Toxic. in Vitro Vol. 3, No. 2, pp. 85-90, 1989 0887-2333/89 $3.00+0.00 Printed in Great Britain. All rights reserved Copyright © 1989 Pergamon Press plc EFFECTS OF SOME PURIFIED SAPONINS ON TRANSMURAL POTENTIAL DIFFERENCE IN MAMMALIAN SMALL INTESTINE J. M. GEE, K. R. PRICE, C. L. RIDOUT, I. T. JOHNSONand G. R. FENWICK Agricultural and Food Research Council Institute of Food Research, Norwich Laboratory, Colney Lane, Norwich, Norfolk NR4 7UA, England (Received 26 May 1988; revisions received 28 September 1988) Abstract--The effect of a range of saponins, commonly present in foods or dietary supplements, on the potential difference (PD) across the mucosa of the rat small intestine/n vitro has been examined. Saponins from Gypsophila, guar, alfalfa, Quillaja, clover and liquorice together with glycoalkaloids from the potato and tomato were examined. The typical response was an immediate reduction in PD, although there was considerable variation in the response to particular compounds. Amongst the factors affecting the nature and magnitude of the de-polarizing effect were pH, solubility and the chemical form of the saponin. In agreement with the findings of others, glycyrrhizic acid, isolated from liquorice root, was found to exhibit a protective effect against the activity of a more potent saponin. The observations are discussed in the light of the known physiological activities of plant saponins and the regular, or excessive, consumption of certain foods or dietary supplements. INTRODUCTION Saponins are widespread in the plant kingdom, being found in more than one hundred different families (Bader and Hiller, 1987). Their occurrence in foods and animal feedstuffs has recently been reviewed (Price et al., 1987) and their biological activities have been considered. Although most saponins share common surface-active properties because of their lipophobic monosaccharide or oligosaccharide and lipophilic aglycone moieties, they include a great diversity of chemical structures that, in turn, dictate the biological and chemical properties of the com- pound (Adler and Hiller, 1985; Bader and Hiller, 1987; Hiller and Voigt, 1977). There have been many investigations which have purported to show the biological effects of saponins and which have been rightly criticized for their lack of information about the nature, origin or purity of the saponins used (Malinow, 1984; Price et al., 1987). In addition, claims as to the behaviour of individual saponins in in vivo and in vitro assays have been conflicting. We have described the permeabilizing effect of a number of isolated saponins, using an in vitro assay procedure (Johnson et al., 1986). This work demonstrated that the monodesmosidic soya- saponins (i.e. those possessing a single carbohydrate chain) had a weak--but detectable--permeabilizing effect on the rat intestine, although this was much less than those produced by either ~-tomatine or a mix- ture of Gypsophila saponins. In subsequent studies the effects of Gypsophila and soya saponins have been examined in rive (Gee and Johnson, 1988; Southon et al., 1987 and 1988), and the mean daily intake of saponins in the UK diet has been deter- mined (Ridout et al., 1968). Abbreviation: PD ffi potential difference. The present paper reports more detailed findings on the effects of a range of structurally-diverse saponins isolated from plants used as foods and dietary supplements. Factors affecting the observed gut permeabilizing behaviour of the saponins are examined, and the possible consequences of the presence of these compounds in the human diet are discussed. MATERIALS AND METHODS Materials The physiological studies were conducted using commercially available glycyrrhizin and ~t-tomatine (Sigma Chemical Co. Ltd, Poole, Dorset), and the glycoalkaloids at-solanine and ~t-chaconine, which were obtained from potato (Solanum tuberosum) sprouts as reported by Coxon et al. (1979). The isolation and purification of Gypsophila, alfalfa and clover saponins has been described by Southon et al. (1988) and Jurzysta et al. (1988). Crude Quillaja saponin was kindly supplied by Dr D. G. Oakenfull (Food Research Laboratory, CSIRO, Australia) and guar saponin was isolated by the method of Curl et al. (1986). Transmural potential difference measurements Male Wistar rats (200-250 g) were fed a commer- cial pelleted non-purified diet (RM3 Expanded, Special Diet Services Ltd, Witham, Essex) and water ad lib. prior to sacrifice by an ip injection of sodium barbiturate (Euthetal, May & Baker Ltd, Dagenham, Essex) and cervical dislocation. The manufacturer's specification indicated that the diet was unlikely to contain saponins. The entire small intestine of the rat was removed, rinsed with Krebs bicarbonate Ringer solution, and the proximal 40-cm portion was everted. Following removal of the proximal 10% portion of the small intestine, three 5-cm segments of 85 T.LV. 3/~-A