Lipophilic Xenobiotic Conjugates - American Chemical Society

9 Lipophilic Xenobiotic Conjugates 1

2

Gary B. Quistad and David H. Hutson 1

Biochemistry Department, Zoecon Corporation, Palo Alto, CA 94304 Department of Biochemistry, Shell Research Ltd., Sittingbourne, Kent ME9 8AG, England

Downloaded by UNIV OF ARIZONA on July 1, 2014 | http://pubs.acs.org Publication Date: January 24, 1986 | doi: 10.1021/bk-1986-0299.ch009

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Although foreign compounds generally are rendered more hydrophilic by the vast majority of metabolic conjugations, occasionally xenobiotics enter path ways of lipid biosynthesis and thereby are con verted to more lipophilic derivatives. Specifi cally, the xenobiotic participates in normal routes of fatty acid metabolism. Foreign acids may mimic natural fatty acids (directly or after chain -extension to homologous acids), thereby incorporat ing into glycerides or forming esters of choles terol. Xenobiotics themselves may be esterified with natural fatty acids. This report summarizes the occurrence of this recently discovered class of nonpolar conjugates in animals and plants. H i s t o r i c a l l y , metabolic conjugations have been regarded as an organism'8 attempt to render an intruding chemical innocuous by increasing i t s polarity, thus f a c i l i t a t i n g excretion. While this maxim i s s t i l l generally correct, Important exceptions have been described in the last decade. There are now several examples of polar metabolites which are toxicologically s i g n i f i c a n t . For example, within certain chemical classes, glucuronidation, s u l f a tion, and glutathione conjugation have each been implicated i n the formation of reactive metabolites which i n i t i a t e carcinogenesis. The generalization that conjugates are more hydrophilic than their precursors must also be examined i n the l i g h t of the discovery of an increasing number of xenobiotics forming derivatives of natural l i p i d s . These xenobiotic l i p i d s , formed from both acids, amines, and alcohols, are certainly conjugates by virture of their mechanisms of formation, but their physical properties distinguish them from the c l a s s i c a l (hydrophilic) conjugates. This review emphasizes recent examples of novel l i p o p h i l i c conjugates. Previously reviewed nonpolar conjugates (1-3) are considered only b r i e f l y and although certain r e l a t i v e l y simple conjugations (e.g. methylation, formylation, and acetylation) also 0097^6156/86/0299-0204$06.00/0 © 1986 American Chemical Society

In Xenobiotic Conjugation Chemistry; Paulson, G., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1986.

9.

QUISTAD AND HUTSON

Lipophilic Xenobiotic Conjugates

205

produce more l i p o p h i l i c metabolites, these reactions have been known for many years and are not reviewed here. Chain Extension of Acids The f i r s t example of chain extension of a xenobiotic involved f u r f u r a l which was reported i n 1887 to add a 2-carbon unit thus producing 3-(2-fury1)acrylie acid which was isolated i n rabbit urine as i t s glycine conjugate (4). Subsequent workers were not able to confirm this observation (see 3_ f ° leading references). The hypotensive drug, 5-(4-chlorobutyl)picolinic acid, i s converted by rats into four metabolites which each contain the net addition of one C 2 unit (5, Figure 1). The i s o l a t i o n of the 0-keto acid and a, 3-unsaturated acid metabolites implies that a l k y l substituted p i c o l i n i c acids are a l t e r n a t i v e substrates i n the enzymatic chain elongation of natural fatty acids. S i m i l a r l y , horses excrete benzoic acid i n urine as chain-extended metabolites (6, Figure 1). These 3-keto and ^-hydroxy acids also could be viewed as intermediates i n a pathway toward the f u l l y saturated extended-chain metabolite. Thus, while 3-hydroxy, 3-keto, and a, $unsaturated intermediates are usually not abundant species i n natural fatty acid chain elongation ( i . e . the f u l l y saturated product predominates), these intermediary metabolites may be quantitatively important for xenobiotic acids. Because of i t s structural s i m i l a r i t y to acetate, i t i s not surprising that cyclopropanecarboxylic acid (from the miticide eye lop rate) i s incorporated readily into normal fatty acid path ways. Six homologous u>-cy clop ropy 1 fatty acids were i d e n t i f i e d from treatment of rats, dogs, and a cow with eye lop rate (7-10). Generally, these u>-cyc lop ropy 1 fatty acids were not free, but rather were e s t e r i f l e d to produce modified hybrid l i p i d s (vide inf ra). There are few examples of chain elongation of xenobiotic acids by plants. The herbicides 2,4-DB [4-(2,4-dichlorophenoxy)butanoic acid] and 2,4-D (2,4-dichlorophenoxyacetic acid) are both converted by a l f a l f a to 6-(2,4-dichlorophenoxy)hexanoic acid (11,12). Fluoroacetate i s converted by plants (13) and rats (l4) into longchain a>-fluoro fatty acids. In plant seeds 18-fluorooctadecenoic acid was the main constituent from fluoroacetate. Unsaturated long-chain to- eye lop ropy 1 fatty acids also have been isolated from apples and oranges treated with eyeloprate (15, Figure 1).


