1098

Metaboliccharacteristics

of diacylglycerolAn edible oil

that is less likelyto become body fat

This article is byNohoru Matsuo and

tchiro Tckimiteu of theBiological Science Laboratories

at Kao Corporationin Todrigi, [apan,

Volume 12 • November 2001 • Inform

Biochemistry

Triacylglycerol Diacylglycerol

- ......- • .:'"'1' ............

EI:~HzOCOR,

~HOCORzCH,OCOR,

•-,,.0:,.-.-

9H,OCOR,

9HOCORzCHzOH

1(l).2-~

R'.I : F.tty Acid

9HZOCOR,

CHOH,CH,OCORz

1.3-~

Figurf! I. $chf!matk structu", of dlacylglycf!rols and triacylglycf!rols.

Obesity is a growing problem world-wide, nor only in industrialized coun-tries but also in the urban areas ofdeveloping countries. According toWorld Health Organization statistics,the number of obese adults increasedfrom 200 million in 1995 to over 300million as of 2000.

Obesity is associated with a numberof health risks, such as heart disease,diabetes mellitus, hypertension, gall-bladder disease, and some types of can-cer. The importance of the prevention ofbody fat accumulation in the preventionof lifestyle-related diseases h.1S beennoted by many experiments and sur-veys. Clinical studies have suggestedrhar weight loss in the range of 5-10%of initial weight can confer significantreductions in obesity-related disorders.

As for countermeasures, functionalfoods and materials that improve fatmetabolism have been extensively stud-ied. Diets generally recommended forweight loss promote energy (caloricintake) restriction and limitation oftotal and saturated fat intake. In con-trast to these approaches, we havefocused on the glyceride backbone,instead of the fatty acid composition ofoils, and have found that diacylglycerol(DAG), particularly in the 1,3·isoforrn,

has metabolic characteristics distinctfrom triacylglycerol (TAG) that mayproduce beneficial effects with regard to

the prevention and management of obe-sity. DAG is a natural component ofvarious edible oils and is currently usedin foods as an emulsifier. A cooking oilproduct containing 80% (w/w) orgreater of diacylglycerol has been on themarker in Japan as a "Food forSpecified Health Usc" since 1999. Inthis article, the nutritional chnmcteris-Ticsof DAG arc outlined.

DAG content in edible oils,its structure and energy valueThe largest constituent of cooking oil isTAG, but it generally also containsbetween a few percent to about 10%DAG, with the relative content depend-ing on rhc origin of thc cooking oil. Forexample, olive oil may contain 5.5%DAG and corronsccd oil, 9.5%. DAGproduced in the digestive tract as ametabolic intermediate after ingestionof TAG is 1,2 (or 2.3)-diacyl-slI-glycerol(1,2-DAG), bur DAG present in cook-ing oil is converted to 1,3-diacyl-slI-glycerol (1,3-DAG) by migration of thencyl group during the manufacruringprocess. Figure I illustrates structuresof DAG in comparison with TAG. Acyl

250 -0- DAG........ TAG

~ 200o•~o'"

150

100

501------...]o 2 4 6 8

Time after fat loading ( hrs )Figure 2. Differentiill effects of DAG andTAG on po$tprandia/s"rum TAG concen-tration. ~rvm TAG WU meu..,~ ilke.....single ingenlon of lipid emulsion (dose oftest 011 '" ]0 &lml of body surface inhealthy men; Hean t SO, n '" 6, -: P < O.OS,":P < 0.01).

migration is an equilibrium reaction,and in oils with :1 common fatty acidcomposition, approximately 70% ofOAG exists as J .J-DAG.

We have studied the differencebetween DAG and TAG (I) regardingfood energy. 111e heal of combustion ofcooking oil containing 87% DAG wascompared with that of TAG oil consistingof the same Farry acid composition byusing a bomb calorimcrcr. As shown inTable I, the food energy value of DAGwas 2% less than that of TAG. Therefore,when 10 g per day of cooking oil (averagecooking oil intake of a Japanese adult) isreplaced with DAG oil, the difference inenergy intake is approximately 0.1 %,which m3Y be considered to be negligible.Furthermore, the absorption coefficientsof the DAG and TAG oils 3S determinedin rats were similar, These results suggest-ed that the physiological differencesbetween DAG and TAG observed in ani-mals and humans, described below, arecaused b)' the different metabolic fatesafter absorption into the gastrointestinalepithelial cells.

