6
Downloaded by UNIV OF CALIFORNIA SAN DIEGO on February 1, 2016 | http://pubs.acs.org Publication Date: January 16, 1986 | doi: 10.1021/bk-1986-0296.ch006
Polyhydroxy Plant Alkaloids as Glycosidase Inhibitors and Their Possible Ecological Role L. E. Fellows, S. V. Evans, R. J. Nash, and E. A. Bell Jodrell Laboratory, Royal Botanic Gardens, Kew, TW9 3DS, United Kingdom
Polyhydroxyalkaloids which structurally resemble monosaccharides have been found in a variety of organisms, including higher plants. Many are potent inhibitors of glycosidase activity in insects, mammals and micro-organisms and it is suggested that these properties contribute to the natural chemical defences of those plants in which they accumulate. In recent years, representatives o f 3 new classes o f a l k a l o i d have been i s o l a t e d from both plants and micro-organisms, namely polyhydroxy derivatives o f i n d o l i z i d i n e , p y r r o l i d i n e and p i p e r i d i n e . Their r e l a t i v e l y l a t e discovery i s probably due to t h e i r f a i l u r e to react with most colorimetric reagents used to detect a l k a l o i d s , such as Dragendorff reagent, but most do react with ninhydrin to give a yellow o r brown colour. Evidence i s growing that these compounds are widely d i s t r i b u t e d i n plants, representatives having now been i s o l a t e d from four separate families, v i z . Koraceae ( l ) Leguminosae (2-8), Polygonaceae (£) and Aspidiaceae (10). In some legumes we have observed that these a l k a l o i d s may accumulate as 2# or more o f the plant's dry weight (11) which suggests that they have an ecological r o l e . Many other types o f a l k a l o i d are o f course known to be e f f e c t i v e deterrents to prédat i o n , p a r t i c u l a r l y by mammals. The unhydroxylated piperidine alkal o i d s o f Hemlock (Conium maculatum) poison the mammalian nervous system, as Socrates well knew, but when a polyhydroxy p i p e r i d i n e (Figure 1,) was i s o l a t e d a t Kew from the legume Lonchocarpus sericeus (2), no a c t i v i t y o f t h i s nature was detected. 1^ i s an isomer o f t
2-hydxOxymethyl-3,4,3-trihyo^xypipendine,
which can a l s o
be
as 1,5-dideoxy-l,5-imino-D-mannitol; i t i s i n f a c t an analogue o f 1 - d e o x y m a n n o s e i n which the r i n g oxygen i s replaced b y nitrogen. This suggested that 1^ might mimic the action o f mannose and thereby i n h i b i t carbohydrase, p a r t i c u l a r l y mannosidase, a c t i v i t y . Ve were unaware a t t h i s time that Japanese workers had i s o l a t e d from c e r t a i n mulberry (Morus) spp. a compound (Figure 2) being an epimer of 1^ at C5, which they named moranoline ( l ) . I t was l a t e r also found i n b a c t e r i a (12-13) and described as deoxynojirimycin, the name which now p r e v a i l s . This compound, 2_ i s an analogue o f described
%
0097-6156/ 86/ 0296-0072506.00/ 0 © 1986 American Chemical Society
In Natural Resistance of Plants to Pests; Green, Maurice B., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
Downloaded by UNIV OF CALIFORNIA SAN DIEGO on February 1, 2016 | http://pubs.acs.org Publication Date: January 16, 1986 | doi: 10.1021/bk-1986-0296.ch006
6.
FELLOWS ET AL.