r

Glycerides Several xenobiotic acids are known to form hybrid t r i a c y l g l y c e r o l s ( i . e . a xenobiotic acid e s t e r i f l e d with a natural d i a c y l g l y c e r o l ) and adequate reviews of these conjugates have appeared elsewhere (1-3). Such hybrid t r i a c y l g l y c e r o l s have been reported for 3phenoxybenzoic acid (rats; 16) and a c i d i c metabolites of cycloprate (rat, cow, dog; 8-10), methoprene (chicken; 17), dodecylcyclohexane (rat; 18), and ethyl 4-benzyloxybenzoate (rat; 19). After discovering the formation of hybrid t r i a c y l g l y c e r o l s from 4benzyloxybenzoate, Fears et a l . (19) used an i n v i t r o system containing rat l i v e r s l i c e s to probe the generality of this pathway. When f i f t e e n aromatic acids were screened, four acids


XENOBIOTIC CONJUGATION CHEMISTRY


206

were incorporated into f a t more e f f i c i e n t l y than was 4benzyloxybenzoate while most of the remaining acids (including n i c o t i n i c and a c e t y l s a l i c y l i e ) were incorporated poorly or not at a l l . Pharmacologically important acids such as the antiinflamma tory drugs fenoprofen and ibuprofen formed novel CoA thioesters and were incorporated into glycerolipids by substituting for natural fatty acids. Reports of the formation of hybrid diacylglycerols ( i . e . glyc erol e s t e r i f i e d with a xenobiotic acid plus one natural fatty acid) are rare. When chickens were dosed with the insect growth regu lator raethoprene, 4% of the radiolabel i n egg yolks was i d e n t i f i e d as a mixture of two hybrid diacylglycerols (17). When rats were dosed at 1 mg/kg with the pyrethrold insecticide fluvalinate i n corn o i l (20), 2% of the radiolabel i n the fecal extract (0-1 day) was i d e n t i f i e d as hybrid diacylglycerols resulting from e s t e r i f i c a tion of both oleoylglycerol (Figure 2) and l i n o l e o y l g l y c e r o l by an a n i l i n o acid metabolite. Hybrid 1,2-diacylglycerols were more abundant than the corresponding 1,3-adducts. Interestingly, o l e i c and l i n o l e i c acids are the primary fatty acid constituents of the l i p i d s in corn o i l (the dose vehicle). A reduction of l i p i d i n the dosing medium resulted in the disappearance of these diglycerides, demonstrating the e f f e c t of diet on their formation. The a n i l i n o acid from fluvalinate forms the only reported conjugate between a xenobiotic and glycerol (20; Figure 2). This monoacylglycerol represented 2% of the radiolabel in the 0-1 day f e c a l extract from rats dosed with fluvalinate i n com o i l . Curiously, although actively sought, hybrid t r i a c y l g l y c e r o l s could not be i d e n t i f i e d in the above studies with fluvalinate in rats. Cholesterol Esters This class of l i p i d conjugates i s the most nonpolar yet i d e n t i f i e d , a c h a r a c t e r i s t i c which is often useful i n pursuing the i d e n t i f i c a tion of unknown metabolites. The f i r s t cholesterol ester of a xenobiotic was reported i n 1976 for a saturated methoprene metabo l i t e which contributed 15% of the total ^ - r e s i d u e i n the l i v e r of a chicken given a single oral dose of methoprene at 64 mg/kg (17). The a)-eye lop ropy 1 fatty acids derived from cycloprate also form esters of cholesterol. Three such esters contributed 5% of the t o t a l residual radiocarbon i n rat carcasses four days a f t e r a single oral dose of cycloprate at 21 mg/kg (8). A cholesterol ester forms in the l i v e r of rats given an oral dose (250 mg/kg for seven days) of the hypolipidemic drug l - ( 4 carboxyphenoxy)-10-(4-chlorophenoxy)decane (21). This xenobiotic cholesterol ester represented about 11% of the total l i p i d i n the l i v e r and was neither further metabolized nor transported by l i p o proteins. An additional 1% of the total l i v e r l i p i d s consisted of hybrid t r i a c y l g l y c e r o l s containing this xenobiotic. The authors of this work suggest that the hypocholesterolemic a c t i v i t y of the drug in rats results from hepatic accumulation of the xenobiotic choles terol ester which appears to promote the hydrolysis of natural cholesterol esters and thereby f a c i l i t a t e clearance of low-density lipoproteins.


