Factors other than phytosterols in some vegetable oils affect the survival of SHRSP rats

Abstract

Unusual survival-shortening activities of some vegetable oils were detected in stroke-prone spontaneously hypertensive (SHRSP) rats, and phytosterol (PS) in the oils and the tissue tocopherol status have been suggested to be the factors for the activities. Here, we re-evaluated the contribution of PS to the survival-shortening, and examined the hepatic tocopherol status. A basal diet for rodents and a test oil were mixed at a 9:1 ratio, and the diet was given to male SHRSP rats upon weaning. The total and major PS contents of the diets and tissue lipids did not correlate with relative survival time. The free fatty acid fractions obtained by lipase and alkaline hydrolyses of canola oil (Can) and the original Can contained PS in comparable amounts but the free fatty acid fractions did not exhibit survival-shortening activities compared with the soybean oil (Soy) group. The activity was not detected in the ethyl acetate extracts of the aqueous phase after the hydrolysis. When a commercially available PS preparation was added to the Soy diet at an amount 2.8-fold higher than that in the Can diet, the mean survival time was shortened but was still significantly longer than that of the Can group. The hepatic tocopherol level was significantly higher in the Can group than in the hydrogenated Soy group and Soy group, but the former two groups exhibited a survival-shortening activity. These results indicate that factors other than PS, tocopherol status and fatty acid composition in some vegetable oils are critical for the survival-shortening activity observed in SHRSP rats.

Introduction

Oil from traditional rapeseed variety contains two anti-nutritional factors, erucic acid and thyrotoxic sulfur compounds from glucosinolates. In 1974, a new rapeseed cultivar that is low in both erucic acid and glucosinolate contents was released (Stefansson and Kondra, 1975), which is now available as canola. Even with the use of rapeseed oil of the low-erucic type as dietary supplement, myocardial lesions were observed in rats (McCutcheon et al., 1976). However, a triacylglycerol fraction, highly purified from low-erucic rapeseed oil by repeated molecular distillation, resulted in similar cardiac lesions even though the preparation lacked phytosterol (PS) (Kramer and Sauer, 1983). Based on the results of extensive studies, the observed myocardial lesions in male rats (Sprague-Dawley, Wistar and Sherman) were ascribed to triacylglycerols and their particular fatty acid composition (Kramer and Sauer, 1983).

Perilla seed oil, flaxseed oil and fish oil with very low n−6/n−3 ratios extended the survival of the SHRSP rats, an animal model of human-cerebral hemorrhages, by approximately 10%, and exhibited other beneficial effects compared with high-linoleic oils (Shimokawa et al., 1988; Okuyama et al., 1996; Miyazaki et al., 2000; Minami et al., 1997), indicating that dietary n−3 fatty acids are beneficial to SHRSP rats. Unexpectedly, canola oil (Can) with a relatively low n−6/n−3 ratio (∼2.5) shortened the survival time of SHRSP rats compared with the Soy diet (Huang et al., 1996). Some other oils such as high-erucic rapeseed oil, olive oil, evening primrose oil, high-oleic safflower oil, high-oleic sunflower oil, corn oil, hydrogenated Can and hydrogenated Soy (Hyd.Soy) also exhibited comparable survival-shortening activities (Huang et al., 1996, Huang et al., 1997; Miyazaki et al., 1998a, Miyazaki et al., 1998b; Ratnayake et al., 2000a, Ratnayake et al., 2000b). However, the free fatty acid fractions obtained by the lipase hydrolysis of Can, Hyd.Soy and high-oleic safflower oil did not exhibit such an activity, although the fatty acid compositions, tissue fatty acid compositions and growth rates were very similar between the groups fed these oils and derived free fatty acid fractions. These observations suggested that minor components other than fatty acids are responsible for the survival-shortening activity (Miyazaki et al., 1998a, Miyazaki et al., 1998b). In neonatal piglets raised on a Can-based milk replacer diet, tissue tocopherol levels were lower and signs of vitamin E deficiency were severer than those in piglets raised on a Soy-based milk replacer diet (Sauer et al., 1997). However, the factors in Can that increased the demand for vitamin E have not yet been determined.

Recently, the SHRSP rat strain as well as its parent WKY strain has been reported to exhibit severe phytosterolemia in comparison with other rat strains (Wistar, WKA) (Ikeda et al., 2001), the condition is associated with a mutation in an ATP-binding cassette (ABC) transporter with a presumed activity of excreting PS into the intestine (Scoggan et al., 2003). By comparing some vegetable oils, phytosterol (PS) was proposed to be involved, at least in part, in the survival-shortening activity, although olive oil was exceptional in that its PS content was the lowest but resulted in the shortest survival time among the oils examined (Ratnayake et al., 2000a). A mixture of PS, extracted from canola deodorant distillates and re-crystalized, was shown to exhibit a survival-shortening activity (Ratnayake et al., 2000a). If PS is the major factor affecting survival, the impact of the observed survival-shortening activity of some vegetable oils on human nutrition may be considered relatively small because the effects may be confined to particular animal strains, and PS is generally recognized as a beneficial nutrient for its ability to suppress cholesterol absorption in the intestine. However, our data (Huang et al., 1996, Huang et al., 1997; Miyazaki et al., 1998b) were not consistent with this interpretation; hence, we re-evaluated the correlation between survival time and PS content in dietary and tissue lipids, attempted to extract the presumed factors from the aqueous phase after the lipase and alkaline hydrolyses of Can by modified methods, and evaluated hepatic tocopherol status in relation to the reported vitamin E deficiency in piglets (Sauer et al., 1997).