On-line version ISSN 1678-4936
J. Venom. Anim. Toxins vol. 2 n. 1 Botucatu 1996
LINAMARIN - THE TOXIC COMPOUND OF CASSAVA
1 The Center of Tropical Roots, CERAT-UNESP, São Paulo State University, Botucatu, State of São Paulo, Brazil, 2 Department of Pathology of the School of Medicine of Botucatu, State of São Paulo, Brazil.
ABSTRACT. Cassava is a widely grown root crop which accumulates two cyanogenic glucosides, linamarin and lotaustralin. Linamarin accounts for more than 80% of the cassava cyanogenic glucosides. It is a ß-glucoside of acetone cyanohydrin and ethyl-methyl-ketone-cyanohydrin. Linamarin ß-linkage can only be broken under high pressure, high temperature and use of mineral acids, while its enzymatic break occurs easily. Linamarase, an endogenous cassava enzyme, can break this ß-linkage. The enzymatic reaction occurs under optimum conditions at 25ºC, at pH 5.5 to 6.0. Linamarin is present in all parts of the cassava plant, being more concentrated on the root and leaves. If the enzyme and substrate are joined, a good detoxification can occur. All the cassava plant species are known to contain cyanide. Toxicity caused by free cyanide (CN¯) has already been reported, while toxicity caused by glucoside has not. The lethal dose of CN¯ is 1 mg/kg of live weight; hence, cassava root classification into toxic and non-toxic depending on the amount of cyanide in the root. Should the cyanide content be high enough to exceed such a dose, the root is regarded as toxic. Values from 15 to 400 ppm (mg CN¯/kg of fresh weight) of hydrocyanic acid in cassava roots have been mentioned in the literature. However, more frequent values in the interval 30 to 150 ppm have been observed. Processed cassava food consumed in Brazil is safe in regard to cyanide toxicity.
KEY WORDS: cyanide, cassava, linamarin, cyanogenic glucoside, residues, waste water, culinary and industrial uses.
The past 35 years have provided new knowledge of the process of cyanide biochemistry (5). Cassava roots provide an important source of energy-rich food for millions of people, including those in Brazil who lack appropriate nutrition. However, one of their nutritional shortcomings is their potential toxicity due to their two cyanogenic glucosides: linamarin and lotaustralin.
Cassava is a widely grown root crop in Brazil as well as in several other countries lying at 30º north and 30º south of the equator. In Brazil, cassava is broadly used in industry in the processing of a typical cassava flour named farinha as well as in starch extraction, and at homes for culinary purposes (3).
Processed cassava food consumed in Brazil is safe in regard to cyanide toxicity as cyanide is water-soluble and thus is carried by solid and liquid waste from the processing industries (3).
The amount of cassava waste produced depends on the processing method used. When cassava is used at homes for culinary purposes, the amount of waste is small and does not cause environmental hazards. In contrast, when cassava is industrially used, even the small flour factories, the so-called Casas de Farinha produce a considerable amount of waste since they are traditionally concentrated on a certain place. In Paranavaí, State of Paraná, for instance, 150 flour factories of different sizes produce amounts of residues which bring about a drastic environmental and economic impact (3).
LINAMARIN - THE TOXIC COMPOUND OF CASSAVA: According to Cooke (6), linamarin and lotaustralin, are the two different cyanogenic glucosides in cassava plant. Roots and leaves contain the highest amount of linamarin (8,14). Linamarin produces the toxic compound hydrogen cyanide (HCN) which can be hazardous to the consumer. Toxicity caused by free cyanide (CN¯) has already been reported, while toxicity caused by glucoside has not. Oke (12) reports that linamarin and lotaustralin are ß-glucosides of acetone cyanohydrin and ethyl-methyl-ketone-cyanohydrin, respectively. Linamarin is the most representative glucoside accounting for about 80% of the total cassava glucoside. The structure of linamarin is shown in Figure 1. Oke (12) also reports that linked glucosides are not toxic to the plants which contain them.
FIGURE 1. Linamarin structure.
Toxicity in higher animals results from the combination of cyanide with Fe which accounts for the formation of cyanohemoglobin. In regard to higher plants and microorganisms (10), cyanide interferes with the oxidative phosphorilation pathway by combining with cytochrome-oxidate and inhibiting electronic transportation, and consequently, the ATP (adenosine triphosphate) formation.
The average lethal dose of cyanide for higher animals was experimentally obtained and expressed in milligram per kilogram (mg/kg) of live weight. Oke (12) ascertains that the lethal dose is 1 mg/kg of live weight. Hence, cassava roots are classified into toxic and non-toxic depending on the amount of cyanide in the root. Thus, if the cyanide content is high enough to exceed such an average dose, the root is regarded as toxic. In the literature, values from 15 to 400 ppm (mg CN¯/kg of fresh weight) of hydrocyanic acid in cassava roots are reported, however, more frequent values in the interval 30 to 150 ppm (2) are observed. In addition, there are varieties of cassava which contain more than 1,000 ppm of CN¯(3).
O'Brien et al. (11) relate that in the processing of cassava roots, linamarase - the hydrolytic enzyme - remains active and catalyzes the reaction which releases one molecule of glucose, acetone and hydrocyanic acid. Linamarase has optimum pH at 5.5 to 6.0. Microorganisms which have ß-hydrolytic enzymes can also cause glucoside hydrolysis (12).
In general, animals have a detoxification mechanism which can avoid death when cyanide release is slow. This mechanism, depending on the pH, is present in swines (monograstic with pH 3.0 in the stomach), but not in bovine (polygrastic with pH 7.0 in the stomach) (12).
Microorganisms can grow in cyanide-containing substrates due to their anaerobic metabolism, their alternative metabolism regarding the respiratory chain (4) and their capacity to detoxify cyanide by splitting the CN¯ radical into carbon and nitrogen (7).
