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The Effect of Long Chain N-3 PUFA on Obese Individuals - Desk Review
Shilpa Goel1, Monica Chaudhary2, Renu Khedkar1 and Shweta Khandelwal2*
1Department of food and nutrition, Amity Institute of Food Technology, India
2Public health foundation of India (PHFI), India
Submission: June 8, 2018;Published: August 31, 2018
*Corresponding author: Shweta Khandelwal, Senior Research Scientist & Associate Professor, PHFI, 47, Sector 44, Gurugram, Haryana-122002, India
How to cite this article: Shilpa G, Monica C, Renu K, Shweta K. The Effect of Long Chain N-3 PUFA on Obese Individuals - Desk Review. Nutri Food Sci Int J.
002 2018; 7(3): 555714. DOI:10.19080/NFSIJ.2018.07.555714.
Context: Current recommendations for counteracting the action of obesity advocate the consumption of a healthy diet and participation in regular physical activity. Studies have indicated towards a potential role of long-chain omega-3 polyunsaturated fatty acids (LC N-3 PUFA) in a number of effects which ameliorate the condition of obesity.
Objective:To assess the effect of LC N-3 PUFA (Docosahexaenoic acid and Ecosapentaenoic acid) on obese individuals.
Data sources: Using Cochrane guidelines, electronic databases- Google Scholar and PubMed-NCBI were searched for evidence from the last 10 years (March 2007-17).
Study selection:Randomized Controlled Trials (RCT) based on LC N-3 PUFA supplementation involving obese adult were included.
Results: The association between change in body weight/ body mass index (BMI)/ waist circumference and fish oil supplementation in 31 RCTs involving overweight or obese adults was investigated. Of the 31 studies, 9 did not provide any evidence for the biomarkers of LC N-3 PUFA intake like EPA/DHA fatty acids plasma concentration and N-3 FA percentage in serum phospholipids. The preliminary evidence suggested that LC N-3 PUFA supplementation combined with energy-restricted diets or exercises prevents weight regain. Treatment groups showed a higher drop in BMI and body fat percentage (0.24kg/m2; 0.49%) than controls.
Conclusion: While there is growing evidence that LC N-3 PUFA can improve body composition, contradictory findings have also been reported. There is an urgent need for long-term studies and meta-analysis in this area of research to generate conclusive evidence on the effects of LC n-3 PUFA supplementation on obesity..
Keywords: Obesity; LC N-3 PUFA; Fish oil; Weight loss; Body weight; Body mass index; Anti-inflammatory biomarkers; CVD risk
Abbreviations: EPA: Eicosapentaenoic Acid; DHA: Docosahexaenoic Acid, LC N:3 PUFA: Long Chain Omega 3 Polyunsaturated Fatty Acids: BW: Body Weight; BCA: Body Composition Analysis; HR: Heart Rate; WC: Waist Circumference; CVD: Cardio Vascular Disease; HRV: Heart Rate Variability; HOMA-IR: Homeostatic Model Assessment-Insulin Resistance; SBP: Systolic Blood Pressure; DPB: Diastolic Blood Pressure; PCOS: Polycystic Ovarian Syndrome; RCT: Randomised Control Trial; T2DM: Type 2 Diabetes; HC: Hypo Caloric Diet; BMI: Body Mass Index; CRP: C: Reactive Protein; IL:6: Interleukin 6; WHR: Waist to Hip Ratio; HDL: High Density Lipoproteins; LDL: Low Density Lipoproteins; VLDL: Very Low Density Lipoproteins; TC: Total Cholesterol; TAG: Triacyl Glyceride
Globally, 2.1 billion people are overweight . Worldwide obesity prevalence rate has more than doubled since 1980 . The 2013 Global Burden of Disease study, published in May 2014, showed that 37% of men and 38 % of women had Body Mass Index ≥ 25 kg/m2. It stands for a rise of 28% adult and 47% children since 1980. Considering the demographic factors, it has been observed that the prevalence of overweight individuals is higher in urban settings as compared to rural settings .
