Effects of cooking using multi-ply cookware on absorption of potassium and vitamins: a randomized double-blind placebo control study

This 2-week interventional study involved a randomized allocation of subjects into three groups: Group A (daily ingestion of 350 g vegetables cooked without water using multi-ply [multilayer-structured] cookware), Group B (daily ingestion of 350g vegetables; ordinary cookware) and Group C (routine living). Before and after intervention, each subject underwent health examination with 24-h urine sampling. Blood vitamin C significantly increased after intervention from the baseline in Group A (P < 0.01) and Group B (P < 0.05). β-Carotene levels also increased significantly after intervention in Group A (P < 0.01) and Group B (P < 0.01). Oxidized low-density lipoprotein decreased significantly after intervention in Group A (P < 0.01). In Group A, 24-h urinary potassium excretion increased significantly (P < 0.01) and 24-h urinary sodium (Na)/K ratio improved significantly (P < 0.05) after intervention. In conclusion, a cooking method modification with multi-ply cookware improved absorption of nutrients from vegetables and enhanced effective utilization of the antioxidant potentials of vegetable nutrients.


Introduction
Ingestion of vegetables in the daily diet prevents lifestyle-related diseases (Hooper 2001). In Japan, the National Health Promotion Campaigns for the 21 st Century( HealthyJ apan 21), advocatedb yt he Ministryo fH ealth, Labour and We lfare (2000), recommendadultsingest 350 gormore vegetables per day for maintenance of good health. In practice, however, the amount of vegetables eaten daily by individuals aged between 20 and 39 yearsi sa pproximately 250 g, which is below the target level (Ministry of Health, Laboura nd We lfare 2009).T he goal of ingesting 350 go rm ore of vegetables per day can be achievedi fv egetables eaten are cooked (rather than raw) andi ft he amountr equired daily is divided into threem eals. However, individuals, particularly students, increasingly skip breakfast or relyonfast food as one of their three meals (Osakoetal. 2005).For these individuals, daily ingestion of 350 gv egetables is difficult to achieve, ands ome investigatorsr eported that young mena nd women consumed an average of 130 go fv egetables per day.A dditionally, the use of water to cook vegetables shouldb ea voideda sf ar as possible so that the nutrients, which are likely to be elutedinto water, are preserved (Kimura and Itokawa 1990;Agte et al. 2002;Gupta and Bains 2006).T he purpose of the presents tudy was to examine whether the absorption of vegetable nutrients is higher when vegetables are cooked using traditional methods or using modified cookware to preserve water-solubleo r heat-sensitive nutrients. We asked the randomized volunteerst ou se modified cookware thate nabled more efficient nutrient intaker athert han ingesting vegetables cooked by ordinarycookware, and analyzed changes in the levels of 24-hour urine sample (24 U) K excretion, blood vitamins and oxidized low-density lipoproteincholesterol (ox-LDL), among othervalues.

Subjects
Of the undergraduate and graduate students of Kyoto University,those living alone and capable of cooking at homew ere invited to an orientation meeting during which the study design and details were explained. The criteria of exclusion were (1) the student whowas notl ivinga lone,( 2) thes tudent whod isliked vegetables and(3) the student living in aroom without ak itchen. And 90 students understood the study design and details and provided informed consent after the orientation, and were enrolled in the study. Alls ubjects underwent healthe xamination before the startofthe study,and subjects were randomly divided intot hreeg roups: Group A, daily ingestiono f3 50 g vegetables cooked without water using a' multi-ply cookware' (fully multilayer-structuredc ookware #5123,Vita Craft Japan Ltd, Kobe, Japan); Group B, daily ingestion of 350 gv egetables cookedu sing an ordinarycookware and Group C, ordinarydaily living. This study was performedu nder ethical considerations in compliance with 'The Declarationo f Helsinki -E thicalP rinciples for Medical Research Involving Human Subjects' and 'The Ethical Guidelines on Epidemiological Studies'( Ministry of Education,C ulture, Sports, Science, andT echnology andMinistryofHealth, Labour and We lfare). During the study,wetookadequate care of the life, health and privacy of individualc ontributors,s taff membersa nd subjects. Informed consent was obtained from each subject before the starto ft he study.T his study was approved in advance by the Ethics Committee of the KyotoU niversityG raduate Schoolo fH umana nd Environmental Studies and the Ethics Committee of Mukogawa Wo men'sU niversity.

Schedule
Of the subjects randomly allocated to the three groups, those allocated to Groups Aa nd B( groups to which cookware was provided) received cookingt raining (separate training for each of these two groups) so that thes ubjectsw ould uset he providedc ookware appropriately.N ext, the apparently similar cookware (one unit) was provided to individuals ubjects in Groups Aa nd B. The cookware and first seto f vegetables were supplied simultaneously,a nd the subjects began to consume the supplied vegetables on the day of allocation. To retain freshness of the supplied vegetables, vegetables were supplied twice per week (six supplies during the study period). Amounts of unusedvegetables anddiscarded amounts were recorded; full ingestion of supplied vegetables was preferred. Health examinations, identical to those performedb eforet he starto ft he study, were performed2weeksa fter thes tart of vegetable consumption( Figure 1).

Te st diet
To ensure that au niforma mounto fv egetables was ingested daily by eachs ubject, consistent amounts of vegetables were supplied at six time points during the study period. Ta ble Il ists the vegetables supplied during the study.

