Cyanide In Cassava: A Review - Gavin Publishers

1. Introduction

Cassava (Manihot esculenta Crantz) is an important tropical root crop providing energy to about 500 million people [1,2]. Almost all the cassava produced is used for human consumption and less than 5 percent is used in industries. As a food crop, cassava fits well into the farming systems of the smallholder farmers in Nigeria because it is available all year round, thus providing household food security. Compared to grains, cassava is more tolerant to low soil fertility and more resistant to drought, pests and diseases. Furthermore, its roots store well in the ground for months after maturity. Cassava is important, not just as a food crop but even more so as a major source of cash income for producing households. As a cash crop, cassava generates cash income for the largest number of households, in comparison with other staples, contributing positively to poverty alleviation. The presence of cyanogenic glycosides in cassava which when broken down through enzymatic reaction librates hydrogen cyanide poses a great concern in cassava utilization as food and as industrial raw material. 

With respect to Cyanide levels, cassava varieties are broadly divided into two groups; the sweet cassava known for low cyanide content and the bitter cassava with its high characteristic content of Cyanogenic Glycosides (CGs) that is highly toxic when consumed [3-5]. Total cyanide in cassava products exists in form of CGs (linamarin and lotaustralin), cyanohydrin and free hydrocyanic acid (HCN). Notwithstanding the CGS, according to FAO, FAO, 2001 [6] 172 million tons of cassava were produced world-wide in 2000 with Africa accounting for 45%, Asia 28% and Latin America and the Caribbean 19%. The five main producing countries are Nigeria, Brazil, Thailand, Congo (DRC) and Indonesia.

The on-going challenge is to ensure that the presence of these cyanogenic glycosides is minimized through proper understanding and possibly control of factors that affect cyanogenic glycoside content of cassava. Roots and leaves contain the highest amount of linamarin [7]. 

2. Cyanide in Plants 

The cyanogenic glycosides are a group of nitrile-containing plant secondary compounds that yield cyanide (cyanogenesis) following their enzymatic breakdown. The functions of cyanogenic glycosides remain to be determined in many plants; however, in some plants they have been implicated as herbivore deterrents and as transportable forms of reduced nitrogen [8-10]. It is estimated that between 3,000 and 12,000 plant species produce and sequester cyanogenic glycosides. The major edible plants in which cyanogenic glycosides occur are almonds, sorghum, cassava, lima beans, stone fruits and bamboo shoots [11,12]. In certain sapindaceous seeds, HCN may arise during cyanolipid hydrolysis. More frequently, HCN production in higher plants results from the catabolism of cyanogenic glycosides. The approximately 75 documented cyanogenic glycosides are all O-β-glycosidic derivatives of ahydroxynitriles. Depending on their precursor amino acid, they may be aromatic, aliphatic, or cyclopentenoid in nature. Most are cyanogenic monosaccharides in which the unstable cyanohydrin moiety is stabilized by glycosidic linkage to a single sugar residue. Alternatively, in the cyanogenic disaccharides [e.g. (R)-amygdalin, (R)-vicianin, and linustatin] or trisaccharides (e.g. xeranthin), two or three sugar moieties, respectively, are involved in such stabilization. Sulfated, malonylated, and acylated derivatives of cyanogenic glycosides are also known. are also known. Cyanogenesis is not exclusive to those plant species accumulating cyanolipids and cyanogenic glycosides. All higher plants probably form low levels of HCN as a coproduct of ethylene biosynthesis [13]. This might explain why even 'acyanogenic' plants contain significant levels of the cyanide detoxifying enzyme β-cyanoalanine synthase. Cyanogenesis is also known in animals, but is restricted to the arthropods, notably to certain centipedes, millipedes, and insects. In fungi and bacteria, HCN may originate via oxidative decarboxylation of glycine.

