algae source of docosahexaenoic acid increases (n-3) fatty acid status

8/4/2019 Algae Source of Docosahexaenoic Acid Increases (N-3) Fatty Acid Status

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Human and Clinical NutritionSupplementation with an Algae Source ofDocosahexaenoic Acid Increases (n-3) Fatty Acid Statusand Alters Selected Risk Factors for Heart Disease inVegetarian Subjects1'2

JULIE A. COHQER AND BRUCE J. HOLB3Department of Human Biology and Nutritional Sciences, University of Guelph,Guelphi, Ontario , Canada NI G 2W1

ABSTRACT The purpose of this double-blind studywas to inv estig ate th e influenc e o f die ta ry supplementation with an algae source of docosahexaenoic acid[DMA;22 :6 (n -3 )], d evoid o f any e icosapentaenoic acid[EPA; 20:5(n-3)], on serum /platelet DHA status, theestim ated retroconversion of DHA to EPA, and riskfactors for heart disease in vegetarian subjects.H ealthy vegetarians (12 male, 12 female) consum ednine capsules daily of either DHA (1.62 g/d) or cornoil for 6 wk. Consumption of DHA capsules increasedDHA levels in serum phospholipid by 246% (from 2.4to 8.3 g/100 g fatty acids) and in platelet phospholipidby 225% (from 1.2 to 3.9 g/100 g fatty acids). EPAleve ls in crea sed in serum pho spho lipid by 117% (from0.57 to 1.3 g/100 g fatty acids) and in platelet phospholipid by 176% (0.21 to 0.58 g/100 g fatty acids)via metabo lic retro conv ersion; th e estimated ex tent o fDHA retroconversion to EPA was 11.3 and 12.0%,based on the serum and platelet analyses, respecti ve ly . Arach idon ic acid [AA; 20:4 (n -6 )J leve ls in s erumand pla telet ph osph olipids d ecrea sed mod era tely du ring the trial period (DHA group) as did both docosa-p enta eno ic ac ids |22 :5 (n -6) and 22:5(n -3 )|. A lthoughn o significan t chang es w ere found in th e to ta l and LDL -cho le ste ro l le ve ls w ith DHA supplementa tion, th e to ta lcho le ste rohHDL-cho le ste ro l ratio showed a moderatedecrease over tim e as did the LDL -cholesterol:HDL -cho le ste ro l ratio and serum tr ig lycr id econcentrations. DHA supplem entation did not alter the variousthrombogenic factors m easured. In conclusion, DHAsupplemen ta tion mark edly enhanced th e DHA sta tus(of serum and platelets), provided for the formationof substantial EPA, and lowered the total and LDL-cholesterohHDL-cholesterol ratios. J. Nutr. 126:3032-3039,1996.INDEXING KEY WORDS:humans cholesterol vegetarian DHA supplementationhrombogenic factors

In contrast to the omnivorous diet, the vegan diet isdevoid of the long-ch ain (n -3) fa tty acids, eico sapentaenoic acid [EPA ,4 20:5(n-3)] and docosahexaenoic acid[DHA , 22:6(n-3)], commonly found in fish and fish oils.Although vegans consume no EPA or DHA (Pan et al.1993, Sanders et al. 1978), vegetarians consuming eggs(Simopoulos and Salem 1992) consum e small amountsof both. Thus, in the vegetarian diet, a-linolenic acid[aLNA, 18:3(n-3)] is the major dietary (n-3) fatty acid.The efficiency of conversion of this fatty acid to DHA,in healthy adults, via desaturation and elongation, appears to be ~5% (Emken et al. 1994). Various studieshave confirmed that vegans/vegetarians have lower serum and/or platelet phospholipid levels of DHA thando omnivores (Reddy et al. 1994).DHA is found in high levels in the brain and retina,where it functions in mental performance and visualacuity, respectively (reviewed in Neuringer et al. 1988).Several investigators have shown that dietary fish/fishoils containing EPA plus DHA may help reduce therisk for developing cardiovascular diseases (CVD ) (forreview see Herold and Kinsella 1986). Fish oils con-

1Su pp orted b y th e Hea rt an d S tro ke F ou nd ation o f Ontario . J.A .C ,w as a recipient of a H eart and Stroke Foundation of O ntario researchfellowship.2 The costs of publication of this article were defrayed in part bythe paym ent of page charges. This article m ust therefore be herebymarked "advertisement" in accordance with 18 USC section 1734solely to indicate this fact.3 To whom correspondence and reprint requests should be addressed.4Abbrevia ti on s u sed: AA, a ra ch idon ic a cid ; ACD, aci d c it ra te d extrose; CVD , cardiovascular disease; DHA , docosahexaenoic acid;EPA , e ico sap en ta en oic ac id ; a LNA, a-lin ole nic a cid ; PPP , p latelet-poo r p la sma; PRP , p la te le t- ric h p la sma; PUFA, polyunsatu ra ted f att yac id s; T xA2, th rombox an e A2; TxB2; th rombox an e B2.

