Chemistry:

is the study of matter, including its composition, structure, physical

properties, and reactivity. It is divided into five fields: organic, inorganic,

physical,

biochemical, and analytical

الكيمياء: هي دراسة المادة ، بما في ذلك تكوينها وبنيتها وخواصها الفيزيائية وتفاعلها. وهي مقسمة إلى خمسة مجالات: عضوية ، وغير عضوية ، وفيزيائية ، وكيميائية حيوية ، وتحليلية.

1

1-

Beaker كأس

2-

Watch glass ساعة زجاجه

3-

Washing bottles غسل قنينة

4-

Conical flask حجمية قنينة

5-

Graduated cylinder مدرجة إسطوانة

6-

Test tube إختبار إنبوبة

7-

Brush

فرشاة

Chemistry:

is the study of matter, including its composition, structure, physical

properties, and reactivity. It is divided into five fields: organic, inorganic,

physical,

biochemical, and analytical.

Laboratory Equipment 8-

Funnel قمع

9-

Separating funnel

فصل قمع

10-

Dropper قطارة

11-

Rack حامل

12-

Pipettes ماصة

13-

Burette سحاحة

14-

Volumetric flask حجمية قنينة

15-

Spatula ملعقة

16-

Burette clamp and stand سحاحة وماسك حامل 3

Basic Laboratory Safety

1-

Personal safety

2-

Eye safety

3-

Handling of needles and sharps

4-

Handling of biologically hazardous material

5-

Material Safety Data Sheet (MSDS)

Balances :- used to weight the chemical reagent .

a-

Rough balance b-Analytical balance

In case of Accident

Accident Needle Stick

1-

Bleed wound

2-

Wash wound thoroughly with soap

3-

Notify the supervisor of the incident

4-

May need to get blood tested for hepatitis

Analytical chemistry consists of:

1-

(A) Qualitative analysis which deals with the identification of elements,

ions, or compounds present in a sample (tells us what chemicals are present in a

sample).

2-

(B) Quantitative analysis which is dealing with the determination of how

much of one or more constituents is present (tells how much amounts of

chemicals are present in a sample).Carbohydrates

Carbohydrates:- are aldihydic or ketonic multy hydroxylic compound, which

gives when hydrolyses multy hydroxyl alcoholic aldihyde or ketone, its general

structure (CH2O)n , when n about three to many thousands, and can be devided

into:-

1- Monosaccharides: Is a one molecule saccharide cannot be divided to smaller

parts.

2- Disaccharide: is a saccharide consist of tow monosaccharides linked by

glycosidic (etheric) bond( C-O-C) .

3- Oligosaccharides: a saccharide consist of (3-10) units of monosaccharide

linked by a glycosidic etheric bond C-O-C as: Raffinose, Maltotriose.

4- Polysaccharides: is asaccharides with high molecular weight , consist of a

large number of monosaccharide linked by a glycosidic etheric bond C-O-C

as:starch, cellulose, in plant and glycogen in human and animals. And the

main role for polysaccharides is to store energy. All polysaccharides are non

reducing sugar because of their high molecular weight, although containing

free carbonyl group.5

Some important points

1- Most of the tests and reactions described are not quantitative and volumes are

approximate, despite these facts some tests do not work if quantities greatly in

excess of those stated are used.

2- DO NOT place your pipette in reagent bottle as this lead to contamination.

3- In most tests, it is important to apply a control test using water instead of the

solution under examination. If you are in doubt about the result of a test,

perform the reaction with a suitable known compound.

4- In this experiment, sugar sample are given in their solid state. To perform each

procedure, you should prepare your own sugar solution by taking very small

amountof solid sugars.

5- When you need to boil your sample in a test tube, prepare a hot water in large

beaker and put your test tube inside the beaker. DO NOT forget to put boiling

chips in the beaker.

A carbohydrates is an organic compounds with the general formula (CH2O)n, that

is, consist only of carbon, hydrogen and oxygen as the structure above.