Measurement at the time of health examination and during the study period
Health examinationb efore anda fter thes tudy included the measurement of height, body weight, body fatr atio, blood pressure, heartr ate (using an automatedb lood pressurem easurement system; HEM-907 Omron,K yoto, Japan),f asting blood sampling and by 24 Uc ollectionu sing an aliquot cup.T he photography of meals before dieting and diarytracking (to record supplied vegetableconsumption) were collected during health examination before Baseline Group A: Eating 350g of vegetables per dayf or using multi-ply utensils by waterless cooking , aspartate aminotransferase, alaninea minotransferase, g -glutamyl transpeptidase, vitamin C, b -carotene and insulin. The levels of Na, K, creatinine and 8-hydroxydeoxyguanosin were measured in the 24 Us ample. Photographs taken before taking meals were used to assess the amount of vegetables consumed before and after the starto ft he study.P hotographs of all vegetables consumedo vera 3-day period before the startofthe study and for 3days before the completion of the vegetable consumption period (6 days in total) were acquired using am obile phone camera by individual subjects. Photographs takenw ere saved on ar ecording medium for mobile phones (microSD) supplied by the researchersa nd collected at the time of the last health examination. Each subjectm aintainedadiary to record the vegetables consumed relative to those supplied, the vegetables discarded without being consumed, physical conditiona nd supplement intake, as well as other relevant factors, everyd ay during the study period.

Adverse events
One subject withdrew from participation in the study after the pre-studyh ealthe xamination and before randomizeda llocation forapersonalr easonn ot related to thes tudy design. One subject from Group Aq uit the study because of fever onset on the day of post-study healthexamination. One subject quit thes tudyo nt he dayo ft he post-study health examination for ap ersonal reason not related to the study design.
Allo ft hese events were judgedt oh ave no association with the test diet according to the principal investigator andt he physician responsiblea tt he testing facility.

Statistical analysis
At hirdp arty who was not parto ft he study staff evaluated the amounts of vegetables consumedb y individual subjects using the photographs of meals takeno ver a6-day period before and after the startof the study.A nalysis included5 7s ubjects, and subjects eachi nG roups Aa nd Bt ow hom vegetables were supplied increased vegetable consumption and subjects in Group Cd id not increase the amounto f vegetables ingested.Analysis of the data obtained from 24 Uanalysis included45subjects (16 from Group A, 15 from Group Ba nd 14 from Group C) in whom 24 Uc ollectionw as objectively rated as successful on the basis of self-reportsa nd urinaryc reatinine levels ( Figure 2). The health examination data before and after the starto ft he study in these subjects were evaluated using the t -test. The magnitude of change after the starto ft he study was compareda mong the threeg roups using analysis of variance. Statistical analysis was performedu sing SPSS Ve r. 15.0. P , 0.05 was regarded as statistically significant.

Magnitude of change between the vitaminlevels before and after vegetable intake
The magnitude of change in vitamin Cl evels before anda fter vegetable intake in Groups Aw as significantly different ( P , 0.05)f romG roup C. The magnitude of changei n b -carotene level was significantly different between Groups Aa nd C( P , 0.05) and between Groups Ba nd C(P , 0.01; Ta bleI II).

Results of 24 Usample test
The amountofKexcreted daily into urine (calculated as K) increased significantly after the study compared with thel evelsb eforet he study in GroupA (1.6^0.9 ! 2.0^1.1 g; P , 0.01), while it showed no statistically significant changes in Group Bo r C. The Na/Kratio decreased significantly in Group A (3.7^2.0 ! 2.6^0.9; P , 0.05), while no statistically significant changes were observed in Group Bor C( Ta ble IV).

Discussion
The presents tudy was designed to evaluate the differential effecto nt he absorptiono fv egetable nutrients by using two cookware and to objectively analyze the differenceb ye xamining blood and 24 U biomarkerso fn utrients. Cooking using multi-ply cookware, designated as Group A, was characterized as water-free cooking, includingb oiling vegetables using only the moisture contained in the vegetables. Ordinaryheating or boiling of vegetables using water is likelyt or esult in the elution of ah igh percentage of vitamins andm inerals in the vegetables (Kimura and Itokawa 1990) andt hese nutrients are decreased in ready-to-eat foods ( Agte et al. 2002) comparedw ith traditional foods ( Gupta and Bains 2006),w hile water-free cooking minimizesthe elution of nutrients, indicating that increased Kintake and decreasedNa/K ratio concomitantly observedp articularly in Group A maycontribute to the possible risk reduction of stroke. Moreover, serum total and LDL cholesterol observed in Groups Aa nd Bt akingm ore vegetables confirmed the well-known benefite ffect of vegetable intake (Djoussé et al. 2004) andt he related risk reduction of coronaryheartdiseases (Yamorietal. 2006).Blood vitaminCand b -carotenel evels also increased significantly afteri ntervention in GroupA , accompanied by as ignificant reduction in blood ox-LDLl evel. There is ar eportw hich clearly demonstrated that vitamin Cprevented increased endothelial permeability caused by ox-LDL (May andQ u2 010). Increased endothelialp ermeabilityr esults in the development of atherosclerosis vascular lesions (Fujita et al.2 009). Therefore,v itamin Ce levation and concomitant ox-LDL reduction observedi nG roup A indicate that the ingestion of vegetables cooked by the modified cookware is expected to reduce the risk of cardiovascular diseases and major outcomeo f atherosclerotic vascular lesionsint he long run.

Conclusions
This randomizedd ouble-blind study comparing the intake of similar amounts of provided vegetables cooked with water-freee fficient cookware with the intake of vegetables cooked with ordinaryc ookware (placebo) has confirmed that using this water-free cooking method enables more efficient utilization of thea ntioxidant potential of vegetables,w hich is expected to haveb ettere ffects on cardiovascular risk reduction than ingesting the vegetables cookedb y ordinarym ethods.