0022-3166 /96 $3.00 1996 Ameri can Ins tit ute o f Nut rit ion.M anuscript received 5 June 1996. Initial review com pleted 5 July 1996. R evision accepted 22 A ugust 1996.3032


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DMA AND HEART DISEASE RISK FACTORS 3033taining EPA plus DHA lower serum triglycridelevelsin humans (for review see Harris 1989) and reduceblood platelet reactivity (for review see Weaver andHolub 1988). These latter effects have often been attributed indirectly to EPA due to its predominance in mostfish oil supplem ents/concentrates studied to date. Isolated studies with DHA-enriched preparations containing significant levels of EPA have shown a reduction in platelet aggregation (von Schacky and Weber1985 ) and a lowering of serum triglycride conc entrations in human volunteers (Sanders and Hinds 1992).DHA has been found to inhibit human platelet reactivity in vitro (Gaudette and Holub 1991).A lthough vegetarians tend to be at overall decreasedrisk for CVD (Dwyer 1988), due partly to their lowerserum cholesterol levels, their thrombogenic risk factors may be significant (Pan et al. 1993). Consideringalso the lower DHA status in vegetarians, it was ofinteres t to determ ine th e effect of oral supplementationwith a DHA-rich/EPA-free source (from micro-algae,non-fish derived) of triglycride oil (DHASCO ) inthis group. In addition to evaluating the potential forDHASCO to enhance the DHA and EPA (via retro-conversion) status in vegetarians, the influence of dietary DHA on conventional (serum total cholesterol,lipop roteins an d trig lycrides) a nd thrombog enic cardiovascular risk factors (platelet aggregation, throm-boxane production, serum viscosity, factor Vile and fi-brinogen levels) w as determ ined. Measures of total cho-lestero l:HDL-cholesterol and LDL-cho lesterol:HDL-cholesterol ratios were of particular interest becauserecent reports indicate that they are superior predictorsof coronary heart disease risk com pared with total cholesterol and LDL-cholesterol levels (Kinosian et al.1994 and 1995).

SUBJECTS AND METHODSSubjects and experimental design. The subjectswere 24 healthy vegetarians (12 male, 12 female) selected from the Guelph community who reported having no meat, poultry or fish for a period of at least 6mo. Approval for this double-blind study was granted

by the Human Ethics Committee of the University ofGuelph and written informed consent was obtainedfrom each subject. The 24 subjects (aged 29.6 1.7 y,mean SE)were randomly assigned (12/group) to thetwo supplem entation groups, DHA-supplemented andcontrol (6 males, 6 females each). The DHA-enrichedencapsulated triglycride oil, DHASCO (algae derived), and placebo (corn oil) capsules were kindly provided by David Kyle of Martek B iosciences, Columbia,MD. As confirmed by quantitative gas-liquid chro-m atographic analyses, the DHASCO oil containedDHA as the major fatty acid (39 g/100 g fatty acids)

VariableHeight,mWeight,kgBMI,2g/m?Protein,% ofnergyFat,% ofnergyCarbohydrate,% ofnergyAlcohol,% of energyControl1.7

.067.7.422.9.011.80.625.4.758.4.14.21.9DHASCO1.767.122.910..3

TABLE 1Sub je ct c ha ra cter istics and e stimated d ieta ry in ta ke durin gdocosahexaeno ic aci d o r p lacebo supplementat ion!