Carbohydrates make up the bulk of organic substances on earth and perform

numerous roles in living things.

The carbohydrates (saccharides) are divided into four chemical groups:

monosaccharides, disaccharides, oligosaccharides, and polysaccharides.

Polysaccharides serve for the storage of energy (e.g.,starch in plants and

glycogen in animals) and as structural compounds (e.g., cellulose in plant and

chitin in arthropods).structural polysaccharides are frequently found in combination with proteins

(glycoproteins or mucoproteins) or lipids (lipopolysaccharides).

The 5-carbon monosaccharide ribose is an important compound of coenzymes

(e.g.,ATP,FAD and NAD) and the backbone of the genetic molecules known as

RNA.

The related deoxyribose is a component of the DNA. Saccharides and their

derivatives include many other important biomolecules that play key roles in the

immune system, fertilization,preventing pathogenesis, blood clotting and

development. This experiment aims to introduce you with the identification of

unknown carbohydrates. To gain maximum benefit, observation should be

related, as far as possible, to the structure of the substance examined

Source of carbohydrates:

1- Rice , corn and potatoes.

2- Sucrose (cane and beet sugar).

3- Lactose (milk and milk products).

4- Glucose (honey , corn syrup and fruits).

5- Fructose (fruits , honey).

Metabolism of Glucose :

Glycogenesis : conversion glucose

glycogen .

Glycogenolysis : breakdown glycogen

glucose and other

intermediate products.7

Glycolysis : the oxidation of glucose to lactate or pyruvate .

Gluconeogenesis : formation glucose from non – carbohydrate sources (amino

acids and fatty acids).

Citric acid cycle : the final common pathway of oxidation of carbohydrates .

lipids and proteins , through which acetyl CoA is completely oxidized to CO2 and

H2O .

The hormones are important in the regulation of blood glucose

concentration :

1.

Insulin.

2.

Growth hormone.

3.

Hydrocortisone.

4.

Epinephrine .

5.

Glucagon .

6.

Thyroxin .

Clinical significance ꞉

The most common disease related to carbohydrate metabolism is diabetes

mellitus which characterized by insufficient blood level of active insulin .

Deficiency of insulin is result in inability of glucose to enter muscle and liver

cellsImportant Notes :

FBS : Fasting blood sugar.

RBS : Random blood sugar.

PPG : Two hours postprandial glucose.

The normal Cerebrospinal fluid (CS F) glucose concentration ranges from 40 -

70mg/dl

Glycolysis should be avoided and its prevented by fluoride-oxalate.

Glucose is more stable in plasma when blood is drawn , permitted to clot and

to stand uncentrifuged at room temperature . The average

rate of decrease in serum glucose is approximately 7% in one hour.

Principle꞉

Glucose oxidase (GOD) catalysis the oxidation of glucose to gluconic acid

.The formed hydrogen peroxide (H2O2) , is detected by a chromogenic oxygen

acceptor phenol –aminophenazone in the presence of peroxidase (POD).Qualitative tests for Carbohydrates

1- Molisch`s test :-

Purpose: - General test for all carbohydrates.

Reagent: - α - naphthol (1% in ethanol), Conc.H2SO4.

Principle: - Conc. H2SO4 Hydrolyze glycosidic bonds to give the

monosaccharides which are then dehydrated to Furfural and its derivatives. These

products then combine with sulphonated

α - naphthol to give a purple ring.

Procedure:-

1-

Place 2 mL of a known carbohydrate solution in a test tube, add 1 drop of

Molisch`s reagent (1% α – naphthol in ethanol).

2-

Pour 1-2 mL of Conc. H2SO4 down the side of the test tube, so that it

forms a layer at the bottom of the tube.

3-

Observe the color at the interface between two layers and compare your

result with a control test.

*A brown color due to charring must be ignored and test should be

repeated with a more dilute sugar solutionQuestions:-

-Write the reaction equation for molisch`s test.