Alkaloids
as Glycosidase
inhibitors
73
1-deoxyglucose, being an epimer of 1^ at C2 i f each molecule i s con sidered as a monosaccharide, and i n h i b i t s both a - and Β -glucosidases from varied sources, i n c l u d i n g fungal trehalose (12-1$). I t also i n h i b i t s the postprandial r i s e i n blood glucose i f administered to r a t s with food and a patent was f i l e d on 2 as an a n t i d i a b e t i c agent (16). Recently i t has been shown to be a potent i n h i b i t o r o f mouse gut digestive disaccharide a c t i v i t y , p a r t i c u l a r l y with substrates incorporating an α -linked glucopyranosyl residue (17)· The mannose analogue !L i n h i b i t s α -mannosidase from various sources, and also bovine epididymis α -L-fucosidase 0 Λ , 18-19)* No glucosidase i n h i b i t i o n has been reported. An a l k a l o i d c l o s e l y related to both !1 and 2, but l a c k i n g a hydroxy group at C5 (G2 i f considered as a sugar) was detected i n buckwheat (Fagopyrum eseulentua) and named fagomine (Figure £ ) , (£). Recently we have i s o l a t e d a glucoside o f fagomine, *f-0-(&-D-gluco pyranosyl)-fagomine, (£) from the legume Xanthocercis sambesiaca. Neither fagomine nor i t s glucoside showed any i n h i b i t i o n of a range of glucosidaees tested which confirms previous findings of the impor tance o f the hydroxy group at C2 (piperidine C5) i n c a t a l y s i s ( 2 0 ) . It was r e a l i s e d that a polyhydroxy p y r r o l i d i n e , DMDP (Figure 4 ) , i s o l a t e d from D e r r i s e l l i p t i c a ( 2 1 ) , was an analogue o f 0-D-fructofuranose, and at Kew we were able to i s o l a t e k from r e l a t e d legumes (Lonchocarpus spp.) i n s u f f i c i e n t quantity to carry out a range o f b i o l o g i c a l t e s t s . ( I t has also been synthesised and i t s absolute configuration confirmed as 2R-,5R-dihydroxymethyl-3R,^R-dihydroxyp y r r o l i d i n e ) ( 2 2 ) . DMDP also proved a potent glucosidase i n h i b i t o r , but i t s a c t i o n d i f f e r s from that o f 2 i n several respects: f o r example, while k i n h i b i t s yeast invertase (50* i n h i b i t i o n at 5 · 2 5 χ 10~^M), 2 has no e f f e c t at 5 x 10"^M. (1*0. Conversely, 2 i s a more e f f e c t i v e i n h i b i t o r than 4^ of mouse gut invertase (50# i n h i b i t i o n at 7.6 10~°M and 4.6 χ 10"5M respectively) ( 1 7 ) . I n h i b i t i o n of l y s o somal β-D-mannosidase and o f a mannosidase o f i n t e s t i n a l e p i t h e l i a l glycoprotein processing by k has also been reported (23-2*0. DMDP also i n h i b i t s insect trehalase (iff). Since trehalose, 1-i* -D-glucopyranosyl-a-D-glucopyranoside) i s an important storage carbohydrate i n i n s e c t s but not i n mammals, i n h i b i t i o n o f insect trehalase could t h e o r e t i c a l l y provide the basis of a r e l a t i v e l y safe i n s e c t i c i d e . At Kew we have been p a r t i c u l a r l y i n t e r e s t e d i n the chemical basis o f the i n t e r a c t i o n between bruchid beetles and t h e i r legume hosts. Bruchid beetles normally l a y eggs on green pods; the larvae penetrate pod and testa, develop i n the seed and emerge as adults. Crop g r a i n legumes have few natural chemical defences o f t h e i r own and are potential prey for many bruchid spp. but, provided the bruchid r e t a i n s i t s requirement for a green pod f o r o v i p o s i t i o n , a low l e v e l o f contamination at harvest i s no problem since emerging adults do not multiply. This requirement has been l o s t , however, by c e r t a i n spp. which can breed i n storage and are now major economic pests ( 2 5 ) . Wild legumes are p o t e n t i a l sources of anti-bruchid chemicals which might be used for crop protection. When k was incorporated i n t o a r t i f i c i a l 'seeds' made o f cow pea f l o u r packed i n gelatine capsules, l e v e l s o f only 0 . 0 3 * were l e t h a l to larvae o f the pest bruchid Callosobrusus maculatus ( 2 6 ) . (Fagomine and i t s gluco side had no e f f e c t , even at 1%). In contrast, mice fed a diet con t a i n i n g % k f o r one week showed no i l l e f f e c t s other than a s l i g h t
In Natural Resistance of Plants to Pests; Green, Maurice B., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
74
NATURAL RESISTANCE OF PLANTS TO PESTS
N^CHoOH Downloaded by UNIV OF CALIFORNIA SAN DIEGO on February 1, 2016 | http://pubs.acs.org Publication Date: January 16, 1986 | doi: 10.1021/bk-1986-0296.ch006
Π
Figure 1.