 A cyanogenic food of particular economic importance is cassava (Manihot esculenta), which is also known by the names manioc, yuca and tapioca. Cassava is by far the most important cyanogenic food crop for humans and is an important source of dietary energy in tropical regions. The predominant cyanoglycoside in cassava is linamarin. It is present in leaves and tubers, both of which are eaten. Linamarin is also present in beans of the lima or butter type. Amygdalin is the cyanogenic glycoside responsible for the toxicity of the seeds of many species of Rosaceae, such as bitter almonds, peaches and apricots. Sweet almonds are low in amygdalin as a result of breeding processes. Their use in marzipan is common but the preparation procedure should eliminate most of the cyanide. Cyanogen levels can vary widely with cultivar, climatic conditions, plant part and degree of processing. Typical levels for some plant materials consumed by humans are found in (Table 1) below:

In areas of the world where cyanogenic plants such as cassava and lima beans comprise the major item of the diet, chronic cyanide poisoning and associated pathological conditions still exist [16]. It is highly desirable that the toxicity of cyanogenic plants to humans and livestock be reduced. This is achievable by: (a) selective breeding to produce low-cyanogen varieties, as was accomplished for almonds, (b) screening of natural populations for low-cyanogen varieties, (c) mutagenesis of protoplasts or cell cultures with subsequent regeneration of plants having desired mutant genotypes, or (d) genetic engineering. 

3. Role of Cyanogenic Glycosides in Plants 

A common feature of cyanophoric plants is that cyanogenic glycoside hydrolysis occurs at a significant rate only after their tissues have been disrupted by herbivores, fungal attack, or mechanical means. Although other explanations are possible, it is generally assumed that the glycosides and their catabolic enzymes are separated in the intact plant by compartmentation at either tissue or subcellular levels [17]. These possibilities have been extensively tested in a single organism, namely the leaves of 6-day old light-grown sorghum seedlings [18]. Somewhat unexpectedly, the authors demonstrated that the substrate and its catabolic enzymes were localized within different tissues. The cyanogenic glycoside dhurrin was sequestered in the vacuoles of epidermal cells, whereas the 3- glycosidase and hydroxynitrile lyase were present almost entirely in the underlying mesophyll cells. These two enzymes were located in the chloroplasts and cytosol, respectively. It therefore seems likely that the large-scale hydrolysis of dhurrin, which probably provides a defense mechanism against herbivores by liberating HCN, occurs only after tissue disruption allows the mixing of contents of different tissues.

Available evidence from other plant species, however, favors compartmentation of components of the 'cyanide bomb' at the subcellular level. In cassava, cells throughout the entire root cross-section possess both cyanogens (principally linamarin) and linamarase [19]. As in sorghum, highest glycoside levels are found in outer cell layers, again suggesting the involvement of cyanogens in defense against herbivores or pathogens, but the subcellular localizations of linamarin and linamarase remain unknown. In Phaseolus lunatus, the low recoveries of linamarin, linamarase, and hydroxynitrile lyase in leaf mesophyll protoplasts pointed to other tissues, perhaps the epidermis, as the principal site for these components [20]. Although these data cannot unequivocally distinguish between an epidermal or mesophyll location, it seems certain that the P. lunatus linamarase is apoplastic. Leaf discs hydrolyzed externally supplied linamarin, and about one-third of the total linamarase activity was extractable by multiple infiltrations of the leaves. The T. repens linamarase was detected by immunocytofluorescence in cell walls, especially those of the epidermis, and in the cuticle. More recently, protoplast isolation and tissue filtration experiments with Hevea endosperm showed that linamarin and the hydroxynitrile lyase were intracellular but that linamarase occurred both intra- and extracellularly. The apoplastic distribution of most linamarases contrasts with the intracellular location of sorghum dhurrinase, a fact perhaps related to the nonglycoprotein character of the latter [17]. 

