on the surface of endothelial cells, monocytes, and

lymphocytes. This is the basis of its antiinflammatory

effect. LTB

, which is a potent chemoattractant for

monocytes, important in regulation of inflammatory

response and cytokine synthesis, declines in the pres-

ence of excess 20:5n-3. Also, PGE

, which normally

increases during infections and acute inflammation,

declines, and the series-3 prostaglandins and series-5

leukotrienes, which have a less potent effect, increase.

These effects are attributed to 20:5n-3, rather than

22:6n-3.

0023 Because of anecdotal and epidemiological evidence

suggesting that FO had a positive influence on

decreasing the incidence of CHD, in 1993, the US

Food and Drug Administration (FDA) conducted a

study to determine whether to allow a health claim

for n-3 fatty acids. At the time, the FDA concluded

that findings were inconsistent in the general popula-

tion, and there was insufficient evidence to support a

health claim for n-3 FA and CHD. Some suggested an

adverse effect. A part of the problem with the early

epidemiological and intervention studies is that

plasma total cholesterol or low-density lipoprotein

cholesterol (LDL-C) was used as the biomarker for

reduced risk of CHD. Almost all studies with fish oil

showed no significant decline in LDL-C, and some

showed an increase. More recently, the FDA again

considered whether to allow the health claim, and in

this review, different endpoints and clinical measures

of the risk of CHD were evaluated. These included

the endpoints of fatal or nonfatal myocardial infarc-

tion and clinical measures of reduction in fasting and

postprandial serum triacylglycerol levels, reduction in

platelet aggregation and adhesion, and changes in the

composition of serum lipoproteins. In the mid- to late

1990s, all intervention trials of persons with diag-

nosed CHD supplemented with fish oil or n-3 deriva-

tives showed no decrease in LDL-C but decreases in

the risk of CHD using the mentioned endpoints. After

reviewing the relevant data on beneficial effects on

the risk of CHD, adverse effects of any type and the

intakes associated with these effects, the FDA found

that the use of 20:5n-3 and 22:6n-3 as dietary supple-

ments is safe, provided the intake does not exceed

3g per day from all food and supplement sources.

Seealso: Fatty Acids:Properties;Metabolism;Gamma-

linolenicAcid;Analysis;DietaryImportance;Trans-fatty

Acids:HealthEffects; Fish Oils:Compositionand

Properties;DietaryImportance