Chamal (Dioon edule Lindl.) in the State of San Luis Potosí, México Laura Yáñez-Espinosa* INTRODUCTION many D. edule populations, because the (0.1% fresh weight) and macrozamin (0.6% pproximately 20% of world cycad species is considered extremely toxic to fresh weight) in Dioon edule seeds and Aspecies are found in Mexico, 80% animals, and exterminating these plants is concluded that the seeds are highly toxic of which are endemic (Vovides, 2000). considered an effective option to prevent for rodents. At least nine species are present in the harm to livestock. One portion of the Huasteca region is state of San Luis Potosí: Dioon edule, According to the farmers, cattle con- inhabited by the indigenous Xi’iuy people, Zamia fi scheri, Z. loddigesii, Ceratoza- suming young leaves or cones is fatal; whose major activity is agriculture, based mia hildae, C. kuesteriana, C. latifolia, the effects start with diarrhea, progress on cultivation of maize, beans, and sugar C. mexicana, C. microstrobila, and C. to paralysis of the hind legs, and fi nally cane. Hunting does not represent a major zaragozae (Yáñez-Espinosa, 2006) [Ed’s result in death. This could be attributed source of food, but gathering products, note: Zamia vazquezii is also found in to toxic secondary metabolites produced like maguey (Agave spp.) and chamal San Luis Potosi]. Dioon edule LINDL. is the in cycads that apparently serve as protec- (Dioon edule) derivatives are consumed. most widely distributed, particularly in tion against predators. Although it has The harvest of chamal has played a major the Huasteca zone located in the Sierra been suggested that these compounds role in the Xi’iuy diet from time imme- Madre Oriental. Habitat deterioration of are responsible for the control of devel- morial, since this is a substitute when D. edule is evident, since many natural opment and maintenance of herbivores maize and bean yields are low. It is com- forests have been converted into either associated with cycads, two of these mon for the Xi’iuy to consume tamales, pastureland for cattle or sugarcane fi elds. compounds, cycasin and macrozamin, can gorditas, and tortillas made from chamal, Cattle breeding has been devastating for cause severe intoxication in addition to the dough made from boiled and ground ______carcinogenic and neurotoxic effects in seeds. Some recipes alert the maker that *Universidad Autónoma de San Luis Potosí, Instituto de Investigación de Zonas Desérticas mammals (Schneider et al., 2002). One improperly prepared chamal foodstuffs Altair 200, Col. del Llano, 78377 San Luis Potosí, study has reported low values of cycasin may cause vomiting, gastrointestinal dis- S.L.P., México