The physiological importance of cyanogenic compounds in plant metabolism is currently receiving renewed interest. As with other secondary products, cyanogenics were originally viewed as excretory substances, but their turnover (seasonal and even diurnal) argues strongly against this hypothesis. 

Given the well documented toxicity of HCN, a role in plant protection against herbivores, pathogens, and competitors is appealing. Much evidence, indeed, favors a defence function for cyanogenics against certain animals including insects [21]. 

4. Cyanide in Cassava

Of all cyanogenic crops, the most agronomically important, is the tropical root crop, cassava (Manihot esculenta, Crantz). More than 153 million tons of cassava is produced annually, and it is the major source of calories for many people living in the tropics, particularly sub-Saharan Africa [22].

Cassava leaves have higher protein content, contain vitamin C and vitamin A and provide some dietary fiber [23]. Much of the protein in the leaves is made up of linamarase, the enzyme that detoxifies the cyanogenic glycosides in cassava [24]. However, each parts of cassava plants (leaves, stem, root) contains high levels of cyanogenic glycosides; linamarin, lotaustralin, and amygdalin [25,26] (Figure 1), with linamarin been the most predominant cyanogen. Linamarin is rapidly hydrolyzed by linamarase to glucose, acetone cyanohydrin, and hydrogen cyanide.

Under neutral conditions, acetone cyanohydrin decomposes to acetone and hydrogen cyanide.

The cyanide level of cassava varies from about 75 to 350 ppm but can be up to l000 ppm or more depending on the variety, plant age, soil condition, fertilizer application, weather, and other factors [27-29].

Studies have shown that the levels of cyanogenic glycosides in cassava roots are generally lower than that in the leaves and stems [30,31]. Cassava roots have been reported to contain cyanide content of 10-500 mg/kg of dry matter [32] and the leaves were reported to contain 53-1300 cyanide equivalents/kg of dry matter [33].

Cassava cultivars are classified as “bitter” or “sweet” depending on the level of cyanogenic glucoside (hence hydrogen cyanide). Values from 15-400 mg of hydrogen cyanide per kilogram of fresh weight of cassava roots have been reported for bitter varieties. Sweet varieties of cassava (low cyanide content) will typically contain approximately 15-50 mg hydrogen cyanide/kg fresh cassava. Sweet varieties of cassava can be processed adequately by peeling and roasting, baking or boiling, while bitter varieties of cassava (high cyanide content) require more extensive processing such as drying, fermentation etc. Bitter cassava varieties are more drought resistant and thus more readily available and cheaper. However, owing to food shortage in times of drought, less time is available for the additional processing required for cassava products. Highly toxic hydrocyanic acid (HCN) is released from the cyanogenic glucosides during hydrolysis by the enzyme linamarase (present in the root peel of cassava).

The World Health Organisation (WHO) has set the safe level of cyanogens in cassava flour at 10 ppm or 10 mg HCN /kg, while in Indonesia the acceptable limit is 40 ppm [34-37]. Consumption of cassava and its products that contain large amounts of cyanogens may cause cyanide poisoning with symptoms of vomiting, nausea, dizziness, stomach pains, weakness, headache, exacerbates goitre and diarrhoea and occasionally death [37-47].

Although processing methods can reduce linamarin and cyanide in food, improperly processed cassava products would contain some amount of residual linamarin and hydrogen cyanide. This would result in the potential toxicity of the cassava products. Indeed, cases of cyanide toxicity from the consumption of inadequately processed cassava products have been reported [40].

5. Factors Affecting Cyanide Content of Cassava

5.1. Cultivar

Thousands of cassava cultivars have been developed that are adapted to local conditions and differ in their ability to tolerate pest and diseases, yield, nutritional and cooking qualities of food products. Cassava is propagated clonally from stem cuttings so there is minimal variation between individuals of one cultivar when grown under the same environmental conditions. All cassava cultivars contain cyanogenic glucosides, however, a wide variation in the concentration of cyanogens exists among different cultivars. This can range from 1 to 2,000 mg/kg [37]. Cultivars with 100mg/kg are called bitter [43]. A study in Fiji by [44] on 17 different cultivars grown in the same environmental confirmed the influence of cassava variety on levels of cyanogenic glucosides (and hence hydrogen cyanide) content. The 17 different cultivars had cyanide levels of 14 -121 mg/kg.

