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United States Department of Agriculture

Agricultural Research Service


Location: Cell Wall Biology and Utilization Research

Title: Efficacy of reducing sugar and phenol-sulfuric acid assays for analysis of soluble carbohydrates in feedstuffs)

item Hall, Mary beth

Submitted to: Animal Feed Science And Technology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 6/24/2013
Publication Date: 8/30/2013
Publication URL:
Citation: Hall, M. 2013. Efficacy of reducing sugar and phenol-sulfuric acid assays for analysis of soluble carbohydrates in feedstuffs. Animal Feed Science and Technology. 185:94-100.

Interpretive Summary: Water-soluble carbohydrates include sugars, oligosaccharides, and fructans, all of which can be excellent energy sources to dairy cows. Increasingly, nutritionists have been analyzing feedstuffs for these soluble carbohydrates to more accurately formulate diets for dairy cattle, but questions have come up regarding how well the assays measure this diverse set of carbohydrates, which vary by feedstuff. We evaluated how accurately various modifications of the assays could measure specific carbohydrates commonly in feeds. We found that the various methods were specialists: different methods were good at measuring different subsets of the water-soluble carbohydrates, but not all of them at once. This information will allow labs to tailor the analyses they do to the type of sample they are analyzing to provide the most accurate reflection of feed quality for more accurate diet formulation for livestock.

Technical Abstract: Reducing sugar (RSA) and phenol–sulfuric acid (PSA) assays are commonly used to analyze water-soluble carbohydrates. However, questions have arisen as to their accuracy for measurement of feedstuffs with diverse carbohydrate profiles. This study evaluated the efficacy of RSA and PSA as they would commonly be applied in feed analysis laboratories in measuring a variety of purified carbohydrates. Carbohydrates analyzed were glucose (Glc), fructose (Fru), galactose (Gal), sucrose (Suc), maltose (Mal), lactose (Lac), raffinose (Raf), and inulin (Inu). Variations on the methods used were PSA using Suc (PSA-Suc) or Glc (PSA-Glc) as standard sugars, and RSA with a 50:50 Glc:Fru blend as the standard with four hydrolysis methods: acid hydrolysis with 0.037 M sulfuric acid (RSA-H2SO4) or 0.5 M hydrochloric acid (RSA-HCl), or enzymatic hydrolysis with invertase (RSA-Inv) or an enzyme blend including sucrase, a-glucosidase, and beta-galactosidase (RSA-EnzBl). Recovery of carbohydrate was calculated on a dry matter (DM) basis as (carbohydrate detected g/kg DM)/(carbohydrate present kg/kg DM), with ‘close to’ complete recovery defined as values falling within the range of 920–1080 g/kg. Monosaccharide recovery did not differ between unhydrolyzed vs. hydrolyzed samples in RSA, indicating no destruction of carbohydrate by the hydrolysis method. For RSA, recoveries of Glc, Fru, and Gal were 979, 1042, and 706 g/kg, respectively. Such response differences among monosaccharides are inherent to RSA, and can affect carbohydrate recovery values. Methods that provided close to complete recovery by carbohydrate were: PSA-Suc and all RSA for Suc; PSA-Glc and RSA-EnzBl for Mal and Lac; PSA-Suc, RSA-H2SO4, RSA-HCl, and RSA-Inv for Raf; and RSA-H2SO4 and RS-HCl for Inu. None of the assays gave complete recovery of the diverse set of purified carbohydrates. Allowing a range of 920–1080 g/kg for recoveries on individual carbohydrates, RSA-H2SO4 and RSA-HCl would give the closest to complete recovery values for feeds such as forage and soybean in which Suc, Raf, and Inu were important. However, RSA-EnzBl would be useful in feeds such as forages or dairy products when Suc, Mal, and Lac are of interest. The allowed 920–1080 g/kg range of acceptable recoveries addresses the point that given very diverse carbohydrate complements of feeds, these assays will not be extremely precise, but may still be serviceable for diet formulation. The most accurate measurements will be achieved by selection of detection method, hydrolysis method, and carbohydrate standard to give greatest recovery of predominant carbohydrates in feedstuffs.

Last Modified: 8/24/2016
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