1Values are reported as means SEM ,n = 12 (height, weight,BMI) o r n = 10 ( protein , fat, c arb ohyd ra te , an d a lc ohol a s a p erce nta geo f ene rgy in take ). No s ignif ic an t d if fe rences between the g roups werefo und fo r th e ab ov e v ariab les.2 BMI, body m ass index.followed by myristic acid (21%), palmitic acid (16%),oleic acid (11%) and lauric acid (8%); no EPA was detected. Linoleic acid (53%), oleic acid (21%) and palmitic acid (10%) predominated in the vegetable oil(co rn oil) placebo. E ach DHASCO capsu le c ontained0.5 g oil and was estimated to provide 180 mg DHA.The experimental group consumed nine DHA-containing capsules per day (total 1.62g DHA /d) w ith theirmeals, and the control group consumed nine vegetableoil placebo capsules per day. Each group consumed thecapsules for a period of 42 d beginning on d 0. After 42 dof capsule ingestion, both groups completed a washoutperiod for 21 d during which there w as no supplem entation. Subjects were weighed on each visit (d 0, 21, 42and 63) and height was measured at entry; there wereno significant differences in these or other characteristics betw een the groups at entry Table1). The amountof fat consumed in encapsulated form (4.5 g/d) represented


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3034 CONQUER AND HOLUBdextrose (ACD) [25 g/L Na citrate, 20 g/L dextrose, 14g/L citric acid (all from Fisher Chemicals, Nepean,C anada)], orN a citrate (3.2% ). Whole blood was treatedas follows: Tube without anticoagulant. W hole bloodwas centrifuged at 1250 x g for 15 min to obtain serum.Serum was used for measurement of plasma total-,HDL- and LDL-choles terol levels, triglycride levels,serum total phospholipid fatty acid content and serumviscosity. Serum was stored at -20 Cu ntil all sampleswere collected and thawed just before analyses of lip-ids. Serum was stored at room temperature until viscosity w as analyzed (see below ).

Tubes containing Na citrate. Whole blood was centrifuged at 200 X g for 17 min to obtain platelet-richplasma (PRP) which was removed, and the remainingblood was centrifuged at 1250 x g for 15 min to obtainplatelet-poor plasma (PPP). The PRP was used in aggregation studies and the PPP was used for analyses offactor VII and fibrinogen levels. The PPP was storedbelow -20 Cu ntil analysis by Med Chem Laboratories(Scarborough, Ontario) .Tube containing ACD. Washed platelet suspensionswere prepared according to the method of Turini et al.1994.Platelet aggregation. Platelet aggregation was performed within 2 h of blood collection. Platelets werecounted in PRP using a Coulter Counter, model ZM(Coulter Electronics, Burlington, C anada) and adjustedto a final concentration of 2.0-2.5 x IO11 platelets/Lusing autologous PPP. A liquots (0.5 mL) of adjusted PRPwere preincubated for 1 m in in siliconized cuvettes w ithstirring at 900 rpm at 37 Cn a dual-channel 800B aggre-gometer (P ayton Instrum ents, Ion Trace, S carborough,C anada) before addition of the aggregating agent. Collagen (Hormone-Chem ie, Mnchen ,Germany) was addedat two levels with the final concentrations of 2 and 10mg/L. The platelets were allowed to aggregate for 2 minfollowing the addition of agonist and then the level ofagg regation was measu red (Born 1962).Cholesterol and triglycride measurement. Totalcholesterol was measured enzymatically with a diagnostic test (Sigma Diagnostics Procedure No. 352, St.Louis, MO). HDL-cholesterol was isolated by using adextran sulfate and Mg ion solution to precipitate theVLDL and LDL from the serum sample. The HDL fraction was then assayed by an enzymatic assay (SigmaDiagnostics Procedure No. 352-3). Triglyc ride wasmeasured enzymatically with a diagnostic test (SigmaDiagnostics Procedure No. 339). LDL-cholesterol wascalculated using the formula validated by Friedewaldet al. (1972). Analyses of all samples for each volunteerwere performed in a single assay.Total phospholipid analysis. The fatty acid compositions of total phospholipid from serum and washedplatelet suspensions were determ ined following lipidextraction, TLC, transmethylation of fatty acid resi