Deoxymanno j i r i m y c i n
Figure 2.
Deoxynojirimycin
OH
Η Figure 3· Fagomine
HQ
OH
HOH cA A»CH,QH 2
N
Η
Figure k.
DMDP
In Natural Resistance of Plants to Pests; Green, Maurice B., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
Downloaded by UNIV OF CALIFORNIA SAN DIEGO on February 1, 2016 | http://pubs.acs.org Publication Date: January 16, 1986 | doi: 10.1021/bk-1986-0296.ch006
6.
FELLOWS ET AL.
Alkaloids
as Glycosidase
Inhibitors
75
weight l o s s and recovered completely when returned to a normal diet (27). α -Glucosidase a c t i v i t y i n C. maculatus l a r v a l gut was strongly i n h i b i t e d by k (Kj 3.6 χ 10"/) (2oT"but that of mouse gut, (e.g. K j for maltase cT 10-^M) far l e s s so (17). The biochemical mechanisms by which insects select food plants are not f u l l y understood. I t has been suggested that an insect glucosidase may be involved which, when i n contact with the plant, releases free sugars which t r i g g e r the feeding response (28). A glucosidase i n h i b i t o r might therefore be expected to be an e f f e c t i v e feeding deterrent. Blaney et a l . (29) presented 5th i n s t a r nymphs of the a c r i d i d s Schistocerca gregaria (polyphagous) and Locusta migratoria (oligophagous) with glass f i b r e discs impregnated with known phagostimulants for l o c u s t s . On discs containing 5* sucrose, for example, DMDP l e v e l s as low as 0.001* were s u f f i c i e n t to reduce feeding, yet when locusts were force-fed (lmg/g body weight) there was no d i s c e r n i b l e e f f e c t . Higher concentrations of DMDP were required to deter feeding i n f i n a l i n s t a r larvae o f two Spodoptera spp. (armyworms), the oligophagous S. exempta being more e a s i l y deterred than the polyphagous j5. l i t t o r a l i s . Both species had high mortality when presented with sucrose-laden discs containing 1* DMDP; discs with 0.01* DMDP caused mortality only i n j>. l i t t o r a l i s owing to i t s having consumed more DMDP-laden food. Results o f electrophysiol o g i c a l studies on the Spodoptera gustatory s e n s i l l a could be i n t e r preted to indicate that DMDP was competing with a glucosidase recep tor s i t e , p a r t i c u l a r l y i n S. exempta, but the neurone f i r i n g i n a dose-dependent manner responded s i m i l a r l y to a l k a l o i d s . The authors, i n conclusion, favoured the view that the deterrency of DMDP derived more from i t s properties as an a l k a l o i d than as a sugar analogue. (Similar studies with 2 suggest that i t i s far l e s s e f f e c t i v e as a feeding deterrent A compound related to ? 7 being an isomer of 2-hydroxymethyl-3i4dihydroxypyrrolidine having the D-arabinose configuration, (alterna t i v e l y l,4-dideoxy-l,4-imino-D-arabinitol), (Figure £ ) , has been i s o l a t e d from the legume Angylocalyx boutiqueanus and found to be a more potent i n h i b i t o r of yeast α -glucosidase than either 2 o r 4 (6,8). (An isomer purported to have the D-xylose configuration has been found i n the fern Arachnoïdes s t a n d i s h i i but other authors claim that i t i e i d e n t i c a l with 5) (10, 3 D . The related (2R, 3S)-2-hydroxynaethyl-3-hydroxypyrrolidine (Figure