a

c

b d

Fig. 1. (A) Dioon edule population located in a well conserved habitat; (B) land conversion to agriculture conserving few individuals of D. edule; (C) leaf herbivory by Eumaeus childrenae caterpillars;(D) D. edule population damaged by fi re when Opuntia plants were established. The Cycad Newsletter 32(2/3) June/September 2009 Page 18 ease, or “fatal consequences” in people The following sections represent the crowns. Adult individuals were classifi ed when consumed (Bassler, 2000). results of three separate investigations as male or female from observation of the Besides the common use of chamal as that were conducted to address these mature or immature cones, or their ves- a food source, it also has medicinal, or- three questions. They were written inde- tiges, on the stem. namental, and other uses. Our research pendently, and each has its own title and Analyses of the soil sampled from the approach to study chamal was multidis- authors. six sites indicated that Dioon edule has a ciplinary, and the proposal included eco- preference for clay soils consisting of over logical, ethnobotanical, and toxicological Population Structure of Chamal 50% clay particles, 17% organic matter, investigations. We were aware that this G. Rubio Méndez, R. Mora Cardona and a slightly basic pH (average pH = 7.2). would be a long-term project, but our ex- & L. Yáñez-Espinosa This soil, because of its density, retains periences by the end of the fi rst year in- Preliminary exploration of an altitude moisture well and tends to be more nutri- clude benefi ts like the promotion of con- gradient (400, 1000 and 1200 m above sea ent-rich than other soil types. The aver- servation of existing populations of Dioon level [masl]) in San Luis Potosí indicated age calcium content of 82.43 meq 100g-1 is edule, a species that is poorly known in an abundance of Dioon edule plants, as very high, and the soil is slow-draining and the state of San Luis Potosí; compilation well as a high frequency of use of chamal compacts easily (Barnes et al., 1998). of traditional knowledge of the Xi’iuy for food, medicine, and ornamental and We found two populations of Dioon people related to this species; and ex- trade objects. Planned fi eld routes were edule with conditions of well conserved ploration of possible toxic effects of the traversed in the company of male key in- habitat of high quality. There were two seeds of this species as a food source. formants that were selected on the basis populations with intermediate quality Thanks to the Cycad Society grant in 2007, of their knowledge, leadership, and moral habitat and intermediate plant density, we were able to carry on the research, infl uence on their communities, as well as and two populations with low quality with complementary funds and infrastruc- their willingness to cooperate with us. habitat, mainly caused by cattle and land ture provided by the Autonomous Univer- Six selected sites were located with conversion to agriculture (Fig. 1). In the sity of San Luis Potosí, and collaboration GPS, and the habitat in each was char- six sites, most of the Dioon edule plants of three B.Sc. students from three dif- acterized by defi ning vegetation type, showed signs of herbivory by Eumaeus ferent disciplines—agronomy, geography, fl oristic composition, soil characteristics, childrenae (Lepidoptera: Lycaenidae) and chemistry—that worked together to and threats, such as deforestation, fi re, caterpillars, that have been previously develop the research project. cattle incidence, and others. Dioon edule reported for the Huasteca region (Contre- The study area comprises the munici- population structure and spatial distribu- ras-Medina et al., 2003). palities of Tamasopo, Rayón, and Santa tion was characterized in two sampling Species richness in the high-quality Catarina in the Huasteca region. These transects of 100 m length by 2 m width. habitats—distributed in the ecotone of were selected to carry on the research Plants intercepted in each transect were submontane shrubs and Quercus forest— based on previous knowledge of the utili- counted and their heights and diameters was high and included Flourensia lau- zation of chamal by the Xi’iuy people. We measured; also assessed were the number rifolia DC. (Asteraceae) and Quercus of leaves per plant and the life stage. stated three main questions, as follows: polymorpha SCHLTDL. & CHAM. (Fagaceae). In Dioon edule life stages were classifi ed as: the lower quality habitats—distributed in 1) What is the situation of the natural 1) seedlings – individuals with no visible the ecotone of submontane shrub and dry populations and habitat of Dioon edule? stem, one to few leaves not arranged in tropical forest—were Bursera simaruba a crown; 2) juveniles - non-reproductive 2) What and how many ethnobotanical (L.) SARG. (Burseraceae) and Pseudobom- individuals with no visible stem or barely categories can be recognized for Dioon bax ellipticum (KUNTH) DUGAND (Bomba- edule plants? visible, and leaves arranged in one or caceae). 