dues and gas liquid chromatography by procedures simila r to thos e prev iously describ ed (Donad-oet al. 1 994).Fibrinogen, factor Vile and serum viscosity. F ibrinogen, factor Vile and serum relative viscosity were analyzed by Med-Chem Laboratories LTD. Serum relativeviscosity was determined on serum (maintained atroom temperature) within 16 h of blood withdrawal.The relative viscosity is the flow time of specimen inseconds/flow time of water in seconds in an OswaldV iscosimeter tube (G radwohl 1970). The concentratio nof fibrinogen in plasma samples was determined bya diagnostic test comparing thrombin time values ofdiluted plasma samples (diluted with Ortho Q.F.A.Buffer, Ortho Diagnostic Systems, Johnson and Johnson, Raritan, N J), to w hich O rtho Q.F.A. Throm bin wasadded, to a reference curve (Clauss 1957). The log ofthe thrombin time value is inversely proportional tothe log of the fibrinogen concentration. To determinefactor Vile levels, prothrombin time was determinedusing Thromborel S (B ehring D iagnostics, Westwood,MA) in conjunction with plasma deficient in factor Vile(Quic k 1935).Thromboxane production. Aliquots of PRP (0.5mL) were stimulated with collagen (2 mg/L) for 2 minand reactions stopped by the addition of 250 mo(final concentration) indomethacin (Sigm a) and 4.5 g/L NaCl (final concentration) and were immediatelychilled on ice. Samples were centrifuged at 2000 x gfor 15 min at 0C.The supernatant was removed andstored at -20 Cu ntil analysis using the ThromboxaneB2B iotrack enzyme immunoassay system (AmershamCanada, O akville, Ontario, C anada).Statistical analysis. All data are reported as meansSEM.Data that were not normally distributed weretransform ed (log or square) before analysis to reach normality. When observatio ns were m issin g, lea st-squaredmeans were calculated so that means could be compared. Split-plot design, including tim e and treatm entas factors, w as used in all analyses before exam inationof specific differences by Fisher's protected LSD. Specific differences for preplanned comparisons betw eentreatment means as well as comparison between dietary intakes and subject characteristics were examined by f test (K irk 1968). Statistical analyses w ere doneusing the SAS system (SAS Institute, Cary, NC).Changes w ithin groups as w ell as comparisons betw eengroups were m ade.

RESULTSThe fatty acid compositions of the serum total phospholipid (controls and DHA-supplemented) at entry(wk 0) and after 3 and 6 wk are given in Table 2. Thelevels of the various (n-3) polyunsaturated fatty acids(PUFA) were not different between the two groups at


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3036 CONQUER AND HOLUBTABLE 3

Fatty acid composition of total ph osph olipid in human platelets before and after supplem entation w ith docosahexaenoic acid or placebo!Control DHASCO

WeekFatt y a ci d

g/100 g fatty acids16:018:018:118:2|n-6]20:3|n-6)20:4jn-6|

(AA)220:5(n-3)(EPA)22:4(n-6)22:5(n-6|22:5|n-3)22:6(n-3)

(DHA|(n-6)/(n-3)EPA/AADHA/AA22:5(n-6)/22:6(n-3)16.8

.417.4.4a17.5.5a,b6.10.3a1.3.6a,b24.2.6b0.24.04a2.1.lb0.19.04b1.71.111.60.010.04O.lbO.la0.5CO.OaO.Oa0.17

0.03b,c17.2

.4a,b16.80.3a16.9.3a,b6.70.2b1.30.1a,b25.5.4b0.25.0432.1.lb0.170.05b1.71.111.90.010.04O.lbO.la0.5C0.03O.Oa0.14

0.04b17.117.416.75.71.124.40.290.4a,b0.4a0.4a0.5aO.la0.8b0.0432.2

.lb0.28.03C1.71.111.20.010.05O.lbO.la0.5CO.OaO.Oa0.25

0.03C17.116.917.06.01.323.60.212.40.181.71.210.70.010.050.3aO.Sa0.43,b0.2a.03b18.016.717.56.81.423.80.481.30.3b0.230.5a,b0.2bO

.0030.66.0533.80.lb6.7.3a0.02.Ob0.16.Ob0.00O.OOa17.8

.3b16.3.6a17.9.5b6.7.3a,b1.50.lb21.7.730.58.OSb1.2O.la0.01.Ola0.71.0733.9.2b6.1.330.03.Oc0.18.Oc0.00O.OOa

1V alues are reported as m eans SEM,n = 12. D iffering superscripts in a row indicate significant differences, P < 0.05.2 AA, ara ch id on ic ac id ; EPA , e ic osap entae no ic a cid; DHA , d oco sa hex ae no ic ac id.phospholipids within 3 wk and maintain these levelsover the 6 wk supplementation period (Tables 2 and3). Previous studies have shown that fish oils, whichcontain EPA plus DHA, raise both EPA and DHA levelsin the platelet and serum phospholipids of omnivoroussubjects (Herold and Kinsella 1986, W eaver and Holub1988). The (n-3) fatty acid level in serum phospholipidis regarded as a useful biochem ical index of the dietaryintake and overall nutritional status for EPA/DHA (Bo-naa et al. 1992). The level of DHA attained (8.3 g/100g total fatty acids in serum phospholipid) is above thelevels reported in free-living omnivorous subjects (Bo-

