Gossypol content analysis in Iranian whole cotton seeds and their meal

Cotton ( Gossypium spp.), is the most important textile fiber crop in the world and second only to soybean as the most important oil seed crop (Khan et al , 2002). Cotton is a major crop throughout the Iran and by-products of the cotton industry such as linted whole cotton seed (WCS) and cotton seed meal (CSM) are extensively used as dietary ingredients for dairy, beef cattle and sometimes poultry. Cotton seed and its by-products are fed to dairy cattle because of the nutritional value derived from their fat, protein, and fiber (NRC, 2001) . Cotton seed and cotton seed meal are excellent feeds that are widely fed to ruminant animals. The safe and effective use of these in diets for dairy cattle requires good nutritional information on which to base feeding decisions and an understanding of factors which contribute to variation in their nutrient and gossypol content (Calhoun, 2004).

Whole cotton seed (WCS) is a product of the cotton fiber industry that is extensively used as an energy and protein source in dairy cattle diets (Calhoun et al., 1995) . The advantages of including WCS in the diets of lactating dairy cows are well documented (Coppock, et al, 1987). Cotton seed meal (CSM) is a product of the oil extraction from WCS and is used as a protein supplement in diets for ruminants. Both WCS and CSM contain gossypol, a yellow poly phenolic compound found primarily in the pigment glands of the cotton plant (Robinson et al., 2001, Calhoun et al., 1995) that exists in the free (FG) and bound (BG) forms. In the intact whole seed, gossypol is mostly found in the free form. However, when cotton seed is processed, gossypol binds to proteins, possibly to the epsilon-amino group of lysine (Calhoun et al., 1995) .

Total gossypol (Free + Bound gossypol) content of seed kernels varies among cultivars within a species (Percy et al., 1996). One of the potential negative effects of feeding excess FG is a potential decline in the reproductive performance of dairy cows. Santos et al. (2003) demonstrated that as plasma gossypol concentrations increased, the risk for pregnancy in lactating dairy cows fed diets containing different types of cottonseed declined.

When in the bound form, gossypol is considered non toxic to ruminants because it cannot be absorbed in the digestive tract. However, some of the gossypol that is bound may be released as free gossypol (FG) during digestion, which then can be absorbed by the digestive tract. This phenomenon has been suggested with bound gossy-pol (BG) from processed WCS (Noftsger et al., 2000) and CSM (Wan et al., 1995; Blackwelder et al., 1998) . Due to gossypol content, the amount of cotton products fed to cattle has to be limited to avoid risk of toxicity (Coppock et al., 1987; Calhoun et al., 1995; Arieli, 1998) .

To avoid risk of gossypol toxicity, the objectives of this study was to determine the free and total gossypol content of Iranian whole cotton seed and cotton seed meal as aprotein source in dairy cattle and broiler chickens.

Material and Methods

Four most important cotton cultivars (Varamin, Bakhtegan, Meher and Sahel) were gathered from different parts of Iran. Sahel cultivated in the North of Iran, Bakhtegan mostly cultivated in the Fars province (South of Iran) and Varamin is the most important cotton cultivar that cultivated in the dry and semi dry zone of Iran. Meher is cultivated mostly in the Northwest of Iran (Figure 1). Some native cotton cultivars are cultivated in some villages of Iran too. Samples were refrigerated until analysis for Gossypol and the seed Index was determined. In order to express gossypol values based on whole seed, it is necessary to know the percentage of meats in the seed. The seed Index mean (percentage of meat in the seed for Varamin and Sahel were 51.3% and for Bakhtegan was 52% respectively. For other cultivars it considered 52%. Total and free gossypols were determined by the Official Methods of AOCS (AOCS, 1985 a, b). The Sahel, Varamin and Bakhtegan cotton seed meals (CSM) were obtained from different oil extraction factory that located near the farms. A subsample that was imported by private company from Uzbekistan was analyzed too. They were sub sampled and stored at -15 to -20oC until used for analyses (Calhoun, 1998).