,."

T;tble IComparison of the theoretical and experimental eneray valuesof diaeylglyeerol and trlaeylalyeerol

DIACYlGI.YCEROl. OIl. TRIACYlGI.YCEROL Oil

Comribunon ConrribunonComponents Weight to energy Weighf 10 energygtner.uOO fraCtion value fraCtion "alueby hydrolysis (gig oil) (kJ/g oil) (gig oil) (kJ/g oil)

Fatty acids 0.913 35.9 0.956 37.6Glycerol 0.145 2.6 0.105 I.'TOTAL [theoretical] 38.5 39.5

E.xrERIME/'<.'TAL 38.9 39.6

Postprandial lipemiais less after DAG consumptionOne of the nutritional characrerisrics ofDAG is its postprandial hypcrtriglyc-eridemia reducing effect. Figure 2 showschanges of serum TAG concentrationsafter a single ingestion of a lipid emul-sion by healthy men (2). The two testoils consisted of the same fnrry acidcomposition. We have shown in OTherstudies that TAG concentrations in thechylomicron fraction were markedlylower after ingestion of DAG than afterthat of TAG (3). The lower serum TAGlevels after DAG ingestion may be theresult of the slower rate of chylomicronformation after DAG ingestion com-pared with that of TAG ingestion.Serum TAG is a source of fat accumu-lated in the body, and postprandialhyperlipidemia is a risk facror for car-diovascular diseases. With respect to therisk factor, DAG has been shown todiminish the postprandial increase ofremnant-like particles (RLP), which is amore potent risk factor for cardiovascu-lar diseases than the TAG concentrationin the whole serum (2).

Repeated DAG vs.TAG consumptionreduces body fat accumulationTo determine the long-term effect ofDAG in humans, we also have con-

ducred a comparative study of DAG oiland TAG oil ingesrion in 38 healthyJapanese men (4). In this experiment, 10g of the 50 g daily oil was replaced withthe test oil for a 16-week period in adouble-blind study. Table 2 shows bodyweight, waist circumference, andamounts of visceral and subcutaneousfat in the abdominal region. Abdominalfat was measured by computer tomog-raphy scan. Significant decreases wereobserved in the DAG group comparedwith the TAG group in body weight,waist circumference, visceral fat, andsubcutaneous fat (Table 2).

The Chicago Center for ClinicalResearch in Chicago, Illinois, recentlyconducted a parallel double-blind studyin which 127 overweight or obese menand women ingested DAG and TAGdiets for six months. In this study, 15%of energy intake was ingested as the testoil under a mild hypocaloric conditionestablished by subtracting 500--800 kcalfrom the energy requirement calculatedfrom the body weight, activity level, andage. The decreases in body weight andbody fat were significantly higher in theDAG group than in the TAG group.

The effect of the ingestion of DAGoil has been further studied in Japan.After consumption of approximately10 g of DAG oil/day for nine months,

Volume 12 • November 200 •• inform

""Biochemistry

Table 1

Change of anthropometric values and body composition of the subjects"

DlACYLGI.YCEROl GROUr

Week Change/!

Body weight (kg) 0 72.1 " 1.8

i6 69.5,. 1.7##€ -2.6:t O.jUdWaist circumference' (em) 0 85.0:!: 1.4

i6 80.6 " 1.3#11 -4.4 .. 0.6

Visceral f;ll area (em2) 0 79:t 7.0i6 63.7,. 7.0'" -16.2" Z.O·Ott

Subc",anooliS f~l urea (ern2) 0 \48", II

i6 126:t 10## -22.3 '" J.OMlt

"Values 3,..,mean ...SEM (or. III).b U;."'«k val... mmus O·,......k "31...,.€ SignifonTlr d.ff~...,nl from I.... ",ina! ,-al"" br S,udrnf""-Iesr: /I p" 0.05;.' p" 0.01.d S'!llllflC:lmlyd,f(ncnr from maq'lglI'errol d,... group by 5",d ...,,", NCSf for pai...-d val",,: • P c 0.05 ••• Po( 0.01.

