SCIENTIFIC

ptaquiloside in milk a few hours after start­ ptaquiloside in milk ing to feed and for a period of more than 80 h after the animal has stopped eating SIR - Ptaquiloside, a norsesquiterpene1 in Merida, Venezuela, with an average milk bracken. This is a wide window for found in bracken ( ), is production of 20±2 litres per day. For five ptaquiloside to be present in milk. Despite a potent carcinogen2 that binds covalently consecutive days we gave the cows artificial the fact that cattle do not normally choose to , causing the splitting of diets containing 6.0 kg per day (2,400 to to eat bracken, they can be forced to eat it uncoiled DNA3• Its potential for causing 10,000 mg per animal per day of ptaquilo­ during periods of drought when bracken tumours and other ailments in farm ani­ side) of previously analysed7 freshly col­ remains green, or when natural grasses are mals is well established4. The observation lected young bracken fronds (neotropical overgrazed, or when animals wander into that milk from cows feeding on a diet con­ var. caudatum ). Milk samples were drawn dense bracken thickets. Therefore, there is taining bracken fronds is carcinogenic and each morning, frozen and analysed imme­ a rather high risk to humans of ingesting mutagenic to mice and rats5•6 led to the diately8 until all ptaquiloside disappeared. milk containing ptaquiloside in areas suspicion that a bracken compound was None of the cows showed signs of intoxica­ where bracken is dominant. If a person present in this milk. When ptaquiloside tion during treatment. drinks about 0.5 litres of milk daily from a was first discovered, it was the chief candi­ We detected and quantified unstable, cow producing 20 litres of milk per day, date for this , but has never water-soluble ptaquiloside by selectively and this cow has eaten a subtoxic dose of been demonstrated to be present in milk. extracting it from deproteinated, defatted ptaquiloside of 5,000 mg per day (as would Here we report our attempts to ascertain milk as stable pterosin B, the only alkali­ be contained in 6 or 7 kg of fem, which typ­ its presence in milk, to estimate the rela­ induced decomposition product of ically might be eaten for a few days), this tionship between the amount of ptaquilo­ ptaquiloside. Pterosin B does not occur person will ingest 10.75 ± 1.45 mg side ingested by cows via bracken and the naturally in milk from bracken-fed cows, ptaquiloside per day. amount excreted through milk, and to and appears only after changing the pH of Although we cannot reach any definite assess the risk of drinking raw milk from the defatted milk to 11 for 2 h at 40 °C. conclusions at present about toxicity to cows exposed to bracken. We find that The per cent conversion curves of humans at the intake levels reported here, ptaquiloside is excreted in milk at a con­ ptaquiloside in milk from bracken-fed as there are as yet no data on the toxicolo­ centration of about 8.6 ± 1.2% of the cows and artificially ptaquiloside-contami­ gy of ptaquiloside in humans, the acute amount ingested by the cow, and is linearly nated milk in alkali at 38 °C overlap strik­ toxic effects of this chemical to rabbits at dose-dependent. ingly well over the 70 min of this 50 -100 mg per kg (ref_ 9), as well as the We used milk from six two-year-old experiment. We found no other secondary expected consequences of ingesting a car­ cows from an agricultural research station metabolite of bracken that could produce cinogen and DNA-damaging compound at pterosin B under these mild conditions. this concentration, strongly indicate cause 0 30.00 0 Thus, pterosin B represents only its pre­ for concern. It is certainly likely, in our q_ (/) a ..... 'c 25.00 cursor, ptaquiloside. We performed quan­ view, that ptaquiloside in milk is responsi­ X ., titative assessment by dichloromethane ble for the connection between bracken --"'E a-- 20.00 extraction and reverse-phase HPLC analy­ infestation and the incidence of gastric 8!:i-"'"'., 15.00 sis against standards. We also identified cancer in populations of farmers inhabit­ E?~ 10 o "OD 10.00 pterosin B in milk extracts by comparison ing cattle-range areas in Costa Rica and :t::.£ of retention times in three mobile phases in other countries where bracken growth is ~.g 5.00 with pure compound and by the exact dense (Andean South America, Central i 0.00 coincidence of the ultraviolet spectrum of America and southern Mexico). 0 24 72 120 168 216 the suspected peak ("-max= 218, 260 nm) Miguel E. Alonso-Amelot Hours after beginning of treatment by stopped-flow scanning of the eluate. Uvidelio Castillo _§ 70 We detected ptaquiloside in milk for Grupo de Quimica Ecol6gica, ~ 60 b the first time 38 h after feeding cows with Oepartamento de Quimica,