в Mehr

□ Sahel

□ Baghtegan и Varamin

Figure 1- Cultivated cotton area in Iran in 2008

Result and discussion

The amount of total and free gossypol, expressed as a percentage of total and free gossypol, in cotton seed meats of commercially important cotton varieties that grown in different locations of Iran, were summarized in Table 1 and Figure 2. Also total and free gossypol of Iranian cotton seed meal (CSM) obtained from several oil extraction factories and an imported sample from Uzbekistan were summarized in table 2 and figure 3.

Table 1- Variation in total (TG) and free gossypol (FG) in meats of commercially important cotton seed varieties of Iran
Cultivar	Total gossypol%1	STD	Free gossypol%1	STD
Varamin	1.14	0.023	1.1	0.024
Sahel	1.27	0.085	1.2	0.029
Bakhtegan	1.08	0.005	1.1	0.006
Mehr	1.07	0.124	0.8	0.315
Native	0.58	0.142	0.53	0.103

1 Values are based on 100% dry matter basis

Table 2- Free and gossypol content of cotton seed meal (CSM) obtained from different cotton cultivar and a imported sample
Cotton seed meal	Free gossypol%1	STD	Total gossypol%1	STD
Varamin	0.03	0.01	0.54	0.13
Sahel	0.06	0.02	0.62	0.04
Bakhtegan	0.06	0.02	0.57	0.16
Uzbakestan	0.13	0.01	0.68	0.01

1 Values are based on 100% dry matter basis

Figure 2- Variation of total and free gossypol in the meats of most important Iranian whole cotton seed

0.80

0.70

0.60

0.50

0.40

0.30

0.20

0.10

0.00

Bakhtegan     sahel      varamin    Uzbakestan

• ■    Free Gossypol%

• ■    Total Gossypol%

Figure 3- Distribution of the free and total gossypol content in studied cotton seed meal

As can be seen from figure 2, Native and Sahel varieties had lowest and highest amount of free and total goss-ypol in seed meat among studied varieties respectively. Amount of free and total gossypol content in native variety were 0.53% and 0.58% respectively while in Sahel variety were 1.2 and 1.27% respectively.

As can be seen from figure 3 the lowest and highest amount of free gossypol were observed in Varamin and Uzbekistan varieties and these varieties with 0.54 and 0.68% had the lowest and highest amount of total gossy-pol respectively.

In the 1992 national cotton variety test (Rayburn and et al., 1993) reported that seed Kernel (meat) gossypol ranged from 0.64 to 1.09% (6.4-10.9 g Kg-1) within G. hirsutum cultivars. According the table 1 the gossypol content of Iranian cotton seed is in agreement with this range. The gossypol content of cotton seed affected by variety and environmental conditions (Pons, et al., 1953., Calhoun et al., 1995). Gossypol tends to be lower in seed plants that grown at locations with higher average temperatures while higher with increased rainfall, particularly when moisture is available during the maturation period (Pons, et al., 1953). There are some slight differences among the gossypol content of Iranian whole cotton seed (table 1) that is related to variety and environmental condition (Calhoun.,2004). Sahel is the most dominant cotton cultivar in the north of Iran (<500 mm annual rainfall) that the rainfall is higher than other part of Iran country. Varamin is the most important Iranian cotton variety that cultivated in dry zone with about <100 – 300 mm annual rainfall. Result of the table 1 showed that free and total gossypol in Sahel cultivar is higher than other cultivars.

As Pons et al. (1953) reported that the gossypol content of cotton seeds was negatively correlated with the temperature and positively correlated with the rainfall that the cotton plants were exposed to it when producing the seed, the highest amount of gossypol content was recorded in Sahel cultivar.

Meher variety cultivated in the northwest of Iran and it cultivation has limited in recent years. Free gossypol content of Meher is significantly lower than Varamin (p<0.05) but the difference between total gossypol is low. Although the rainfall in the northwest of Iran is higher than the central parts of Iran and its mean temperature is lower than the central part of Iran, the gossypol content is lower than Varamin that may be due to dominant effect of variety on the gossypol content of cotton seed and effect of location where the cotton is grown and regional differences also. Type of cotton, variety, growing conditions, harvesting and storage conditions certainly are important sources of variation in gossypol content of whole cotton seed (Calhoun, 2004).