naa et al. 1992). The net rise in EPA in serum phospholipid (0.7 g/100 g fatty acids) was ~one eighth that ofthe corresponding DHA rise (Table 2). The EPA risewould reflect fatty acid retroconversion from DHA(Voss et al. 1992) with an estimated efficiency of 11.3and 12.0% based on the mol/100 mol values for serumand platelets, respectively (net rise in EPA/net rise inEPA + DHA as %) as derived from the data in Tables2 and 3. Interestingly, the decrease in the (n-6) polyun-sa tu ra te s [20:4 (n -6 ), 22:4 (n -6 ) and 22:5 (n -6 )] in p la te le tand serum phospholipids (Tables 2 and 3), as observedin our study as well as with fish oils having EPA plus

TABLE 4E ffect of supplem entation with docosahexaenoic acid or placebo on human serum lipid and lipoprotein levels1

Control DHASCOWeek

VariableTotal cholesterol ,mol/LHDL-cholesterol,mmol/LTotalcholesterol:DL-cholesterol,moLtnolLDL-cholesterol,mmol/LLDL-cholesterol:HDL-cholesterol,mol:molTriglycride,mmol/L3.83

.15a1.35.09b3.0.2b2.10.1831.7.2b,c0.810.08a,b,c3.7 8

.15a1.400.1lb,c2.90.2a,b2.02.18a1.6

.2b0.770.10a,b4.160.19b1.45

.lOc3.00.2b2.290.1b1.7

.2b,c0.920.14b,c3.63

.20a1.20.lOa3.20.3b1.99.1831.8

.2C0.960.1ic3.67

.26a1.36.12b2.8.2a,b1.97.2131.6.2b0.750.09a3.63

.24a1.40.13b,c2.7.2a1.86.171.4

0.20.80O.lla,bV alues are reported as m eans SEM,n = 12. D iffering subscripts in a row indicate significant differences, P < 0.05.


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DHA AND HEART DISEASE RISK FACTORS 3037TABLE5

Effec t of supplemen ta tion with docosahexaenoic acid or p lacebo on selec ted thrombogen ic r isk factor s in vegetar ians!Control DHASCO

WeekVariableP la te le t a gg re ga ti on ,2 %fmaximumaggregationwith10 mg/ LollagenTxBjproduct ion, ng/2.510plateletsRelativeserumViscosityrelativ e toater,wherewater =.0Fibrinogen,g/LFactorVile,prothrombin time83.5

196.31.52.3

119.94.626.90.00.110.587.3186.01.42.299.3

2.421.80.10.1

7.389.5189.11.52.3125.3

1.924.30.00.1

10.486.1180.61.51.8117.8

3.228.20.00.1

11.086.9

176.11.41.9

103.12.917.90.10.15.588.7

159.21.51.9

122.01.724.

1 V alues are reported as m eans SE M,n = 12. N o significant differences due to supplem entation w ere found.2 V alues for platelet aggregation are for 2 m g/L collagen as a percentage of m axim um aggregation w ith 10 m g/L collagen.3 T xB 2, throm boxane 82.

DHA, was further accompanied by a marked decreasein docosapentaenoic acid [22:5(n-3)j. The latter fattyacid generally exhibits considerable enrichment in serum and platelet phospholipids w hen fish oils [providing EPA, DHA and some 22:5(n-3)] are fed (Herold andKinsella 1986, W eaver and Holub 1989). Thus, dietaryD HA itself appears to replace both 22:5(n-6) and 22:5(n-3) in circulating and cellular phospholipids in humansubjects, likely by com petition from D HA -containingprecursors, via com bined de novo phospholipid biosyn-thetic pathw ays and deacy lation/reac ylation reactions.Clinical trials with fish oils containing EPA andDHA (Harris 1989) have generally shown a significantd ec reas e in se ru m trig ly crid econ centra tio ns and o ftena rise in LDL-cholesterol. These trials (Harris 1989)have employed fish oil concentrates which usually contain much more EPA than DHA. Because the EPA-freeDHASCO used in this study as well as the EPA-containing concentrate of DHA used previously (vonSchacky and W eber 1985) both lowered serum triglyc rideconcentrations, dietary D HA m ay produce a tri-gly ceride-low ering effect inde pendent o f E PA . T he relatively minor retroconversion of DHA to EPA is likelyinsufficient to account for the observed alterations inserum lipids and lipoproteins. In our study, the use ofE PA -free D HA SC O indicated th at dietary D HA itselfcan provide a moderate reduction in the total-cho-lesterohHDL-cholesterol ratio and the LDL-choles-terohH DL -cholesterol ratio as w ell as a sligh t redu ctionin serum triglyc rideconc entrations of vegetaria n subjects (Table 4). These changes are consistent with afavorable influence of dietary D HA on these recognizedlipid/lipopro tein risk factors for cardiovascular dise ase(D rexel et al. 1994). Recent publications (K inosian et al.