5.2. Climatic Conditions

Cassava, a perennial shrub thrives in tropical and sub-tropical conditions. In general, the crop requires a warm humid climate. Temperature is important, as all growth stops at about 10ºC. Typically, the crop is grown in areas that are frost free the year round. The highest root production can be expected in the tropical lowlands, below 150 m altitude, where temperatures average 25-27°C, but some varieties grow at altitudes of up to 1500 m. The plant produces best when rainfall is fairly abundant, but it can be grown where annual rainfall is as low as 500 mm or where it is as high as 5,000 mm. The plant can stand prolonged periods of drought in which most other food crops would perish. This makes it valuable in regions where annual rainfall is low or where seasonal distribution is irregular. In tropical climates the dry season has about the same effect on Cassava as low temperature has on deciduous perennials in other parts of the world. The period of dormancy lasts two to three months and growth resumes when the rains begin again. Cassava is drought resistant and grows well in poor soil (Java Cassava, 2007). The problem however is that cyanide content of cassava tends to increase during periods of droughts and or prolonged dry weather due to water stress on the plant [24]. For example, in Mozambique, about 55% of the sweet fresh roots were extremely toxic and the remainder moderately so during drought like conditions. Similar observations were recorded in Democratic Republic of Congo [45], and various citations in Africa. Splittstoesser and Tunya (1992) [46] reported that cassava grown in wet areas contain relatively lower amount of cyanide than those grown in drier areas.

5.3. Fertilizer

There is a general consensus that crop yields do increase with application of fertilizer, there is debate however on the relationship between addition of fertilizer and cyanide content of cassava. Studies in the Philippines [47] concluded that application of fertilizer does not significantly affect cyanide content. It further suggested that the amount of nutrient in the soil does not considerably contribute to the cyanogenic character of the cultivar. In Ethiopia, Endris (1977) [48] suggested that the cyanogenic content of cassava roots was significantly reduced by potassium application. In Nigeria, Okwu and Awurum (2001) [49] were able to prove that the value of HCN in the cassava samples decreases as fertilizer levels increases.

5.4. Health Implications of Cyanide

The toxicity of cyanogenic glycosides and their derivatives is dependent on the release of hydrogen cyanide. Toxicity may result in acute cyanide poisoning and has also been implicated in the etiology of several chronic diseases (FAO/WHO, 2012 [50]. Dietary exposure to elevated levels of some cyanogenic glycosides in food has the potential to cause acute cyanide poisoning or a debili‐ tating irreversible neurological condition in the long term.

High and sustained cyanogens intake at sub-lethal concentrations from cassava or cassava flour in combination with a low intake of sulfur amino acids has been reported to cause Konzo in women and children [5]. Konzo is an upper motor neuron disease characterized by irreversible but non-progressive symmetric spastic paraparesis that has an abrupt onset. It mostly affects children and women of childbearing age [42].

6. Tropical Ataxic Neuropathy (TAN)

It is another health problem associated with continuous consumption of improperly processed cassava products. TAN is used to describe several neurological syndromes attributed to toxiconutritional causes. TAN has occurred mainly in Africa, particularly Nigeria [52] and is common among people of 40 years and above (FSANZ, 2004). Dietary exposure to cyanide from the monotonous consumption of inadequately processed cassava products over years is responsible for the cause of the disease. Symptoms of TAN include sore tongue, optical atrophy, neuro sensory deafness, and sensory gait ataxia [52].

7. Goiter and Cretinism

They are common diseases in developing countries due to low intake of iodine (