J . Sci. Food Agric. 1984,35,29-35

Carbohydrate and Phenolic Constituents in a Comprehensive Range of Rapeseed and Canola Fractions: Nutritional Significance for Animals."

Robert Blairb and Robert D. Reichert'

Department of Animal and Poultry Science, University of Saskatchewan, Saskatoon, Saskatchewan S7N OWO, Canada and National Research Council Prairie Regional Laboratory, Saskatoon, Saskatchewan, S7N OW9, Canada

(Manwcript received 14 April 1983)

Cotyledons from 11 varieties of rapeseed and canola seed were extracted with hexane and analysed for tannins, a-amylase inhibition value, starch and available carbohydrate by conventional methods. The sinapine content (total sinapic acid esters plus free sinapic acid) was determined by a spectrophotometric method. Samples of soya bean cotyledons, high-tannin sorghum grain and commercial rapeseed and canola meals were also analysed for comparison. Sinapine in rapeseed and canola cotyledons was measured at mean levels of 2.67 and 2.85% respectively. Tannins were detected in these cotyledons but only at low levels. The a-amylase inhibition values were also low or negative. Starch was detected in negligible amounts, and available carbohydrate at 13-14%. The predicted metabolisable energy (ME) value of commercial rapeseed or canola meal for poultry was found to be numerically close to the determined value when the available carbohydrate value was reduced to 35% of the observed value.

1. Introduction With the introduction of improved varieties of rapeseed containing low levels of glucosinolate and erucic acid (canola cultivars), attention should be paid to other components of the seed that may affect its nutritive value for animal feeding. These components include sinapine, tannins, fibre and carbohydrates.

The main phenolic constituent of rapeseed appears to be sinapine. '-'This compound is the choline ester of sinapic acid, and is present in Canadian rapeseed meals at levels of 1.0-2.5%.3 Sinapine has several undesirable properties as a constituent of animal feeds; it is a bitter-tasting compound, and its components may also contribute unpleasant flavours. It is also involved in the genesis of a fishy odour or taste in the eggs of certain brown-egg laying Tannins have also been linked to egg taint, through an adverse effect on hepatic oxidation of trimethylamine.6

Literature values for the tannin content of rapeseed meal range from 0.1% (dry matter basis)' to 3.0%,8 the variation in part depending on the analytical method used. Most of the tannin has been reported to be present in the According to a recent report" the tannin content of rapeseed and canola hulls is only 0.02-0.22%, which suggests fairly low levels of tannin in the meal. However, the study described in that report did not include the cotyledon, therefore no recent reports appear to be available on the tannin content of the fractions comprising rapeseed meal.

The high proportion of hull and contributed fibre in rapeseed meal results in a metabolisable energy (ME) value for animals that is lower than that of soya bean meal." Yellow-seeded cultivars are lower in crude fibre content13 and contain less lignin than the dark-hulled cultivars." It has been shown that protein digestibility in poultry is reduced when dark-coloured rapeseed hulls are included at a high level in the diet;14 their high lignin content may be responsible for this observation.

* Presented in part at the International Network of Feed Information Centres Second International Symposium, Sydney, Australia, 22-26 August 1982.

29

30 R. Blair and R. D. Reichert

Normally, the component of lipid-extracted seeds which contributes largely to ME is the available ~arbohydrate, '~ composed of sugars plus dextrins and starch. Prediction equations have been derived for the estimation of ME for poultry, based on the chemical composition." The relatively low ME value of rapeseed meal may be caused by its low available carbohydrate content, since it has been shown that starch is present only in negligible amount^.'^

The objective of this work was to determine comparatively the carbohydrate and phenolic contents of a range of rapeseed and canola varieties. The nutritional implications for animals are discussed, and breeding strategies for the improvement of canola are suggested.

2. Experimental

2.1. Materials Cotyledons from 11 varieties of rapeseed and canola seed were obtained by crushing and physical separation. The full fat cotyledon samples were then hexane extracted and ground in a Swedish-Ball tube, and dried. Samples of cotyledons from soya bean, high-tannin sorghum grain, and rapeseed and canola commercial meals were used for comparison of chemical composition. All assays were conducted in duplicate on one representative sample of each material.