A cross: sectional study was conducted by Indian Council of Medical Research (ICMR): ICMR: INDIAB (Indian Diabetic
Study), which suggested that India, the second most popular country in the world, is currently experiencing a rapid epidemiological transition from underweight to obesity . The National Family Health Survey:4 (NFHS:4) also reported that in India, obesity (BMI ≥ 25kg/m2) was more prevalent in the urban areas (Madhya Pradesh, West Bengal, Chhattisgarh, Orissa) and in higher socio: economic groups as compared to the rural areas, especially among women (Men: urban: 26.6 vs. rural: 14.3%; Women: urban: 31.3 vs. rural: 15.0%) [4-6]. Worldwide, Indians were found to have the highest predisposition to abdominal obesity and accumulation of visceral fat. In context of north India (New Delhi), the overall prevalence was categorised in terms of generalized obesity (50.1%), to that of abdominal obesity (68.9 %) [5,7].
With the increase in urbanization, poor dietary behaviours [8-9] compounded by decreased levels of physical activity [10-11] has created an obesity: prone environment. This adds on to the prevalence rate of obesity, especially in the developed countries. Genetic susceptibility combined with chronic positive energy balance due to unhealthy lifestyle behaviours and environmental factors are considered as the key features in the development of adiposity [8,11].
Current recommendations from most public health bodies
for reducing body weight are based on intensifying physical
activity and eating a healthy balanced diet. However, many
people are facing difficulty complying with these lifestyle
changes, particularly over the longer duration. Hence, despite
widespread recommendations to improve diet and physical
activity habits the prevalence of obesity continues to rise.
Thus, there is an interest in exploring dietary options which
could help in keeping body weight in check. One such option
may be use of Long Chain Omega:3 fatty acids (DHA & EPA) in
or along with our diets . As per WHO, globally, adequate
daily intake suggested for omega 3 fatty acid is 1:2 % of total
energy i.e. 2:4gm/day for adults [14-15].
Pharmacokinetics of omega 3: A study conducted by Rusca
et al.  in the year 2009 showed a 2.0fold and 9.3fold rise
in plasma DHA and EPA concentrations on consumption of
3 fish oil (FO) capsules (1g each) daily for 28 days. The half:
life was found to be 4.8 days and 10.3 days for EPA and DHA
respectively by Martijn et al.  study. This study was done
on 58 overweight men, which were randomised in 4 groups
getting an intervention of placebo (9g/d) vs. FO (3g/d) +
Placebo (6g/d) vs. FO (6g/d) + Placebo (3g/d) vs. only fish oil
(9g/d) for 1 year with 6-month washout period.
Obesity or adiposity is all linked with the alteration in the adipocytokine levels. This is regulated by the action of adiponectin hormone which helps in the metabolism of fat [17-18]. Moreover, dysregulated production of adipocytokine plays a role in the pathogenesis of obesity: associated metabolic
syndrome like Cardiovascular Disease (CVD), type 2 diabetes, cancer, inflammation etc [1,8,19]. It has been concluded in several studies that lower levels of adiponectin up: regulate the activity of pro: inflammatory cytokines and increase insulin resistance, oxidative stress, leptin levels and serum C: reactive protein [17,19]. Peroxisome proliferators: activated receptor: γ (PPAR: γ) is a nuclear transcription factor which is highly expressed in the activity of white adipose tissue . It binds to response elements in target gene promoters and regulates the expression of numerous adipose: specific genes [17,19]. A direct relationship between PPAR: γ activity and adipokine differentiation, and its inverse relation to BMI has been found in several observational studies [8,19-20]. Overall, for managing obesity and its related consequences; it is of prime importance to correct the altered activity of adiponectin and other hormones related to adiposity in the body.
In context of Long Chain Omega:3 Polyunsaturated Fatty
Acids (LC N:3 PUFA) and its association with obesity, higher
levels of LC N:3 PUFA in a normal weight individual than an
obese individual have been reported by Jump et al. .
This Weight loss mechanism includes, down: regulating
the transcription of lipogenic genes and up: regulating the
genes that promote lipid oxidation, thermo: genesis, plasma
adiponectin, leptin and gherlin levels and anti: inflammatory
biomarkers (TNF: a, CRP, IL:6, PPAR: γ activity, etc.) [21-23]. As
a result of this, there is a shift in metabolic profile toward one
that favours increased fat oxidation and reduced fat deposition
which ultimately leads to weight management.
This desk review was conducted by 2 research assistants.
To identify relevant studies two search engines Pub Med and
Google scholar were used from January through March 2017.