1994 and 199 5) indica te tha t the total cholesteroh HD L-cholesterol and LD L-cholesterol:H DL-cholesterol ratios are better predictors of heart disease than totalcholesterol or LDL-cholesterol levels alone. Supplementation with fish oils containing EPA plus DHA forseveral weeks has generally not significantly reducedL DL -choleste rol:H DL -c holesterol ratios (H arris 1989).It remains to be studied whether the moderate lipid/lipoprotein shifts as seen in our vegetarian subjects onrelatively low fat diets (23% of energy) are also seen inother population and dietary groups.A lthough m any previous studies, often using intakesof EPA plus DHA ranging from 3 to 6 g/ ,have showna significant reduction in collagen-induced platelet aggregation and thromboxane form ation, this was not observed (Table 5) in our subjects consuming 1.62 gDHASCO/d for 6 wk. One previous study, usingmuch higher levels of DHA (6 g/d) over 6 d, reporteda reduction in collagen-induced aggregation (vonSchacky and Weber 1985). Further, the absence of anysignificant effect of DHA supplementation on otherthrom boge nic risk fac tors (viscosity, fibrinog en, factorVile) in our trial can be compared with various studieswith fish oils (EPA/DHA) in which either no changeor some beneficial effects have been reported (Harris1989, Herold and Kinsella 1986, Saynor and Gillott1992, W eaver and Holub 1988). Future studies whichemploy varying dietary levels of DHA, including varying EPA/D HA ratios, will be of interest.The marked rise in the DHA level of serum andplatelet phospholipids of our subjects is of interest inview of recent epidemiological studies which haveshown an inverse relationship between DHA levels inthe population (both diet and blood) and the risk of


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3038 CONQUER AND HOLUBCVD (Leng et al. 1994, Simon et al. 1995). Thus, partof the c ardioprote ctive effect of fish/fish oils containing(n-3) PUFA appears due to DHA in addition to EPA.The cardioprotective effects of (n-3) fatty acids are considered to be mediated by a number of physiologicaland biochemical mechanisms; included among theseare studies which indicate that the enrichment of hearttissue in DHA provides an antiarrhythmic effect (Pepeand McLennan 1996), which may account for the reduction in cardiac arrest and sudden cardiac death inthose having a higher DHA status. In addition to theabove, dietary DHA intakes and increased status inthe body have been implicated in favorable effects onattentio n-deficit disorders (S tevens e t al. 1 995), depression and anxiety disorders (Hibbeln and Salem 1995),as well as protection against breast cancer in postm eno-pausal women (Zhu et al. 1995).In conclusion, the consumption of 1.62 g of an animal-free source of DHA per day by vegetarians readilyenhances their DHA status, provides for EPA formationbased on serum and platelet phospholipid analysis, andexerts moderately favorable (low ering) effects on thetotal cholesterol:HDL -cholesterol ratio, the LDL-cho-lesterol:HDL-cholesterol ratio, as well as serum triglycrideconcentrations. The present studies, conductedwith young non-hyperlipidemic vegetarian adults ofboth genders, should now be extended to other groups(e.g., omnivorous normolipidemics and hyperlipid-emics).

ACKNOWLEDGMENTSWe would like to thank Margaret Berry, IndManiand Melinda Gooderham for their help in all aspects ofthis investigation. We would also like to thank DavidKyle and Eeva-Kaarina Koskelo of Martek BiosciencesCorporation (Columbia, MD) for supplying both thecontrol and DHA capsules for this investigation. Appreciation is also extended to Martek for covering thecosts of the fatty acid analyses as provided by LipidAnalytical Laboratories, University of Guelph Research Park (Guelph, Ontario, C anada).