Lipophilic Xenobiotic Conjugates

QUISTAD AND HUTSON

r

>

^ ° \

H

C

°2H

COOH R=COCH2C0 H = CH=CHC0 H = CH CHp0 H 2

2


2

2

rat c o w

[>C0 H

_ ^

2

[>(CH }COOH

n=4,6,8,IO,IZ.I4

2

dog from

cycloprate apples

[>CH=CH(CH ) n=l2.,l4 fcOfcH 2

oranges

ftv)COJt\

85% of the fatty acid component (39, Figure 5). An experimental acaricide, Ro 12-0470, is converted by apples into esters of C i 6 , C i 8 , C 2 0 , and C 2 2 fatty acids (40). These l i p o p h i l i c esters represented up to 22% of the applied dose just one day a f t e r treatment of f r u i t and f o l i a g e . A unique angelic acid conjugate was i d e n t i f i e d in carrots treated with carbofuran (41). This conjugate increases with time and has a h a I f - l i f e s i g n i f i c a n t l y longer than that of other carbofuran residues (42). The angelic acid conjugate appears to occur only in carrots (not found in potato or radish). Since angelic acid is abundant in the essential o i l of carrots, the i d e n t i f i c a t i o n of this l i p o p h i l i c residue i s a good example of how knowledge of an organism's natural constituents can lead to discovery of novel conjugates. The Significance of Xenobiotic L i p i d Conjugates Consideration of the biochemical and t o x i c o l o g i c a l significance of these conjugates (3,43) has been largely speculative and the sub j e c t cannot be treated in depth at this time. Clearly, the a b i l i t y of several classes of xenobiotics to participate i n the processes of l i p i d biochemistry has been demonstrated. In view of the high rate and quantity of turnover of natural l i p i d s , i t would seem unlikely that competitive effects of xenobiotics w i l l cause many problems for l i p i d metabolism. L i p i d conjugation is a mechanism for retention of xenobiotic acids and alcohols (32). However, where measured, the rates of removal of xenobiotic acylglycerols (8,43) and fatty acids (8) are similar to those of the natural analogs. The cholesterol esters appear to be more persistent. The only unequivocal example of a causative relationship between a tissue lesion (granulomata) and a xenobiotic l i p i d i s described later i n


QUISTAD AND HUTSON

Lipophilic

Xenobiotic

Conjugates

palmltate and


llnoleate

detergents

esters

n= 6, 9

apples

CH OC

Lipophilic Xenobiotic Conjugates - American Chemical Society

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