2.2. Methods 2.2.1. Tannin content Defatted and full fat samples were analysed for extractable tannins by the vanillin-HCI, methanol extraction method" and by a modified vanillin-HC1, 1% HC1 in methanol extraction method," using catechin as the standard. Reagent blanks were used as described by Price et aL2'

2.2.2. Sinapine content The methanol extracts used for the tannin assay were diluted (1:50) and analysed for sinapine content using spectrophotometry. Crystalline sinapine bisulphate was used as a standard, and samples prepared in two different laboratories showed virtually identical spectroscopic properties. The concentration of sinapine in the methanol. extracts was calculated using the formula c=AIEZ, where c=concentration in mol litre-', A=absorbance at 330 nm, E=extinction coefficient at 330 nm and [=pathlength of the spectroscopic cell. The extinction coefficient of sinapine in methanol was found to be 21 390. The sinapine content determined by this procedure includes all sinapic acid esters plus free sinapic acid.

2.2.3. Other components a-Amylase inhibition assay was conducted by the method of Daiber.21 Starch values were determined by the dual-enzyme semi-micro method of Banks et a1.,22 and the available carbohydrate content was determined by the method of Bolton."

3. Results

The results shown in Tables 1 and 2 are all on a dry matter basis. The table also includes data of Mitaru et ul." on hull composition.

3.1. Tannin Tannins were detected in rapeseed and canola cotyledons, but only at low levels. The a-amylase inhibition values were also low or negative. Tannin values obtained with the modified vanillin method were higher than those found with the unmodified vanillin method. It was found that sinapine did not react with the vanillin-HC1 reagent to form a colour, and therefore did not interfere with the tannin analysis. Canola cotyledons appeared to contain somewhat higher tannin levels than rapeseed cotyledons; the mean values with the vanillin and modified vanillin methods were 0.10 and 0.36% (defatted rapeseed) and 0.15 and 0.53% (defatted canola). The yellow-hulled rapeseed, cv. R500, showed the lowest tannin content.

Carbohydrate and phenolic constituents of rapeseed and canola 31

Table 1. Levels of phenolic constituents in canola and rapeseed mealP

Tannin content a-amylase Sinapined (% DM) inhibition(%) (% DM)

m o d i f i e d vanillin vanillin method method

Rapeseed cotyledons Midasb

Targetb

Zephyrb

Polar'

Span'

TorchC

R500'

full fat defatted full fat defatted full fat defatted full fat defatted full fat defatted full fat defatted full fat defatted

0.05 0.18 0.11 0.41 0.03 0.17 0.07 0.36 0.09 0.19 0.18 0.39 0.06 0.18 0.13 0.39 0.04 0.20 0.08 0.36 0.05 0.22 0.10 0.43 0.005 0.09 0.01 0.18

n.a. 2.4 n.a.

n.a.

n.a.

n.a. -2.5 n.a. -3.4 n.a.

0.1

-2.3

-4.1

2.4

Canola cotyledons Altexh

Regentb

Tower'

Candle"

full fat defatted full fat defatted full fat defatted full fat defatted

0.09 0.23 0.20 0.54 0.08 0.24 0.16 0.50 0.08 0.27 0.17 0.62 0.03 0.23 0.07 0.46

n.a. -7.9 n.a.

0.3 n.a.

ma. -2.9

-4.0

1.8 4.0 1.2 2.5 1.7 3.5 1.1 2.4 1.3 2.3 1.3 2.6 0.7 1.4

1.4 3.3 1.3 2.7 1.3 2.9 1.3 2.5

-

Soya bean cotyledons full fat 0.00 0.00 ma. n.a. defatted 0.00 0.00 -13.6 n.a.

Sorghum grain P570 full fat 3.8 3.7 100.0 n.a. AKS614 full fat 2.1 1.3 86.6 0.09' AR3003 full fat 1.2 1.3 50.2 0.07'

Commercial meals Canola Regent Soya bean

ma. n.a. n.a. n.a. n.a. -1.1 n.a. n.a. n.a. n.a. n.a.

Hulls" Rapeseed (Torch') Rapeseed (R500') Canola (Regentb) Soya bean

0.02 0.09 0.00 ma. 0.10 0.06 0.00 ma. 0.11 0.22 0.00 n.a. 0.00 0.00 0.00 ma.

'mean of duplicate analyses of the same sample. 'Brarsica naplrs. 'B . campestris. dincludes all sinapic acid esters plus free sinapic acid 'probably not sinapine. DM=dry matter. n.a. =not assayed.