LITERATURE CITEDBonaa, K. H., Bjerve, K. S. & Nordoy, A. (1992) Habitual fish consump tio n, plasma p ho sp ho lip id fa tty ac id s, a nd se rum lip ids : th eT rom so study. Am. J. C lin. N utr. 55: 1126-1134.B orn, G .V .R. (1962) A ggregation of blood platelets by adenosined iph osp hate a nd its re versa l. N atu re (L on d.) 1 94 : 92 7-9 29.C lauss, A . (1957) G erinnungsphysiologische S chnellm ethode zurbestimmung des fibrinogens. A cta H aem atol. 17: 237.Donadio, J. V ., Jr., Bergstrath, E. J., Offord, K. P., Spencer, D. C. &Holley, K. E . (1994) A controlled trial of fish oil in IgA ne-p hro pa th y. N . E ngl. J. Med . 3 31 : 11 94 -1 19 9.

D rexe l, H ., Amann, F . W., Ber an , J ., Ren ts ch , K ., Cand-nas ,R ., Mun t-wyler, J., Luethy, A., Gasser, T. & Follath, F. (1994) Plasmat rig ly cr id esand th re e li poprot ei n cho le st ero l f ra ct ions a re independent risk factors of the extent of coronary atherosclerosis.C ircu la tion 90: 2230-2235 .Dwyer, J. T . (1988) H ealth aspects of vegetarian diets. Am. J. C lin.Nut r. 48: 712 -738 .Emken, E. A., Adlof, R. O. & Gulley, R. M . (1994) Dietary linoleica ci d i nfl uences des at ur ati on and acy la tion o f deu te ri um-l abell edlinoleic and linolenic acids in young adult m ales. B iochim . B iophy s. Acta 1213: 277 -288 .Ferrier, L. K., Caston, L. J., Leeson, S., Squires, J., W eaver, B. J . &H olub, B . J. (1995 ) a-Linolenic acid- and docosahexaenoic acid-enriched eggs from hens fed flaxseed: influence on blood lipidsand platelet phospholipid fatty acids in humans. Am. J. Clin.Nutr. 6 2: 81 -8 6.Friedew ald, W . T., Levy, R . I. & Frederickson, D . S . (1972) E stim atio n o f th e c on ce ntratio n o f low-de nsity lip op ro te in ch olestero lin plasma w ithout use of the preparative ultracentrifuge. Clin.Chem. 18: 499 -502 .Gaudette, D. C. & Holub, B. f . (1991) Docosahexaenoic acid(DHA) and human platelet reactivity. J. Nutr. Biochem. 2116-121.Gradwohl, R.B.H. (1970) Gradwohl's Clinical Laboratory M ethods and Diagnosis. A Textbook on Laboratory Procedures andTheir Interpretation (Frankel, S., R eitm an, S. & Sonnenw irth,A . C ., eds.), 7th d.p. 663-664. M osby, Saint Louis, MO.Harris, W . S. (1989) Fish oils and plasma lipid and lipoprotein metabolism in hum ans: a critical review . J. L ipid R es. 30: 785-807.Herold, P. M . & Kinsella, J. E . (1986) Fish oil consum ption and decreased r isk of cardiovascu la r d isease : a compar ison of f indings f romanimal an d h uman fee din g trials. Am. J. C lin . Nutr. 4 3: 5 66 -5 98 .Hibbeln, J. R . S.alem, N. (1995) Dietary polyunsaturated fattyacids and depression: w hen cholesterol does not satisfy. Am. JClin. Nutr. 62: 1-9.Kinosian, B., C lick H. & Garland, G. (1994) Cholesterol and coronary heart disease: predicting risks by levels and ratios. A nn.I nt er n. Med . 121 : 641 -647 .K in osian , B ., C lick , H ., P reiss, L .& .Pu de r, K . L . (1 99 5) Cho le stero land coronary heart disease: predicting risks in men by changesin levels and ratios. J. Invest. M ed. 43: 443-450.K irk, R . E . (1968) Experim ental D esign: Procedures for the Behaviora l S cien ce s. B ro ok s Cole P ub lish in g Compa ny , B elmon t, CA.Leng, G. C., Horrobin, D. F., Fowkes, F.G.R., Smith, F. B ., Lowe,G.D.O., Donnan, P. T. & Ells, K. (1994) Plasma essential fattya cids , cig are tte smokin g, a nd d ietary an tio xid ants in p erip hera la rt er ia l d is ea se . A rt er io sc le r. Thromb. 14: 471 -478 .Neuringer, M ., Anderson, G. J. & Connor, W . E. (1988) The essentiality of n-3 fatty acids for the development and function of theretina and brain. A nn. R ev. N utr. 8: 517-541.Pan, W .-H ., Chin, C.-J., Shen, C.-T. &. L ee, M .-H. (1993) Hemo-static factors and blood lipids in young B uddhist vegetarians andom nivores. Am. J. Clin. N utr. 58: 354-359.Pepe, S. & McLennan, P. L. (1996) Dietary fish oil confers directantiarrhythmic properties on the myocardium of rats. J. Nutr.126: 34-42.Quick, A. J. (1935) A study of the coagulation defect in hemophilia and in jaundice. Am . f. M ed. Sci. 190: 501.Rper,N. R., Cronin, F. J. & Exler, J. (1992) Omega-3 fatty acidC ontent of the U .S. food supply. J. Am. C oll. N utr. 11: 304-308.Reddy, S., Sanders, T.A.B. & Obeid, O. (1994) The influence om aternal vegetarian diet on essential fatty acid status of the newborn. Eur. J. C lin. N utr. 48: 358-368.S anders, T.A .B ., Ellis, F. R . & D ickerson, J.W .T . (1978) Studies ovegans: the fatty acid com position of plasm a choline phospho-g ly cerid es, e ryth ro cy te s, ad ip os e tis su e a nd bre ast m ilk an d someindicators of susceptibility to ischem ie heart disease in vegansand controls. Am. J. Clin. N utr. 31: 805-813.