~ Siglllf"",mly d,ffnnll frum lrL;Kylgl)'Cnoi d'<1 gmup by anal)";' of CO'"3riance:t p" 0.05.

the physical characteristics such as waistcircumference and the subcutaneous fatthickness were found to be significantlydecreased from the initial values.Furthermore, consumption of a relativelylarge amount of DAG (20 glday for threemonths) did nor affect the fat-soluble vit-amin StaTUS in humans (5).

4

I

0'-'---'-ConL High High

TAG DAG

Figu~ ). Effec.t$ of diet on body fa.ta.ccumuluion In CS7BU6J mice a.fter fiyemonths. (Cont .• stand:ard chow controldiu containln. S" TAG; HI&hTAG, TAGdiet containln& )0"TAG + I)" 5UCrGse;High OAG, OAG diet containing )0" OAG+ I)" SUCrG5e(n ". S each, mean t SO;WAT, white adipose tissue).

Volume 12 • November 200 I - inform

Anti-obesity effectsalso proven in animal studiesAnimal models have been used TO con-firm the effects of DAG and to elucidatethe mechanism of the effects.Substituting DAG for TAG preventedincreases in body far, insulin, and leptinconcentration associated with a high-fat and sucrose diet in obesity- anddiabetes-prone mice. Figure 3 shows thereduction of body fat (white adipose tis-sue, WAT) by feeding DAG for fivemonths to obesity- and diabetes-pronemodel mice (6). After five months of adlibitum feeding, body weight and fatwere significantly elevated with thehigh-TAG diet bur not with the high-DAG diet, as compared with controls.Insulin and leptin (peptide product ofthe OB gene, which is associated withobesity in mice as well as in humans) con-centrations were similar to those in rhecontrol group in the mice fed the high-DAG diet, bur significamly higher withthe high-TAG diet (Figure 4). Therefore,the mice consuming the DAG diet wereable to maintain smaller fat stores, sug-gesting that DAG consumption producedan increase in energy expenditure.

TRIACYLGI.YCEROl GMOUP

Change

68.1 :t J.J67.0:t 1.5'81.0:t 1.079.5:t 1.2"

56 :t 6.051 :t 6.0

126:t 101I8:t 13

-\.I :t 0.4

-1.5", 0.6

-5.0", 3.0

-8.0", 4.0

Events in the digestive tractand the liverDuring digestion, the major componentof common cooking oil, TAG, normallyis hydrolyzed to 2-monoacylglycerol(MAG) via I(or 3},2-DAG by the actionof pancreatic lipase and incorporatedinto mucosal cells. The digestion/absorption process of DAG has beeninvestigated in an experiment tracingtime-course changes in the lipid compo-sition ratio after perfusion of trioleinand diolein (1,2-:1,3-diolein = 3:7) inthe intestinal tract of rats (Figure 5). Inthe diolein-perfusion group, monooleinand oleic acid were produced as diolemwas degraded, and unlike the trioleinperfusion, 65% of the rnonoolein wasl-monoolein 60 minutes after initiationof perfusion. Thus, the production of1(3}-MAG may be a characteristic ofDAG metabolism.

Focusing on resynthesis of TAG inepithelial cells of the small intestinefurther clarifies the characteristics ofDAG metabolism. The TAG resynthe-sis pathway in the small intestinalepithelial cells include the 2-MAGpathway and the glycerophosphate

A. Serum Insulin Levels

1.2

'i 1.0

".s 0.8.s~ 0.6

0..

Cont. High HighTAG DAG

B. Serum Leptin Levels

•• ••

W 20

".s is.~

10:l5

0 eo.L High HighTAG DAG

Figu~e <4. Insulin ilnd leptln (oncent~iltlonsIn the mice fed DAG and TAG dietsfor five months.

pathway, with the former being con-sidered the mam pathway (7).Whereas 2-MAG is a good substrate ofthe reactions in the 2-MAG pathway,the reactivity of 1(3)-MAG is lower(8). Free glycerol is a substrate of theglycerophosphate pathway, but thcreaction rate is slower than that in the2-MAG pathway, and its contribution[0 TAG resynthesis is small (7).Although it has not been quantitative-ly concluded what ratio of 1(3)-monoolein produced from DAG isdirectly absorbed, the amount of 2-MAGproduced is lower after the ingestion ofDAG than after ingestion of TAG;thus, the resynthesis rate of TAG in thesmall intestinal epithelium may be

