Determination of carbohydrates

Contents Introduction Distribution of carbohydrates in food Definition Classification Structure of carbohydrates Distribution of carbohydrates in food Function of carbohydrates in food processing Determination of carbohydrates

Introduction Definition Chemically, carbohydrates are organic molecules in which carbon, hydrogen and oxygen bond together in the ratio: Cx(H2O)y where x and y are whole numbers that differ depending on the specific carbohydrate to which we are referring 多羟基醛或者酮及其衍生物和缩合物 Carbohydrates are the main energy source for the human body. Carbohydrates are the most abundant class of organic compounds found in living organisms.

Classification（从组成上分） Simple Sugars Monosaccharides:不能再被水解的多羟基醛或者酮，是碳水化合物的基本单位。如葡萄糖、果糖、半乳糖等。 Complex Carbohydrates Oligosaccharides ：2～10个单糖分子缩合而成，水解后生成单糖。 如蔗糖、麦芽糖、乳糖等。 Polysaccharides：由许多单糖分子缩合而成，如淀粉、纤维素、果胶等。

Monosaccharides Two of the most common monosaccharides are glucose and fructose. Both glucose and fructose have the same chemical formula (C6H12O6) however they have different structures as seen below

Fructose Glucose

Disaccharides have two sugar units bonded together. For example, common table sugar is sucrose a disaccharide that consists of a glucose unit bonded to a fructose unit: 

Lactose Lactose: is found exclusively in the milk of mammals and consists of galactose and glucose in a b-(1,4) glycosidic bond

maltose Maltose: the major degradation product of starch, is composed of 2 glucose monomers in an a-(1,4) glycosidic bond

Complex Carbohydrates Definition polymers of the simple sugars.  long chains of simple sugar units bonded together ，often referred to as polysaccharides).  Several polysaccharides Starch Cellulose pectin

amylose

Cellulose

Characteristic of monosaccharides Another characteristic of monosaccharides is that they can act as mild reducing agents. This is because the aldehydo group that is present can be oxidized to form a carboxylic acid group, or in the presence of a base, a carboxylate ion group.

Fructose can also act as a reducing sugar, even though it has a ketone group instead of an aldehyde group. Under basic conditions, the fructose molecules can, essentially, have the location of the carbonyl bond switched to convert them into a glucose molecule. This occurs in a number of steps involving removing hydrogens from the #1-C and its oxygen and moving them to the #2-C and its oxygen.

Equilibrium of Ring form and open form any monosaccharide molecule that's in a ring form will, within a fraction of a second, be in the open form and, thus, be able to react with the oxidizing agent and reduce it. Equilibrium of Ring form and open form reducing sugars not reducing sugars

Characteristics of starch Insoluble in hot water Blue complex compound by reaction with iodine. Insoluble in 30% of ethanol solution or above. Hydrolyzed by acid and enzyme, the ultimate product is glucose. Specific rotatory power is between +201.5 and +205

Characteristics of cellulose Insoluble in water No reducibility(无还原性) Uneasy to be hydrolyzed.concentrated H2SO4 is needed in the hydrolysis of cellulose for a long time at high temperature.(水解比淀粉困难得多，需用浓硫酸在一定时间下长时间加热水解) Can not be digested by body, but can be used by some animal, such as cows.(人体不能产生分解纤维素的酶，一些草食动物可以消化纤维素。 )

Distribution of carbohydrates in food Carbohydrates comprise more than 75% of the dry matter of plants, corn ,vegetable, fruits and so on. Monosacchrides and oligosaccharides is usually found in vegetables and fruits Polysaccharides can mainly be found in corn, seed, root and so on.

Function of carbohydrates in food processing texture of food mouth-feel of food Sweet taste Flavor for food

Determination of carbohydrates Determination of reducing sugar Determination of sucrose Determination of total sugar Quantitative determination of different carbohydrates Determination of starch Determination of cellulose Determination of pectin

Determination of reducing sugar Preparation of samples 40-50℃， soluble sugar water Above 40-50℃ ,Soluble sugar, starch,dextrin, etc 70-75% ethanol—protein,dextrin,starch can’t be soluble Sample containing oil—extract fats, then separate sugar Adding clarifier to remove the interruption(no effect on sugar)

Determination of reducing sugar Reducing Sugars carboxylate Red precipitate aldehyde

Common test reagents are : Benedicts reagent (CuSO4 / citrate) Sugars that contain aldehyde groups that are oxidised to carboxylic acids are classified as reducing sugars. Common test reagents are : Benedicts reagent (CuSO4 / citrate) Fehlings reagent (CuSO4 / tartrate) They are classified as reducing sugars since they reduce the Cu2+ to Cu+  which forms as a red precipitate, copper (I) oxide. In order for oxidation to occur, the cyclic form must first ring-open to give the reactive aldehyde. So any sugar that contains a hemi-acetal will be a reducing sugar. But glycosides which are acetals(乙缩醛） are not reducing sugars.

hemi-acetal => reducing sugar b-D-glucose methyl b-d-glucopyranoside acetals => non-reducing sugar sucrose

碱性铜盐法 原理： 氧化还原反应－滴定法－次甲基蓝为指示剂 注意： 1）已知的还原糖溶液标定碱性酒石酸铜溶液 2）在酒石酸铜溶液中加入少量亚铁氰化钾，络合氧化亚铜 3）沸腾条件下进行滴定 4）快速完成测定

3，5－二硝基水杨酸比色法测定还原糖 原理： 3，5-二硝基水杨酸溶液与还原糖(各种单糖和麦芽糖)溶液共热后被还原成棕红色的氨基化合物，在一定范围内，还原糖的量和棕红色物质的颜色深浅的程度成一定比例关系。在540nM波长下测定棕红色物质的消光值，查标准曲线，便可求出样品中还原糖的含量。