3) How can toxins present in the seeds of more well-developed crowns; and 3) All populations showed a low fre- Dioon edule affect the Xi’iuy people adults - reproductive individuals with quency of female adults (Table 1), but it that eat them? a visible erect stem and one or more is interesting that populations inhabiting conserved habitats were located closer to Xi’iuy indigenous communities, and less conserved habitats were farther away from Xi’iuy communities. Ironically, fe- male cone harvest pressure is higher in the closer populations. The sex ratio in the medium and highly conserved habi- tats averaged 2:1 (M:F). Populations from more distant habitats have cattle pres- ent, and all plants in the population are b threatened in order to avoid toxic dam- age. Seedlings and juveniles showed a low frequency in the lower quality habitats, and the sex ratio was of 1.4:1. We also evaluated cone and seed devel- opment, seed production, and sporophyll morphology in 13 mature female cones harvested from two sites authorized by the Mexican government (SEMARNAT No. a c 00507/08) (Fig. 2). Average weight of female cones was 1.4 kg, and the aver- Fig. 2. (A) Female cone of Dioon edule being weighed and (B) dissected; age number of sporophylls was 220, with (C) sporophylls with attached seeds removed for characterization. an average of 180 well-developed seeds. The Cycad Newsletter 32(2/3) June/September 2009 Page 19 These seeds were sowed last July 2008 in regulate its use with the aim of conserving for grinding corn) to produce a dough. The greenhouse conditions to evaluate germi- populations without detriment of every- tamales are small dough balls of 4-7 cm, nation, but this process has not fi nished. one’s benefi t. plain or fi lled with egg or chickpea stew, Food processing of chamal seeds must and occasionally wrapped with leaves of Alimentary Uses of Chamal be done correctly to avoid symptoms like Arundo donax L. Finally, the tamales are E. Tristán Martínez & J. Fortanelli Martínez vomiting, diarrhea, nausea, and motor boiled in water for some minutes until After exploratory trips into the Xi’iuy function loss in people—termed ‘enchama- they fl oat, and then they are ready to region, fi ve localities with abundant Dioon lar’. There are variations in the processing eat (Fig. 3). There are some women from edule populations were selected for this procedures, ingredients, amounts, etc., one community dedicated to cooking the portion of the study. Seventeen individu- and this apparently is related to regional tamales and selling them nearby in pack- als were interviewed, mostly women that variants and economical purposes of the ages of fi ve, and occasionally the tamales agreed to share their knowledge about fi nal product. The most frequent alimen- are sent to another country. the alimentary uses of chamal. The ques- tary product is the ‘tamal of chamal’, The chamal (Dioon edule) is a plant tionnaire included questions pertaining which is described below. strongly related to agriculture-gatherer to ethnobotanical categories, type of Xi’iuy culture because it has allowed use, harvest process, and food process- Seed harvest. Female cones are re- them to subsist in marginalized conditions ing. To include more detailed information moved from the plant during April and by functioning as a substitute for maize in about the processing of the ‘tamales de May, when the chamal is ‘sazón’ (physi- times of food shortage. Nevertheless, the chamal’, as they are locally known, we ologically mature) and then opened with a knowledge of the proper cooking proce- worked with three families who allowed knife or stricken with a rock. dure is endangered because new genera- us to monitor and record—through digital tions are not interested to learn. photos and video—the entire process. Gametophyte removal. Complete seeds Data taken included volumetric and chro- are dried, to store, or consumed immedi- Toxicity of Chamal nometric measurements, organoleptic ately after removing the sclerotesta. The Covarrubias Camarillo & E. García Chávez attributes, and Spanish and Xi’iuy names gametophyte is sectioned in three to four The aim of this component of the study for each activity and kitchen utensil. portions, with or without embryo. was to evaluate the toxicity of the seeds Data processing was merely descriptive of Dioon edule LINDL. (seed, gametophyte, and qualitative because of directed non- Gametophyte cooking. The gameto- and sarcotesta) and the possible damage random sampling. phytes are boiled in bleach prepared with to the kinematics (locomotion) of male The botanical categories related to a mix of water, ash, and lime for 5 to10 Wistar rats by repeated exposure of dif- Dioon edule and its uses were as follows: minutes, and then left to stand for 40 ferent treatments (0.5 g / kg / day) for 15 1) ornamental-ceremonial - the leaves minutes to 2 hours. Favorite ashes come days. Rats received humane care, and the or the entire plant are used to decorate mainly from Quercus laeta LIEBM., Q. poly- experiment adhered to the Institution’s altars; 2) ornamental with trade purpose morpha SCHLTDL. & CHAM., and Cordia bois- guidelines and the Mexican Offi cial Norm - the entire plant is removed from the sieri A.DC. (NOM-062-ZOO-1999) regarding technical root and sold to dealers who transport specifi cations for production, care, and the plant to the city; and 3) alimentary. Grinding. After some testing to assure use of laboratory animals. Different parts of the seed are used to seeds are cooked and have absorbed the The animals were assigned to fi ve process the most frequent made ‘torti- exact quantity of bleach, they are ground groups: one control group and four groups llas’, ‘atoles’, ‘gorditas de horno’ and in a mill and a ‘metate’ (fl at stone used of D. edule – seed, gametophyte, sar- ‘tamales’. Although nearly all people self- consume the tamales, there are also