Table 1 lists the inclusion and exclusion criteria used in the
study. The systematic search identified 487 titles and abstracts
of which the full texts of 104 potentially eligible articles were
critically appraised. Out of all, 31 studies met the inclusion
criteria and were included in the review.
As per the Cochrane collaboration, a search strategy was
followed from title-based screening to abstract screening
and finally screening full text of the articles. The screening
was based on the inclusion and exclusion criteria, which
were formed at the beginning of the study. For this review,
the relevant records were obtained by using the terms “fish:
oil”, “omega 3”, “DHA or docosahexaenoic acid”, “PUFA or
polyunsaturated fatty acid” and “N:3 polyunsaturated fatty
acids” (Figure 1).
After exclusion according to our criterion, 104 studies were retrieved for full: text screening. Of these, 31 studies met the inclusion criteria among which majority of them were 28 parallel studies with double blind placebo trial and rest 3 studies were crossover studies. In more than half of the studies, the results were interpreted in the form of body weight (BW), waist circumference (WC) and BMI. The association of weight loss in the context of change in Leptin, Gherlin and Adiponectin levels is also illustrated in 9 studies. Beside this, change in inflammatory biomarkers; insulin resistance and lipid profile were also measured that are linked to obesity.
There were 13 studies whose prime focus was towards the weight lowering action of LC N:3 PUFA supplementation, out of which 5 studies reported a significant effect of DHA: EPA in the treatment of obesity as a reduction in body weight. The first Randomized control trial in healthy overweight patients was published in 2007 by Parra et al. [21,24], this was a result from same SEAFOO plus young study. The effect of LC N:3 PUFA on body weight was assessed by making a comparison between 4 experimental groups: no seafood vs. cod fish vs. salmon fish vs. fish oil capsule which were randomised among 324 participants. The trial was conducted for 8 weeks along with the addition of lean meat, 30% energy restriction in diet and increase in physical activity level. The result demonstrated that caloric restriction in the diets induced an average weight loss of 5.8 ± 3.2%, which was statistically significant in all treatment groups and slightly higher in the dietary groups including seafood or fish: oil capsules [21,24,25].
Another RCT (study by Munro et al. [26-27]) on overweight individuals investigated the effect of fish oil (2.1g/day) in comparison to that of Sunola oil (placebo) along with energy restriction diet. At 4 weeks (the intervention phase), the mean weight loss was :6.54kg (26.9%) for Placebo group and :6.87kg
(27.7%) for fish oil group. Whereas at week 14 (after 10 weeks
of maintenance phase in which participants were given normal
diet), there was a further mean decrease in weight, :1.57kg
(1.85%) for PB and :1.69kg (:1.9%) for FO. On the top of it, a
reduction in fat mass by 8.95% (-3.43kg) in placebo group to
that of 9.76% (:3.8kg) in fish oil group followed by increase in
fat free mass of 0.67% (0.29kg) for placebo to that of 0.51%
(-0.36 kg) in fish oil was also found.
Hardena et al.  through their study explained that
body mass composition was not affected by treatment,
although a fall in body weight in the DHA group approached
statistical significance (P = .029). Considering the individual
effect of DHA in fat reduction, Neff et al.  (n=49) showed
a significant reduction of abdominal (p=0.007) and gluteal
fat (p=0.001) on supplementation of algal DHA (2g/day) for
4 months as compared to soyabean oil as placebo. Cussons
et al.  and Crochemore et al.  investigated the effect
produced by administration of 4g/day of fish oil and 1.5: 2.5g/
day of fish oil respectively. The former study was conducted on
polycystic ovarian syndrome women whereas the latter was
on postmenopausal Type 2 Diabetic patients. The significant
decrease in levels of waist circumference, body weight, BMI
and fat mass were observed in the fish oil group of PCOS
women. The greater percentage of weight loss was found in B
group (1.5g/d) as compared to the other groups in later study.
Unlike the studies mentioned above, 4 studies showed
contradictory findings. Wong et al.  conducted a study
on 27 healthy overweight individuals for 16 weeks (12 weeks
intervention phase + 4 weeks of maintenance phase). The
randomization was done between 2 groups (control and fish
oil (4g/ day) group), and no considerable difference in weight
loss between the 2 group was found. On the other hand,
Gammelmarka et al.  in their study also showed the same
outcome where they supplemented their one group by fish oil
(1.1g/day) and other by olive oil. In the same manner Defina
et al.  and Tapsell et al.  in their studies found no significant difference in reduction of body weight among the groups.