Bakhtegan and Varamin cultivars displayed less variation for seed kernel gossypol than Sahel from the north of Iran. Bakhtegan were mostly cultivated in Fars province of Iran (100- 200 mm annual rainfall) that is similar with other central parts of Iran that Varamin variety was cultivated. Slight difference in total gossypol is related to regional differences (Percy and et al 1996), variety and environmental condition (Calhoun, 2004). Iranian native cotton seed were low gossypol cultivars, which enhances the value of seed for livestock feeding.

As determined by the AOCS method, the free gossy-pol content of CSM samples from Iranian cotton seed have a narrow distribution (Figure 3) and the difference is not significant.(P>0.05) but the CSM from Uzbekistan have higher gossypol content. Lordelo and et.al (2008) reported that the CSM samples obtained from Egypt and Peru contained a high concentration of gossypol, whereas the samples from Tajikistan and Iran had lower gossypol levels. The free gossypol content of CSM samples obtained from several countries had a wide distribution from 0.01 to 0.39%. Type of cotton, variety and growing conditions certainly are important sources of variation but harvesting and storage conditions and procedures used for oil extraction also have an important effect on gossypol content (Calhoun, 2004). In the United States, cotton seed was processed by screw presses, prepress solvent extraction, direct solvent extraction and by hydraulic pressing (Calhoun and et al, 1995b). When cotton seed was processed, availability of gossypol decreased, and more gossypol was detected in the bound form than in the free form (Mena et al, 1997). When in the bound form, gossy-pol is considered non toxic because it cannot be absorbed in the digestive tract (Mena et al, 2001).

By the mid to late 1980's, the predominate process was direct solvent extraction and this is still the predominate process, but in the last 10 years, there has been a rapid introduction of expanders into the direct solvent process. This process in now commonly referred to as the expander solvent process. Expanders dramatically reduced free gossypol levels compared with the direct solvent process without expanders (Calhoun and et al, 1995 b). Nearly all of the Iranian whole cotton seeds (WCS) are processed by screw pressing.

The higher temperatures and pressures associated with production of mechanically extracted and prepress solvent cotton seed meals lead to lower free gossypol values than for direct solvent meals. Processing method does not appear to affect total gossypol (Calhoun and et al, 1995 b). The mechanically extracted cotton seed meals have higher residual oil content than pre-pressed solvent or direct solvent cotton seed also (Calhoun and et al, 1995 b). The gossypol content of Uzbekistan cotton seed meal sample was higher than the Iranian cotton seed samples that are related to variety of cotton seed, environmental conditions and oil extraction procedures (Calhoun, 1995 a).

The safe and effective use of cotton seed and cotton seed meal in diets for dairy cattle requires good nutritional information on which to base feeding decisions and an understanding of factors which contribute to variation in their nutrient and gossypol content (Calhoun, 2004). Although the reliable nutritional information is essential for formulating and manufacturing of cotton products in animal diet, the low level of Iranian gossypol cotton seed products in cotton seed and especially cotton seed meal as a negative impact on the amounts of fed must be considered.

However CSM containing low levels of gossypol can be successfully used in broiler diets that contain adequate levels of all essential nutrients, without any adverse effects on body weight or mortality (Waldroup et al, 1968; Walkins et al, 1994).

Acknowledgments

The authors would be like to thank Dr. Millard C. Calhoun from Texas A&M university system for his assistance during sample analysis and Dr. M. M. Lordelo from Poultry Science Department of The University of Georgia, Athens for gossypol detection in some Iranian cotton seed. Appreciation is extended to Dr.S.W. Kulmann from Texas A&M university system and Dr. Nike Dale from USDA for their helps and Dr. A. Asadian and Mr. Moshrraf from Isfahan Agriculture Research center for sample preparation too.