some that sell them. It is necessary to mention that the harvest of this plant for alimentary pur- poses is apparently independent of ethnic identity, and is, instead, more related to economic level. In other words, frequent consumers of chamal are typically indig- enous people and poor mestizos. How- ever, chamal also represents a threat for peasants mainly dedicated to livestock, a c because plants “invade” grasslands and grazing lands in forests. The highly toxic new leaves are palatable for cattle, and shortly afterwards the animals lose motor function. Regarding the ornamental use, in one community plants are gathered and sold to buyers from San Luis Potosí. They pay around US$40.00 for four sacks with four to six plants in each. Here, Dioon edule is not only a complement to the inhabitants’ diet but also a source of income. Consid- b d ering that D. edule is an endemic threat- ened species (PROY-NOM-059-ECOL-2001) Fig. 3. (A) Gametophyte color after boiling in the bleach mix; (B) grinding dough in a ‘metate’; (C) fi lling the (Yáñez-Espinosa, 2006), it is critical to tamales with egg stew; (D) tamales ready to eat with some chamal leaves at the bottom of the pot. The Cycad Newsletter 32(2/3) June/September 2009 Page 20 cotesta, and 2-Amino-3-(methylamino)- crease in body weight gain in the group of as a result of the content of BMAA in the propionic acid (BMAA, which is a chemi- rats treated with sarcotesta 6 days after seeds (Banack & Cox, 2003). Studies in cal found in cycads and thought to have exposure, which remained at 9 and 15 animals exposed to BMAA showed altera- neurodegenerative effects on mammals). days (p<0.002). The group of rats exposed tions characterized by inability to extend Control rats received deionized water to BMAA only showed an increase after 9 legs, dragging gait, hyper-excitable state, daily for 15 days by gavage; BMAA rats days of exposure (Fig. 5). However these ataxia, and diffi culty in walking (Karamyan received the dose by IV acutely; and rats changes were not associated with the con- & Speth, 2008). in the other three groups received a water sumption of food, because an increase in extract of seed, gametophyte, and sar- the consumption of food was not observed REFERENCES cotesta fl our by gavage for 15 days. The in the same periods of time (Table 2). The Banack, S.A. & P.A. Cox. 2003. Distribution of experimental dose was based upon previ- group of animals exposed to BMAA signifi - the neurotoxic nonprotein amino acid BMAA ous reports in rats (González, 1990) which cantly reduced their consumption of food in Cycas micronesica. Bot. J. Linn. Soc. showed that steady-state brain concentra- after 12 days of exposure, but this de- 143:165-168. Barnes, B.V., D.R. Zak, S.R. Denton & S.H. Spurr. tions of 10-30 μg/g could possibly cause crease had no effect on the body weight 1998. Forest Ecology. 4th ed. Wiley. 792 pp. alterations in behavioral and locomotor of the animals (Table 2). Bassler, C.H. 2000. Recetario pame de San Luis activity (Fig. 4). In relation to the effect of treatments Potosí y Querétaro. Serie: Cocina indígena y The results showed a signifi cant in- on the kinematics of the animals, the popular. Consejo Nacional para la Cultura y results showed alterations in the locomo- las Artes. México. 160 pp. tion of the rats exposed to seeds, char- Bonta., M., O. Flores Pinot, D. Graham, J. acterized by a signifi cant decrease in the Haynes & G. Sandoval. 2006. Ethnobotany support phase and a signifi cant increase and conservation of tiusinte (Dioon mejiae STANDL. & L.O. WILLIAMS, Zamiaceae) in north- in the length and duration of stride com- eastern Honduras. J. Ethnobiol. 26:228–257. pared with the control group (p<0.001, Contreras-Medina, R., C.A. Ruiz-Jiménez & I. Table 3). These results were translated Luna V. 2003. Caterpillars of Eumaeus chil- into an increase in the velocity of the drenae (Lepidoptera: Lycaenidae) feeding stride. The group of animals exposed to on two species of cycads (Zamiaceae) in the gametophyte showed a signifi cant de- Huasteca region, Mexico. Rev. Biol. Trop. crease in the length and velocity of stride, 51:201-204. compared with the control group. This González, A. 1990. Algunas interacciones en- tre Dioon edule (Zamiaceae) y Peromyscus behavior was also observed in the group mexicanus (Rodentia:Cricetidae). La Ciencia treated with BMAA (Table 3). These results y el Hombre 1:93-112. were associated with a decrease in the Karamyan, V.T. & R.C. Speth. 2008. Animal velocity of the step, and with a slow and models of BMAA neurotoxicity: A critical re- eventful displacement. The results of the view. Life Sci. 82:233–246. phases of fl exion, extension, and balance Vovides, A. 1999. Zamiaceae. P. 16, in: J. Rze- showed no signifi cant differences (data dowsky & G. Calderón (eds.), Flora del Bajío not presented). y de regiones adyacentes. Fasc. 71. Pátzc- uaro, Michoacán, México. Consumption of seeds of Cycas micro- Yáñez-Espinosa, L. 2006. Las Cycadas. Biología nesica by Chamorro people on Guam has y Conservación en México. Univ. Autón. been associated with neurodegenerative Chapingo. Estado de México, México. 208 pp. disease (amyotrophic lateral sclerosis) a Table 1. Dioon edule population density in the Huasteca region, San Luis Potosí, México. Adult Habitat Seedling Juvenile male female Total quality (no. ha-1) (no. ha-1) (no. ha-1) (no. ha-1) (no. ha-1) High 1413 963 238 100 2713 Medium 0550 438 175 088 1250 Low 0363 263 175 125 0925