3.2. Sinapine It was found that the spectroscopic properties of sinapine bisulphate in methanol and the methanol extracts of canola and rapeseed varieties were very similar in the range 250-600 nm, suggesting that spectrophotometry could be used to analyse for sinapine content. A wavelength scan showed a very defined absorption maximum at 330 nm which was used for quantification. Krygier et a1.2 have shown that sinapine constitutes more than 98% of the phenolics in canola, and therefore the contribution of interfering phenolics to the absorption at 330 nm was considered minimal.

32 R. Blair and R. D. Reichert

Table 2. Levels of carbohydrate constituents in canola and rapeseed meals'

ADF ADL Starch AC (%DM) ( % D M ) (% DM) ( % D M )

Rapeseed cotyledons Midasb full fat

Targetb full fat

Zephyr' full fat

Polar' full fat

Span' full fat

Torch' full fat

R500' full fat

defatted

defatted

defatted

defatted

defatted

defatted

defatted

n.a. ma. n.a. n.a. n.a. 0.05 n.a. ma. n.a. n.a. n.a. 0.01 n.a. n.a. n.a. n.a. ma. 0.13 ma. n.a. n.a. n.a. n.a. 0.001 n.a. n.a. ma. ma. n.a. 0.07 ma. n.a. n.a. n.a. n.a. 0.05 n.a. n.a. ma. ma. ma. 0.08

n.a. 14.1 n.a. 14.0 ma. 12.5 n.a. 14.9 n.a. 13.9 n.a. 13.9 n.a. 15.0

Canola cotyledons Altexb full fat

Regentb full fat

Towerb full fat

Candle' full fat

defatted

defatted

defatted

defatted

n.a. n.a. n.a. n.a. n.a. 0.16 n.a. n.a. n.a. n.a. ma. 0.08 n.a. n.a. n.a. n.a. n.a. 0.09 n.a. ma. n.a. ma. n.a. 0.11

n.a. 13.3 n.a. 12.1 n.a. 13.4 n.a. 14.5

Soya bean cotyledons full fat n.a. n.a. n.a. n.a. defatted n.a. ma. 0 35 19.4

Sorghum grain P570 full fat n.a. n.a. 66.5 n.a. AKS614 full fat n.a. n.a. 64.1 n.a. AR3003 full fat n.a. n.a. 66.3 n.a.

Commercial meals Canola Regent Soya bean

n.a. n.a. n.a. 9.3 n.a. n.a. n.a. 10.3 n.a. n.a. n.a. 17.9

Hulls" Rapeseed (Torch') Rapeseed (R500') Canola (Regent') Soya bean

57.3 28.8 n.a. n.a. 34.1 5.5 n.a. n.a. 51.9 26.6 n.a. 4.8 51.2 1.3 n.a. 4.9

ADF=acid detergent fibre. ADL=acid detergent lignin. AC=available carbohydrate. DM=dry matter. n.a. =not assayed. "mean of duplicate analyses of the same sample. bB. napus. 'B. campestris.

Sinapine was detected in high concentration in rapeseed and canola cotyledons, and at low or zero levels in soya bean cotyledons and sorghum grain. The mean content of sinapine in defatted rapeseed cotyledons was 2.7% (range 2.5-3.3%).

3.3. Other components Starch was detected only in trace quantities in the rapeseed and canola cotyledons, although some available carbohydrate was found. The mean available carbohydrate content of defatted rapeseed cotyledons was found to be 14.0% and in defatted canola cotyledon it was 13.3%. The

Carbohydrate and phenolic constituents of rapeseed and canola 33

corresponding values obtained for commercial meals were 9.3% (canola), 10.3% (rapeseed) and 17.9% (soya bean meal).

4. Discussion and conclusions The values obtained for the sinapine content of rapeseed and canola cotyledons were slightly higher than those reported by Krygier et a1.,2 and correspond with values obtained by Mueller et aL3 for rapeseed meals. The variation in sinapine content appeared to be greater in rapeseed cotyledons (range 1.4-4.0% in defatted samples) than in canola cotyledons (range 2.5-3.3% in defatted samples), although fewer samples of canola were assayed. The sinapine assay method used by Krygier et aL2 involved extraction of defatted ground samples with methanol-acetone, and quantitation by gas-liquid chromatography (g.1.c.). The spectrophotometric method used in this investigation may therefore be a suitable alternative method to the g.1.c. method.