1101

TAG Perfusion DAG Perfusion100 tOO

~ ~

" "f.- 8() - 8()c c0 0":i .=•• 50 • 8()c ~

c• TO •• •c c0 40 <3 40U• •> .~"" 20 20.. •• ~" 0 00 2tl 40 60 0 2tl 40 60

Time of Perfusion (min.) Time of Perfusion (min.)

Figure S.Time-coune changes in the lipid composition notio afte~ perfusion of trio"!,inand diolein in the intestinal tract in nou .

Use ofDAGin pathological conditionsType II diabetes, abnormal lipid metab-olism, hypertension, hyperuricemia!gout, arteriosclerotic diseases, and Iartyliver nrc known as complications ofobesity. The importance of diet therapyas a means of improving these lifestylediseases is widely recognized. DAG oilhas been rested in pathological statessuch as dialysis patients, type II diabetespatients, and LPL (lipoprotein lipase}-deficient patients as well as obese chil-dren, and preliminary results have beenreported at scientific meetings.

In patients receiving hemodialysis,abnormal lipid metabolism, such ashigh TAG and low high-density lipopro-tein cholesterol levels, is commonlyobserved, and an association with ahigh incidence of heart diseases hasbeen reponed. When ten dialysispatients with hyperlipidemia ingestedDAG oil as cooking oil for threemonths, the amounts of visceral andsubcutaneous fat significantly decreasedcompared to amounts at the initiationof the study. After the STUdy, as thepatients rerumed to the ordinary cook-ing oil, the amount of intra-abdominalfat increased significantly, suggesting

Volume 12 • November 200 I • inform

lower with the ingestion of DAG. Thismay explain why the posrprandialIipi-dernia is less after DAG ingestion asshown in Figure 2.

In an experiment using rats, therelease rate of resynthesized TAG intothe intestinal lymph, was lower afteradministration of DAG emulsion thanthat of TAG emulsion (9). The concert-rrarion of free fatty acids in the mucosalcells may increase when the rate of TAGresynthesis is slowed after DAG admin-istration. This may explain the increasein free fatty acid levels in the portalblood after DAG administration com-pared to the levels after TAG adminis-tration.

It has been reported that when foodcontaining about 10% DAG was givento rats for 2-3 weeks, the enzymc activ-ity for fatty add synthesis in the liverwas decreased and that for ~-oxidationof fatty acids was increased (10).Hepatic acyl-coenzyme A oxidaseactivity and mRNA for acyl-coenzymeA synthase also were increased, sug-gesting a higher capacity for hepaticlipid oxidation. The characteristics ofthe digestion/absorption of DAGdescribed above may be related to thesefindings in the liver.

1102

that the DAG oil is effective inhemodialysis patients in reducing intra-abdominal fat.

In diabetic outpatients with hyper-nigtyceridcmia, continual ingestion(three months) of about 13 g of DAGoil daily decreased the serum TAG levelfrom 222 :I: 66 to 135 :I: 25 mgldL (/I ==

8, mean :I: SD, I' < 0.05). In contrast, inthe control group who ingested TAGoil, the serum TAG level changed from285:1: 189 to 318:1: 151mg/dL (/I == 8,mean :I: SD), showing no significant dif-ference. DAG oil also has been used inpatients with homozygous LPL defi-ciency showing hypenriglyceridemia,and has been shown to be effective incontrolling the serum TAG level.

DAG also has been used with obesechildren. In II simple obese children(7-17 years old), the daily use of cook-ing oil was replaced with DAG oil, andthe amount of body fat was measuredafter the children had ingested the oilfor five months. Ingestion of DAG oilreduced body fat without affecting thegrowth of the children.

Based on the above studies showingthat long-term ingestion of DAG oilreduces body fat accumulation in vari-ous pathological states in which lipidmetabolism occupies an imporranr posi-riun, expectations for DAG oil areincreasing as a food material not onlyfor healthy people but also in the diettherapy for these pathologies.