b Table 2. Daily food consumption of Wistar rats (g).

Fig. 4. (A) Behavioral and (B) locomotor Treatments Day 1 Day 3 Day 6 Day 9 Day 12 Day 15 activity evaluation in Wistar rats Control 18.0 ± 5.5 21.3 ± 6.5 25.7 ± 3.7 25.7 ± 3.4 27.5 ± 3.8 19.9 ± 7.0 fed various parts of chamal seeds. Seeds 17.0 ± 3.7 23.0 ± 2.1 20.7 ± 2.7 23.9 ± 1.5 26.8 ± 2.0 23.1 ± 2.4 Gametophyte 24.8 ± 3.8* 25.2 ± 4.7 24.2 ± 5.2 23.8 ± 5.2 23.0 ± 6.5 13.0 ± 2.9 Sarcotesta 30.0 ± 0.0* 30.0 ± 0.0* 30.0 ± 0.0 29.6 ± 0.9 27.4 ± 2.1 13.3 ± 3.0 BMAA 18.9 ± 2.8 26.2 ± 2.3 22.9 ± 2.9 25.8 ± 2.9 07.2 ± 7.9* 03.4 ± 2.3* BMAA = beta-methylamino alanine; values represent mean ± SD; *p < 0.000 vs control; n = 6

Table 3. Analysis of kinematics (locomotion) of Wistar rats fed various parts of chamal seeds.

Parameters Control Seeds GametophyteSarcotesta BMAA Phase support (s) 00.5 ± 0.1 00.4 ± 0.1*°b 00.6 ± 0.1 00.5 ± 0.1 0.6 ± 0.1 Length of stride (cm) 09.0 ± 1.3 10.3 ± 1.2*°a 07.2 ± 1.3* 08.2 ± 1.3 6.5 ± 0.9* Duration of stride (s) 00.7 ± 0.1 00.5 ± 0.1*°b 00.7 ± 0.1a 00.6 ± 0.1*° 0.7 ± 0.1 Fig. 5. Body weight of Wistar rats fed various parts of Velocity of stride (cm/s) 13.9 ± 3.7 21.7 ± 6.7*°ab 10.3 ± 3.7*a 13.3 ± 3.6 9.3 ± 3.1*a

chamal seeds. (Values represent mean ± SEM. *p<0.02 a b control vs. sarcotesta; °p<0.02 control vs BMAA; n=6). *p < 0.001 vs control; °p < 0.001 vs BMAA; p < 0.001 vs sarcotesta; p < 0.001 vs gametophyte; n =6 The Cycad Newsletter 32(2/3) June/September 2009 Page 21