酶－比色法 酶－电极法 葡萄糖氧化酶(GOD)在有氧条件下， 催化β-D-葡萄糖(葡萄糖水溶液)氧化， 受过氧化物酶(POD)催化，过氧化氢 与4-氨基安替比林和苯酚生成红色醌亚胺。在505nm波 长处测醌亚胺的吸光度， 酶－电极法 葡萄糖氧化酶(GOD)在有氧条件下催化β─D─葡萄糖(葡萄糖水溶液状态)氧化，生成D─葡萄糖酸─δ─内酯和过氧化氢。过氧化氢与过氧化氢型电极接触产生电流。该电流 值与β─D─葡萄糖的浓度呈线性比例，在酶电极葡萄糖分析仪上直接显示葡萄糖含量。

Determination of sucrose Principle Sucrose----monosaccharides—determine Acid treatment Enzyme treatment

Determination of total sugar Definition Including reducing sugar and sucrose that can be hydrolyzed to reducing sugar at the conditions. Principles As above Methods 蒽酮比色法

Contents Determination of starch Determination of cellulose Determination of pectin

Determination of starch by acid or enzyme hydrolysis Principle Acid amylase Remove impurity Sample with Starch Samples Remove lipids Remove soluble sugar Starch hydrolysis Ethyl ether 碱性铜盐法 Reducing sugar ethanol 酶－比色法 蒽酮比色法

Attention Samples—remove lipids by ethyl ether Samples– remove soluble sugar by 80% of ethanol(dextrin is reserved), if the ethanol concentration is 10%, dextrin is removed with soluble sugar together. Kept starch hydrolyzed completely by controlling the experiments strictly. It can be indicated by iodine.

Protein precipitated by SnCl2 Polarimetry Principle Starch extraction by CaCl2-HAC solution Protein precipitated by SnCl2 Determination of α Calculation

Filtering, collecting precipitate Weighing method Principle Sample + KOH in ethanol filtering Starch 、cellulose Adding KOH in water Calculate the loss filtering Burning at 550 ℃ cellulose supernate HAC-ethanol Filtering, collecting precipitate Drying at 100℃

Determination of amylose Principle Starch + iodine Complex with color Mixing the different concentration of amylose and amylopectin at definite content of total starch 620nm, determine OD Calibration curve

淀粉是葡萄糖的聚合物，由单体葡萄糖通过糖苷键的形式连接而成，线性连接的分子是直链淀粉，分支连接的则为支链淀粉，单纯的直链和支链是不存在的，淀粉分子实际上是由两种形式的混合物构成的. 淀粉与水构成的混悬液在受热的情况下会发生一定的变化。 在较低的温度条件下，淀粉通过氢键作用结合水分子而分散，此时的变化属于物理变化，淀粉的结构不发生变化。 当温度上升到一定程度后，淀粉分子会大量吸收水分而发生急剧膨胀，分子结构发生伸展，此时即使停止加热，分子也不能恢复原来的结构。 随着温度的不断上升，膨胀的程度加大，这就是所谓的糊化，糊化的本质是高能量的热水破坏了淀粉分子内部彼此之间的氢键结合，使分子的混乱度增大，糊化后的淀粉一水体系的行为直接表现为粘度的增加。如果使体系的温度降低，那么随着水分子能量的减小，淀粉分子之间趋向于重新集结，但是不能恢复原有的结构，如果体系的浓度较小，体系变成溶胶；如果浓度较大，体系则成为凝胶。

糊化度测定 原理 淀粉－－淀粉酶作用－还原糖－－测定。

粗纤维测定 性质 人体消化系统所不能消化、分解的物质。包括纤维素、半纤维素等。 稳定，机械强度大，不溶于水，不溶于弱 酸、弱碱，难水解，无还原性。 纤维素－－浓HCl或80%以上浓H2SO4作用－－D－葡萄糖 纤维素――纤维素水解酶作用-- 纤维二糖-- β－水解酶作用 --D －葡萄糖

粗纤维测定 称量法 样品 热稀H2SO4 糖、淀粉、果胶等水解 热KOH醇溶液 蛋白质溶解、脂肪皂化 石油醚 脱脂、色素等 粗纤维 纤维素测定仪

不溶性膳食纤维测定 不溶性膳食纤维：来源于各类植物性食物中不溶于水的半纤维素、半纤维素、木素。 水溶性膳食纤维：指溶解于水的膳食纤维，如水果的果胶、海藻的藻胶 食品中水溶解性膳食纤维含量很少。 测定： 样品－－中性洗涤剂溶液浸煮－－过滤－－热蒸馏水洗涤残渣－－淀粉酶作用除去结合态淀粉－－丙酮等除去色素、脂肪等－－烘干残渣－－不溶性膳食纤维

果胶测定 果胶定义：半乳糖醛酸以α－1，4糖苷键聚合形成的多聚半乳糖醛酸。部分羧基被甲基酯化，部分与离子结合。 果胶的存在形式 原果胶：不溶解于水 原果胶酶或者酸作用 果胶酯酸：溶于水 果胶酶或者酸、碱作用 果胶酸：溶于水，可与离子结合为盐的形式存在。

测定方法 称量法： 比色法： 样品－除去脂肪、可溶性糖、色素等干扰因素－酸或者水提取果胶－果胶酸钙－烘干称重 样品－水解产生半乳糖醛酸－在强酸条件下，与咔唑试剂反应显色－测定OD，与标准曲线对照－计算

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