It was illustrated in the study by Wong et al.  (n=27)
that there is a significant increase in the adiponectin levels
in group supplemented with omega 3 source as compared to
control group. In context of dosage to time effect, Mohammadi
et al. study  in which supplementation of fish oil was given
to PCOS women (n=64), at the rate of 4g/d for 8 weeks, there
was an increase in adiponectin level by 19.5% in FO group.
On the other hand, an increment of 4.5% (+0.37μg/ L) was
observed in the crossover: canola oil multicentre trial (COMIT)
by gravel et al. , in canola +¬ DHA group compared to other
4 interventional groups. A similar trend was marked in the
study by Gammelmark et al. , where the level raised by
0.55μg/ ml in the FO group after 6 weeks.
Gherlin and leptin were found to be having a direct and
inverse relationship with the erythrocyte level of DHA + EPA
respectively . A comparison was made between the3
experimental groups (Hypo caloric diet+ placebo (control)
vs. HC + fish vs. fish + Omega:3 capsules) by Tapsell et al. 
in their study , where group B and C showed a significant
increase in the erythrocyte DHA levels followed by reduction
in leptin levels (p>0.001).
For comparing the effect of type of fish on leptin and gherlin
levels, SEAFOO plus YOUNG study  illustrated that there is
a potential increase in gherlin and decrease in leptin levels in
salmon group followed by supplemented group. Leptin in long
term-controlled trial, is significantly related with fullness and
reduction in desire to eat .
There are total of 13 studies which focus on the potential
effect of DHA: EPA in reducing the risk factors associated with
cardiovascular disease. Out of them, 8 are based on change
in total cholesterol, Low density lipoprotein, Triglycerides,
High density lipoprotein levels; 2 for change in blood pressure
levels; and 3 includes the result based on heart rate variability.
Ramel et al. , showed a decrease in Systolic blood pressure
(:4.4 ± 8.6 mmHg, P < 0.001) and Diastolic Blood Pressure (:4.1
± 7.4mmHg, P < 0.001) with salmon (P=0.032) and fish oil (P
= 0.044) groups more significant than other group, as they
both are rich in omega 3 content. The similar results were
demonstrated by Cusson et al. .
Cardiovascular risk is directly proportional to the levels of serum phospholipids (TG, LDL, and VLDL). Based on the study by Gunnarsdottir et al.  (SEAFOO plus YOUNG study) and Munro et al. [26,27], a significant reduction in the total cholesterol (TC), total triglycerides (TG), low density lipoproteins (LDL) was stated in group fed with higher concentration of omega 3 (DHA + EPA). Also, it has been proven through the theory of gene regulation that on supplementation of fish oil there is rise in level of HDL in both the studies [26,41]. A significant reduction of serum levels of total cholesterol (A:42.8%, B: 54.1%, C: 46.1%), LDL: cholesterol (A: 50%, B: 50%, C: 38.5%), and TG (A: 57.1%, B: 64.3%, and C: 53.8%) among the groups was showed by Crochemore et al. . An increase of serum HDL concentration by 42.9%, 50%, and 38.9% in Group A, B, and C, respectively was observed.
A positive association between N:3 PUFA intake and Heart
Rate Variability has been reported in adults for managing the
high risk for CVD . There was a significant oil treatment
X time interaction (P>0·008), as a result of this there was a
reduction in resting and exercise HR in the subjects. In addition
to this FO intervention had improved HRV by increasing high:
frequency power, representing parasympathetic activity,
compared with placebo (P=0.01).
A significant treatment effect was observed for plasma CRP (C: reactive protein) concentrations (one of the anti: inflammatory biomarker) (P = 0.04), in canola DHA oil supplemented group only . Ramel et al.  and Parra et al.  in SEAFOO plus YOUNG study showed the effect of omega
3 in reduction of hs:CRP & Interleukin:6 concentration (high:sensitivity CRP= :32.0%; IL:6= :18.4%) and oxidative stress respectively. The rise in levels of anti: inflammatory cytokines was also observed in the study by Kabir et al.  & root et al. , where they supplemented Fish oil (FO) approx. at 1.g/d
for 8 and 4 weeks respectively. On the contrary, no overall effect was seen in the study by Gammelmarka et al.  where same amount of FO was intervened for 6 weeks.
