Module 4 Lecture 4: Chemistry of Carbohydrates - Oligosaccharides, Polysaccharides, and Honey

Chemistry of Carbohydrates: Oligosaccharides and Polysaccharides, Case Study: Honey

Learning Objectives

  • Understand the mechanism of hemiacetal reaction to form acetals (covered in Lecture 2).
  • Understand that acetals are more stable than hemiacetals.
  • Recognize that oligosaccharides and polysaccharides are assembled via acetals, identify glycosidic bond(s), and name the type of glycosidic linkage(s).
  • Identify a reducing versus a non-reducing sugar (also in Lecture 3).
  • Recognize that different glycosidic bond connectivity can result in different carbohydrate structures/shapes, affecting physical properties and molecular recognition.
  • Apply previous knowledge (naming glycosidic linkages, formation of imines) to the case study of honey.

Hemiacetal to Acetal

  • Recap: Cyclic hemiacetals react with alcohols to form cyclic acetals.
    • Mechanism:
      • Hemiacetal + Alcohol → Acetal + Water
      • Example: \text{H3C-C(OH)(H3C) + CH3OH \rightarrow H3C-C(OCH3)(H3C) + H2O}
  • If "R-OH" is an alcohol from another sugar, then an acetal is formed, creating a glycosidic bond.
    • \text{C-H(O)(OH) + ROH \rightarrow C-H(O)(OR) + H2O}
    • Sugar 1 + Sugar 2

Glycosidic Bond

  • The new C-O bond formed is called a glycosidic bond or glycosidic linkage.
  • The acetal is called a “glycoside”.
  • Example with D-glucose and methanol:
    • α-D-glucopyranose (hemiacetal) + CH3OH → α-glycoside (acetal) + H2O
    • β-D-glucopyranose (hemiacetal) + CH3OH → β-glycoside (acetal) + H2O
    • \text{C6H12O6 + CH3OH \rightarrow C7H14O6 + H2O}
  • The acetal to hemiacetal equilibrium is reversible in the presence of a strong acid.
  • The alcohol nucleophile can attack either side of the carbocation intermediate forming both α and β glycosides (acetals) without ring opening.

Stability of Acetals

  • Hemiacetals exist in equilibrium with the open-chain monosaccharide form even in neutral aqueous conditions.
  • Acetals are stable in neutral conditions. A strong acid is required to convert between an acetal and hemiacetal.
  • Repeating sugar units linked by acetals are chemically stable in the absence of strong acids.
    • \text{C-H(O)(OR) + H2O \xrightarrow{H^+} C-H(O)(OH) + ROH}
  • Example: D-glucose and methanol
    • Once the α-glycoside is synthesized and purified, dissolving it in neutral aqueous conditions results in no reaction because it is very stable.
  • This glycoside is a non-reducing sugar because there is no hemiacetal, hence no aldehyde in equilibrium.

Higher Saccharides

  • Repeating units of sugars are joined by acetals, which contain a glycosidic bond.
  • Very stable chemically unless a strong acid is present.
  • In a biological setting, acetal hydrolysis is achieved using enzymes.
  • Terminology:
    • Monosaccharide: 1 sugar unit
    • Disaccharide: 2 sugar units
    • Trisaccharide: 3 sugar units
    • Oligosaccharide: 2-10 sugar units
    • Polysaccharide: >10 sugar units, often hundreds
  • Higher saccharides can be branched and have varied linkage stereochemistry.

Glycosidic Linkage – Stereochemistry and Connectivity

  • Two acetal anomers are chemically possible (α or β).
  • Depending on which sugar alcohol reacts, different bond connectivity is possible.
  • Examples:
    • β-1,4-glycosidic linkage: equatorial bond
    • α-1,4-glycosidic linkage: axial bond
  • Biosynthesis of oligosaccharides uses enzymes that selectively make only one isomer.

Glucose Disaccharides

  • Maltose: two D-glucopyranose units linked by an α-1,4-glycosidic bond.
  • Cellobiose: two D-glucopyranose units linked by a β-1,4-glycosidic bond.

Identifying Reducing Saccharides

  • A non-reducing sugar does not contain a hemiacetal.
  • Maltose is a reducing sugar because a hemiacetal will exist in equilibrium with the ring-opened form (aldehyde + alcohol).
  • The ‘silver mirror’ test will still come up positive due to the oxidation of the aldehyde to carboxylic acid.
  • Sucrose: D-glucopyranose and D-fructofuranose linked by an α-1,2-glycosidic bond.
    • It has two acetals and no hemiacetals and is a non-reducing sugar.