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O,:HA AND HEART DISEASE RISK FACTORS 3039Sanders, T.A.B. & Hinds, A. (1992) The influence of a fish oil highin docosahexaenoic acid on plasma lipoprotein and vitamin E

concentrations and haemostatic function in healthy m ale voluntee rs. B r. J. Nutr. 6 8: 1 63 -17 3.Saynor, R. & Gillott, T. (1992) Changes in blood lipids and fibrin-ogen with a note on safety in a long term study on the effectsof n-3 fatty acids in subjects receiving fish oil supplem ents andfo llowed fo r s ev en y ea rs. L ip id s 2 7: 5 33 -5 38 .

Simon, J. A., Hodgkins, M. L., Browner, W. S., Neuhaus, J. M.,Bernert, J. T. & Hulley, S. B . (1995| Serum fatty acids and therisk o f co ron ary h ea rt dise ase . Am. J. E pid em io l. 1 42: 4 69 -47 6.

Sim opoulos, A. P. &. Salem, N., Jr. (1992) Egg yolks as a source oflong chain polyunsaturated fatty acids in infant feeding. Am . J.C lin . Nutr. 5 5: 4 11 -41 4.

S te ven s, L . J ., Z en ta ll, S . S ., D ec k, J . L ., Aba te , M . L ., Watk in s, B .A .,Lipp, S. R. & Burgess, J. R . (1995) Essential fatty acid metabolism in boys w ith a tten tio n-d eficit h yp erac tiv ity d iso rde r. Am. J.C lin . Nutr. 6 2: 7 61 -7 68 .

Turini, M. E., Powell, W. S., Behr, S. R. &. H olub, B. f. (1994) Effects of a fish oil and vegetable oil formula on aggregation andethanolam ine-containing lysophospholipid generation in activated hum an platelets and on leukotriene production in stimulated neutrophils. Am. J. Clin. N utr. 60: 717-724.

von Schacky, C. & Weber, P. C. (1985) Metabolism and effects onp la te le t func ti on o f t he puri fi ed e icosapen ta enoi c and docosahexa en oic a cids in h uman s. J . C lin . Inv est. 7 6: 2 446 -2 450 .Voss, A ., R ein ha rt, M . & Spre ch er, H . (1 99 2) D iffe ren ce s in th e in ter

conve rs ion between 20- and 22- carbon (n-3) and (n-6) polyunsatur ated f at ty acid s i n r at l iv er. B ioch im . B iophys . Act a 1127: 33-40 .Weaver, B. J. & Holub, B. J. (1988) Health effects and metabo

lism of dietary eicosapentaenoic acid. Prog. F ood Nutr. Sci. 12:111-150.Zhu , Z . R ., Agren , J., Mannisto , S ., P ie tin en , P ., E sk eline n, M ., S yrja-

nen, K .& .U usitupa, M . (1995) F atty acid composition of breastadipose tissue in breast cancer patients and in patients with ben ig n b re ast d isea se . Nutr. C anc er 24 : 1 51 -1 60.

algae source of docosahexaenoic acid increases (n-3) fatty acid status

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