Ramel et al.  published in his study, that fasting insulin
and Homeostatic model assessment - insulin resistance
(HOMA:IR) were significantly lower in the fish oil group than in
the control group at endpoint (16.4% and 17.2% respectively)
through his randomized double-blind control trial. Tapsell et
al.  & Cusson et al.  proposed the same result. Omega:3
fatty acids significantly decreased glucose (by 11.4%, p<
0.001), insulin levels (by 8.4%, p<0.05), and HOMA:IR (by
21.8%, p<0.001) compared with placebo in Rafraf et al. 
study (Table 2).
Through various evidences it was found that intake of fish
or fish oil capsules can decrease the weight of adult individuals
with BMI ≥ 25 kg/m2. Among 31 studies there were 9 studies
which did not provide any evidence for the biomarkers of
intake [47-48] (DHA + EPA) that includes plasma concentration
of EPA/DHA fatty acids, increases in the n3/n6 ratio of plasma
FA , percentage of N 3 FA in serum phospholipids and the
ratio of n6: n3 platelet membrane phospholipids etc. These
biomarkers play a key role in achieving the proposed outcomes
of a study, an alteration in any of them act as confounding factor
or/and may create a bias in the study [27,29,30, 36,42,47,50].
Review consist of studies in which participants have went
through some or the other kind of dietary and physical activity
modification which are judged by FFQ, 3: day diary record,
24hr dietary recall etc under free living conditions that cause
attrition in the negative outcomes. As such cases, participants
tend to record some false details that may induce a bias [21,35-
There is conflicting evidence on the effect of LC N:3 PUFA
on weight loss found in the review articles. Du et al.  did
a review on the anti: obesity effect of fish oil, in which 21
studies were included. Through meta: analysis, they concluded
no association between fish oil and weight loss. On the other
hand, Buckley et al. [52-53] who published a review article in
2009:2010 [52-53] with the same objective, but was not based
on meta: analysis. In this review, no evidences were reported
which would conclude the beneficial effect of fish oil in obese
Unlike the reviews mentioned above, a meta: analysis was
done by Bender et al.  in 2014 on 12 RCT’s. A significant
difference was estimated among the intervention and control
group in terms of weight loss (P=0.047), waist circumference
(P=0.003) and body fat. Rather they stated a beneficial role of
long term intervention of fish oil on weight loss (P >0.017) .
In context of the adiponectin level which is. referred
as obesity bio marker, the current review found a positive
correlation between LC N:3 PUFA and serum adiponectin
concentration. On the contrary however, a review conducted by
Gray et al.  on 11 clinical trials and observational studies
showed otherwise. Through their review they concluded with
the potential role of omega 3 in weight maintenance when
given with a calorie restricted diet but no overall effect on
adiponectin levels. In contradiction to the review by Gray et
al.  & Wu et al.  published a meta: analysis on 14 RCT’s
in 2013 for analysing the effect of DHA + EPA on adiponectin
levels. According to this, a potential increase in the circulating
adiponectin levels were found in the group supplemented with
omega 3 (P= 0.02) .
In the present review, there were several studies which
were in agreement of the hypothesis while other was not. All the studies were based on intention to treat analysis, which has its own drawback, because in this case withdrawals and
deviations are usually ignored, leading to biasing in the results.
The study by Ramel et al.  which was a SEAFOO plus YOUNG
study included in this review, demonstrated all the objective of
the review in the form of positive results [21,25,27,41,43,50].
On the other hand, Gammelmark et al.  failed to support
the role of omega 3 in weight loss and another objective. As
the studies included in the reviews were conducted for shorter
duration with low dosage. A huge variation in the supplemented
value was observed in the review studies ranging from 1:6g of
omega 3/day. There are 6 studies in which WHO standards of
recommendation  have not been followed like in the study
by Crochemore et al. , Kabir et al. , Root et al. ,
Hardena et al. , Gammelmark et al. , Neff et al. .
In the present study, some indications were found that the
effect might be greater in men than in women for the outcome
waist circumference, which is a measure for visceral adiposity.