Polysaccharides

  • Amylose: D-glucose only linked by α-1,4-glycosidic bonds, up to 4,000 glucose units; it is one of the polysaccharides in starch.
  • High Fructose Corn Syrup (HFCS) is made from amylose from corn starch using enzymes and heat to isomerize D-glucose to D-fructose.
    • \text{Amylose \xrightarrow{Enzymes, Heat} D-glucose \xrightarrow{Isomerisation} D-fructose \rightarrow HFCS}
  • Cellulose: D-glucose linked by β-1,4-glycosidic bonds, up to 4,000 glucose units; it is a fibrous carbohydrate found in all plants and a structural component of plant cell walls.
    • Has a flat 3D shape and straight ‘molecular rods’ stabilized by a network of hydrogen bonds.
    • Good for structural material and strong (cotton is mostly cellulose).
    • Mammals without a rumen cannot digest cellulose. The human digestive enzyme α-amylase cannot break the β-1,4-glycosidic bonds.

Complex Carbohydrates

  • Gel extracts from Aotearoa’s native Mamaku (black tree fern) used as a rongoā rākau (plant remedy) to soothe and revive skin.
  • The gel extract contains mostly carbohydrates with repeating β-1,4-linkages and α-1,2-glycosidic linkages giving a unique shape and contributing to the gel-like consistency.

Carbohydrates and Molecular Recognition

  • Carbohydrates (via attachment to a protein or lipid) also form a cell surface molecular recognition code based on 3D shape.
  • There is a huge variety in 3D shape.
  • Each shape and color represents a different monosaccharide.
  • Each glycosidic bond can vary in linkage position (e.g., 1,4) and linkage stereochemistry (α or β) = many 3D shapes possible.
  • Many of the initial interactions between human cells and invading microorganisms are governed by cell surface carbohydrates.

Molecular Recognition - Blood Group Example

  • The blood groups A, B, and O have different carbohydrates on the surface of red blood cells.
  • This minor difference in carbohydrate ‘code’ has huge implications for antibodies recognizing foreign red blood cells.
  • Homework: Name the glycosidic linkages (e.g., α-1,3).

Case Study - Honey

  • What is in honey?
    • Mostly monosaccharides (glucose and fructose), smaller amounts of disaccharides and other compounds.
  • Honey bees collect nectar that contains hundreds of compounds including sucrose, that bees break down into monosaccharides using invertase.
    • \text{Sucrose \xrightarrow{Invertase} glucose + fructose}
    • Invertase, an enzyme, cleaves the α-1,2-glycosidic bond.
  • Honey contains a unique ‘chemical signature’ based on the plant’s nectar (including carbohydrates, natural products) and pollen, and is even season and region dependent.

Mānuka Honey

  • Mānuka is taonga (treasure) and has significant cultural heritage and connection to Māori, who are the kaitiaki of Mānuka.
  • Long before honey production, Mānuka extracts were used as a rongoā rākau.
  • Many ‘hive to shelf’ Aotearoa honey businesses incorporate mātauranga Māori (including plant knowledge, environmental stewardship, sustainability) and chemical analysis to produce the end honey product.
  • Chemical analysis includes measuring the sucrose, glucose, fructose levels unique to Mānuka honey, and the natural product leptosperin, a gentiobiose glycoside.
    • Gentiobiose is made up of 2 x D-glucose with a β-1,6-glycosidic linkage.
  • Another important component of Mānuka honey is methylglyoxal (MGO).

Methylglyoxal in Mānuka Honey

  • The level of methylglyoxal (MGO) is thought to contribute to the antimicrobial activity of the honey.
  • MGO is a very reactive compound; aldehydes and ketones can react with an amine to produce an imine, a common covalent reversible bond in the body.
    • Example: MGO can react twice, each time forming an imine, to join small ‘foreign’ compounds to larger proteins, which are recognized by and stimulate an immune response to an infection.

Adulterated Honey

  • Fake or adulterated honey is when colorants, sweeteners, or other substances that were not present when extracted from the beehive are added.
  • Adding ‘synthetic’ methylglyoxal is a concern for the Mānuka honey industry in Aotearoa.
  • Adding high fructose corn syrup (HFCS) is one of the common ’fake’ honey additives.
  • For interest only, not examined - the glucose:fructose ratio is actually very similar in honey and HFCS, so how do we tell a fake?
    • DNA testing: DNA from the plant’s pollen can be identified; HFCS contains small fragments of corn DNA that can be detected.
    • Analyze the carbon isotope ratio in the sugars: Glucose from corn starch has a different 12C13C\frac{12C}{13C} ratio then glucose from plant nectar.

Homework

  • Page 1329, exercise 28.36
  • Page 1330, exercise 28.54