ConclusionSince the caloric value of the normalintake of DAG oil is practically thesame as TAG oil and the absorptionrate is also the same, the effect of DAGoil may be due to metabolic differencesin epithelial cells in the small intestine.Although a more detailed analysis ofthe digestion, absorption, and meta-bolic process is needed. ingestion of

Volume 12 - November 200 I • Inform

Biochemistry

DAG oil has been shown to improvethe lipid metabolism in various states.DAG oil has received GRAS statusfrom the U.S. Food and DrugAdministration. The authors hope thatDAG oil will contribute to the healthimprovement of consumers throughfurther applications of DAG oil in var-ious food products.

Readers wishing to contact the authorsmay reach Dr. Matsuo at BiologicalScience Laboratories, KaoCorporation. 2606 Akabane, Ic/}ikai-machi, Haga-gun, Tocbigi. 321-3497.japan (pholle; 81-285-68-7435;(ax; 81-285-68-7360;e-mail: 384895@kastaflct.kao.co.;p).

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Watanabe, K. Onizawa, N.Matsuo, L Tokirnitsu, and H.lrnkura, Energy Value andDigestibility of Dietary OilContaining Mainly 1,3-Diacylglycerol Are Similar to Thoseof Triacylglycerol, Lipids36:379-382 (200 IJ.

2. Tada, N., H. Watanabe, N.Matsuo, I. Tokimirsu, and M.Okazaki, Dynamics of PostprandialRemnant-like Lipoprotein Particles111 Serum After Loading ofDiacylglycerols, Clin, Cbim. Acta31/:109-117 (2001)

3. Taguchi, H.; H. Watanabe, K.Onizawa, T. Nagao, N. Core, T.Yasukawa, T. Tsushima, H.Shimasaki, and H. ltnkurn, DoubleBlind Conrrolled Study on theEffects of Dietary Diacylglycerol onPostprandial Serum andChylomicron TriacylglycerolResponses in Healthy Humans, l-Am. Coli. Nutr: 19:789-796 (2000).

4. Nagao, T., H. Watanabe, N. Gore,

K. Onizawa, H. Tcgucbi, N.Matsuo, T. Yasukawa, R.Tsushima, H. Shimasaki, and H.lmkura , Dietary DiacylglycerolSuppresses Accumulation of BodyFat Compared to Triacylglycerol inMen in a Double-Blind ControlledTrial,). Nutr: 130:792-797 (2000).

5. Watanabe, H., K. Dnizawa, S. Naito,H. Taguchi, N. Core, T. Nagao, N.Matsuo, I. Tokimitsu, T. Yasukawa,R. Tsushirna, H. Shimasaki, and H.lmkum, Fat-Soluble Vitamin Status IsNor Affected by DiacylglycerolConsumption, AmI. Nutr. Metab. (inpress).

6. Murase, T., T. Mizuno, T. Omachi,K. Onizawa, Y. Komine, H. Kondo,T. Hasc, and 1. Tokirnitsu, DietaryDiacylglycerol Suppresses High Fatand High Sucrose Diet-InducedBody Fat Accumulation III

C57BU6J Mice, l- Lipid Res.42:372-378 (2001).

7. Friedman, LH., and 8. Nylund,Intestinal Fat Digestion,Absorption, and Transport, Am. J.cu« Nutr: 33:1108-1139 (1980).

8. Lehner, R., A. Kuksis, and Y. lrabasbi,Stereospecificity of Monoacylgljceroland Diacylglycerol Acvlrransferesesfrom Rat Intestine as Determined byChira! Phase High-PerformanceLiquid Chromatography, Lipids28:29-34 (1993).

9. Murata, M., K. Ham, and T. Ide,Alteration by Diacylglycerols of theTransport and Fatty AcidComposition of Lymph Chylomicronin Rats, Biosci. Bioteclmo/. Biochem.58:1416---1419 (1994).

10. Murata, M., T. Ide, and K. Hara,Reciprocal Responses to DietaryDiacylglycerol of Hepatic Enzymesof Farry Acid Synthesis andOxidation in the Rat, Br. j. Nutr.77:107-121 (1997).0

I