This is relevant, as visceral fat is strongly associated with
metabolic disease risks [21,45]. Several studies stated that n3:
PUFA had a greater effect on weight loss in men than women
, whereas other studies found stronger effects in women
. Difference among the sexes in the physiological response
to omega: 3 is plausible because men and women have a
different fat tissue anatomy and physiology.
In evidence of some animal studies, supplementation
with DHA+EPA is effective in reducing weight and fat mass
in mice  while others have shown no significant effect
[59,60]. Studies involving human subjects have also stated the
conflicting results, as found in present review. In reference
of the positive outcome, a small number of severely obese
women following a Very Low-density lipoproteins reported
a significant weight loss in the group that also consumed LC
N:3PUFA (2·8g/day fish oil; EPA: DHA = 2:1) compared to the
Reduced levels of CRP, a biomarker of CVD risk, are directly
related to a reduction in weight loss. However, in study by
Ramel et al. , there was a 2: fold greater decrease in CRP
levels for group consuming highest percentage of omega: 3
during the weight loss phase as compared to placebo group,
and CRP continued to decrease during the maintenance phase
also. This would suggest that a greater decrease in CRP for
FO could be attributed to LCN:3PUFA . There were some
studies mentioning or reporting conflicting findings, with one
study stating that DHA+EPA intake but not weight loss was
associated with a significant reduction in CRP , on the
other hand, Madsen et al.  reported that LC N:3 PUFA had
no effect on serum concentrations of CRP . Differences in
the sample size and age of the two groups and the length of the
intervention could account for the conflicting results. Because
of the current limited information, it is difficult to completely
explain why fish oil decreased abdominal fat.
Unlike previous reviews [51,64], which have intervention
period of not more than 6 months whereas in our review
included trials were of longer duration maximum up to 1 year.
In addition to this, among all the studies the highest dose of 6
g of LC N:3 PUFA/ day was also given to seek for the positive
The included parallel and crossover trials had many
differences including subject health status, nutritional therapy,
physical activity modification, fish oil supplement doses (1:6g/
day) and compositions (EPA: DHA), follow: up durations, and
treatment methods (parallel single blind or double blind/
crossover trial). Secondly, the comparison was made within
trials with wide range of difference in treatment durations (4
to 48 weeks). Moreover, the meta: analysis method of analysis
was not used in this review. Also, only 2 electronic databases
were used for searching articles, in comparison to other
reviews that have used Medline, Embase, Cochrane Library
and many others for getting relevant records. This might be an
important reason masking why the anti: obesity effects of fish
oil supplementation were not as obvious in humans as animals.
Omega:3 fatty acid supplementation especially DHA & EPA
has been widely utilised for a variety of medical conditions
that includes CVD, hypertension, T2DM and arthritis. Till
date, results of studies investigating their impact on obesity
development and progression of disease are not clear. In
conclusion, from the results of our review, no effective proof
was found to say that LC N:3 PUFA intake can definitely
decrease body weight in overweight/obese adults.
Although, the majority of studies included in our review
did not concluded the same in their findings. Besides, only few
studies stated the potential role of LC N:3 PUFA in attenuating
the weight gain in maintenance phase when accompanied with
lifestyle modifications (energy restriction and raising physical
activity level). On the other hand, some preliminary evidence
was founded that an increase in LC N:3 PUFA intake within
same dose range might reduce the postprandial sensations
of hunger, which helps in weight management. Data from a
number of studies also suggest that LC N:3 PUFA might promote
increases in plasma HDL levels and heart rate variability thus
potentially increasing the elasticity in large: small arteries and
indirectly assisting with CVD risk reduction. It is also possible
that the increases in plasma omega 3 concentrations, have a
beneficial effect in reducing the blood glucose, HOMA:IR levels
in type 2 diabetic patients even at lower dose levels (1.5g/day).
While there is growing evidence that LC N:3 PUFA can
improve body composition in humans, there is still much of
contradiction as the majority of studies are of relatively short
duration. Accordingly, there is an urgent need for longer:
term studies and meta-analysis with appropriate dosage recommendation as per WHO i.e. 2:4g/day to determine the
result of long chain omega:3 supplementation on body weight
in overweight and obese populations. Moreover, a proper
assessment of parameters like plasma concentration of EPA/
DHA fatty acids, increases in the n3/n6 ratio of plasma FA and
percentage of N3 FA in serum phospholipids should be included
for effective study of proposed outcome. This would help in
getting a definite conclusion.