Waste quality and quantity vary a lot owing to factors such as plant age, time after harvesting and type and adjustment of industrial equipment.
Manipueira is the technical name for the water compound of cassava roots extracted in the pressing of the ground mass in flour processing. The water used in starch extraction dilutes manipueira reducing its organic load and cyanide content, but increasing its volume. The mass balance for cassava flour yields 300 liters of manipueira with less than 10% of dry matter for one ton of roots. Starch industries yield around 600 liters of diluted manipueira with less than 5% of dry matter, 5,000 COD (chemical oxygen demand) and 60 ppm of CN¯ for one ton of roots. Another controversial issue is the presence of the two cyanogenic glucosides, linamarin and lotoaustralin, released during cassava processing. The water used in the cassava processing carries high concentrations of these glucosides, which explains the high amounts of these toxic compounds in the residual liquid waste (13). Linamarin and lotoaustralin hydrolyze in the presence of acids and enzymes producing CN¯, and subsequently, hydrocyanic acid (HCN) (15).
Manipueira composition varies mainly in regard to cyanide content, which depends on the cassava cultivar used, the organic load content (9) and the industrial processing. Since manipueira is the water compound of the root, it contains most of the soluble and some insoluble substances in suspension. Almost all cyanogenic glucosides present in the disintegrated root mass are carried along with this residue.
Sobrinho (13) states that the liquid waste thrown away in the soil and in rivers is highly pollutant. Pollution is usually measured considering the population rate expressed in BOD (biochemical oxygen demand) as: 24 g/person/day (5 days - 20ºC). Pollution brought about by starch extraction industry corresponds to 150-250 person/day. Anrain (1) states that pollution produced by a large cassava processing industry is about 2,500 mg O2/liter in COD, which corresponds to pollution produced by 460/person/day.
CHARACTERIZATION OF CASSAVA LIQUID WASTE: Average data on cassava liquid waste are shown in Table 1. Such waste is characterized by high humidity, COD and cyanide. Manipueira mineral composition is shown in Table 2. High contents of phosphorus and potassium are predominant in manipueira mineral composition.
TABLE 1. Average data on cassava liquid waste (3).
TABLE 2. Mineral composition of manipueira of a farinha processing industry (3).
01 ANRAIN E. Tratamento de efluentes de fecularia em reator anaeróbico de fluxo ascendente e manta de lodo. Congresso Brasileiro de Engenharia Sanitária Ambiental, 12. Balneário de Camboriú, 1983. [ Links ]
02 CARVALHO VD., CARVALHO JG. Princípios tóxicos da mandioca. Inf. Agropec., 1979, 5, 82-8. [ Links ]
03 CEREDA MP. Caracterizacão dos resíduos da industrializacão da mandioca. In: CEREDA MP. Ed. Resíduos da industrializacão da mandioca no Brasil. São Paulo: Paulicéia, 1994: 1-50. [ Links ]
04 CEREDA MP., BRASIL OG., FIORETTO AMC. Atividade respiratória em microorganismos isolados de líquido residual de fecularias. Congresso Brasileiro de Microbiologia, 11. Florianópolis, 1981. [ Links ]
05 CONN EE. Cyanogenesis - a personal perspective. Acta Hortic., 1994, n.375, 31-43. [ Links ]
06 COOKE RD. Enzymatic assay for determining the cyanide content of cassava and cassava products. Cali: Cassava Information Center - CIAT, 1979: 1-14. [ Links ]
07 JENSEN HL., ABDEL-GHAFFAR AS. Cyanuric acid as nitrogen sources for microorganisms. Arch. Microbiol. 1979, 67, 1-5. [ Links ]
08 JESUS VS., MORAES CF., TELLES FFF., SEDIYANA CS. Teor de ácido cianídrico nas folhas e raízes de dez variedades de mandioca Manihot esculenta, GRANTZ, durante o primeiro ciclo. Rev. bras. Mand. Cruz das Almas, 1986, 5, 83-90. [ Links ]
09 LAMO PP., MENEZES TJB. Bioconversão das águas residuais do processamento de mandioca para produção de biomassa. Colet. Inst. Tecnol. Aliment., 1979, 10, 1-14. [ Links ]
10 NARTEY F. Cyanogenesis in tropical feeds and feedstuffs. In: VENNESLAND B., CONN EE., KNOWLES CJ., WESTLEY J., WISSING F. Eds. Cyanide in Biology. London: Academic Press, 1981: 115-32. [ Links ]
11 O'BRIEN MG., TAYLOR AJ., POULTER NH. Improved enzymatic assay for cyanogens in fresh and processed cassava. J. Sci. Food. Agric.,1991, 56, 277-89. [ Links ]
12 OKE OL. The role of hydrocyanic acid in nutrition. World Rev. Nutr. Dietetics., 1969, 11, 170-98. [ Links ]
13 SOBRINHO PA. Autodepuração dos corpos d'água. In: Curso Poluição das Águas. São Paulo, 1975. São Paulo: CETESB/ABES/BNH, 1975. cap.8, 6-9. (apostila). [ Links ]
14 VITTI P., FIGUEIREDO IB., ANGELUCCI E. Folhas de mandioca desidratadas para fins de alimentacão humana. Colet. Inst. Tecnol. Aliment., 1971/72, 4, 117-24. [ Links ]
15 WILLIAMS HJ. Estimation of hydrogen cyanide from cassava at organic solvents. Exp. Agric., 1979, 15, 393-9. [ Links ]
M. P. CEREDA - CERAT, Fazenda Lageado, Caixa Postal 237, CEP 18603-970 - Botucatu - São Paulo - Brasil.