It appears that both rapeseed and canola cotyledons contain tannins, although the a-amylase inhibition values were low or even negative, indicating the absence of tannins capable of inhibiting this enzyme. Slightly higher values for extractable tannin content were found for canola than for rapeseed cotyledons, although fewer canola samples were assayed. The values obtained for cotyledons were higher than those reported for hulls by Mitaru et al.," which does not agree with Durkeeg and Leung et al. lo that most of the extractable condensed tannins in rapeseed meal are found in the hull. However, the overall significance of tannins present in canola and rapeseed meals is that they are probably of minor nutritional importance. The findings of Mitaru et al.14 on growth and protein digestibility in chickens fed high dietary levels of rapeseed and canola hull tannins confirm this conclusion.

Soya bean cotyledons showed a zero tannin value. Mitaru et al. '' also reported a zero value for soya bean hulls. The high-tannin sorghum varieties tested showed expected values for tannin content, which corroborated the a-amylase inhibition values obtained.

Starch was found only in trace amounts in rapeseed and canola cotyledons, which agrees with the findings reviewed by Bell.17 However, available carbohydrate was detected in rapeseed and canola cotyledons and meals at levels consistent with other report^.^^-'^ The main components of the available carbohydrate fraction appear to be sugars such as sucrose, stachyose, D-fructose and

Since commercial rapeseed and canola meals contain low levels of lipid (because of extraction) and have been found to contain low levels of available carbohydrate, they cannot be expected to provide a high level of energy to non-ruminant animals. Recent Canadian estimates" of the ME value of canola meal for poultry are 7.95 MJ kg-' for growing chickens and 8.37 MJ kg-' for adult chickens (air-dry basis). The formula of Bolton16may be used to predict the ME value of canola meal for chickens, based on the content of crude protein, ether extract and available carbohydrate. Using an available carbohydrate value of lO.O%, together with mean values of 38.0% for crude protein and 3.0% for ether extract, gives a predicted value of 8.78 MJ kg-' for chicks and 9.19 MJ kg-' for adult chickens. Therefore, the formula overestimates the ME value by about 10.0%. It has been found with rapeseed meal that the chemical method overestimated the value of available carbohydrate to the and that a chemical value of 11.6% was equivalent to a chick bioassay value of 4.0% (i.e. 35% of total). Substitution of the value of 3.5% available carbohydrate in place of 10.0% in the Bolton formula results in predicted ME values of 7.68 MJ kg-' for the chick and 8.10 MJ kg-' for the adult fowl, which are much closer to the determined values.

According to previous values," the acid-detergent lignin (ADL) concentration appears to be much higher in the dark-hulled varieties of rapeseed and canola than in the yellow-hulled cv. R500 or in soya bean hulls. Mitaru et al. l4 have shown that protein digestibility is reduced when dark-hulled canola hulls are included at a high level in the diet of pigs, suggesting that the ADL content may be detrimental.

The results on sinapine content suggest that canola meal intended for animal feeding should be improved by attempting to develop cultivars with a reduced content of sinapine. As an interim measure, the processing of meal to destroy sinapine should be considered, since various processing techniques (e.g. ammoniation) have been shown to be beneficial in removing

D-glucose.27

3

34 R. Blair and R. D. Reichert

The low ME level in canola meal (relative to soya bean meal) is consistent with low available carbohydrate and relatively high hull contents. The ME content of canola meal would be improved by developing cultivars with a reduced hull content and with an increased content of available carbohydrate in the cotyledons.

Attention should also be paid by plant breeders to the chemical composition of the hull fibre fraction. Findings that fibres containing lignin have an adverse effect on protein and amino acid digestibility in non-ruminant animals14230331 suggest that the lignin content of hulls should be minimised. This would entail selection of yellow-hulled varieties, which have the added advantage of a lower fibre content.

Acknowledgments

The authors are grateful to Dr D. I. McGregor and Dr K. Dabrowski for supplying the samples of pure sinapine bisulphate, and Don Schwab and Kate Goldsmith for technical assistance. The assistance of Mrs Janet Jones in the preparation of this paper is gratefully acknowledged.

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