-Give an example of a carbohydrate structure that would give positive test with

Molisch`s reagent.

2- Benedict`s test :-

Purpose:- General test for reducing sugars. Benedict`s reagent:- (Cu2+ /weakly

alkaline medium).

Principle :- Oxidation-reduction will occur between reducing sugar and Cu2+ in

weakly alkaline medium to form an orange or red precipitate(Cu2O).Reducing

sugar will oxidize to acidic sugar while the cupric ion will reduce to cuprous.

Qualitative tests for Carbohydrates Molisch`s test :-

Purpose: - General test for all carbohydrates.

Reagent: - α - naphthol (1% in ethanol), Conc.H2SO4.

Principle: - Conc. H2SO4 Hydrolyze glycosidic bonds to give the

monosaccharides which are then dehydrated to Furfural and its derivatives. These

products then combine with sulphonated

α - naphthol to give a purple ring.2- Iodine test

Purpose: - Iodine test for polysacchrides such as starch, glycogen, cellulose and

dextrin.

Reagent: - Iodine solution can be prepared by dissolving elemental iodine in an

aqueous solution of potassium iodide(KI).

Principle:- Iodine forms colored adsorption complexes with polysaccharides,

starch gives a blue color with I2 , While glycogen gives red-brown color.

Procedure:-

4drops of polysaccharide+ 2drops of iodine solution

See the color

Starch I2

blue dark Glycogen

I2

red brown Dextrine

I2

russet Water

I2

yellow

Condition of reaction:-

1-The reaction does not require heating, because the heat lead to the

volatilization of I2. And Cannot be performed at very low pH due to the

hydrolysis of the starch

2-The reaction needs to acidic medium or natural, because the basic medium

converts the iodine into ionic form (I- ), thus the reaction do not succeed, as the

reaction direction will go as the equation below Procedure :-

Approximately 1 ml of sample is placed into a clean test tube. 2 ml (10 drops)

ofBenedict's

reagent (CuSO4) is placed in the test tube. The solution is then heated in a

boiling water bath for 3-5 minutes. Observe for color change in the solution and

precipitate.

Important points :-

*carbohydrates with a free or partially free aldehyde or ketone group have

reducing properties.

*All monosaccharides are reducing.

*All disaccharides are reducing except sucrose (because carbonyl group –

responsible of reduction- is associated).

*All poly saccharides are non-reducing.

How to prepare Benedict`s reagent:-

One liter ofBenedict’ssolution can be prepared from100 g of anhydrous sodium carbonate,

173 g of sodium citrate and 17.3 g of copper(II) sulfate pentahydrate.

Questions:-

Give the structure of sodium citrate. What is the medium for benedict`s test?

Why monosaccharides and some disaccharides are reducing, while sucrose and

poly saccharides are not?4- Barfoed`s test

Purpose:- Test for reducing monosaccharides.

Barfoed`s reagent:- (Cu2+ / weakly acidic medium).

Principle:- Barfoed`s reagent is Cu2+ in weakly acidic medium and is only

reducedby monosaccharides because disaccharides is weaker than

monosaccharides in reducing ability. The precipitate of Cu2O is less dense than

with benedict`s test , and its color is abrike red.

Procedure:- 8drops glucose +16drop Barfoed`s reagent

3 min Brike – Red

SOME IMPORTANT POINTS TO KNOW:-

Avoid prolonged boiling, because disaccharides may hydrolyse to

monosaccharide, and give a false positive test.

How to prepare Barfoed`s reagent?

Dissolve 13.3 g of cupper acetate in 200 mL of distilled water and add 1.8 mL of

1 % acetic acid to it.

When you test starch with Barfoed`s reagent, what would be the answer, positive

or negative? Explain your answer by giving reasons and structures.

What is the use of acetic acid?5 Bial`s test

Purpose:- Test for pentoses sugars.

Bial`sreagent:- Concentrated Hydrochloric acid Conc. HCl (500 mL), Orcinol

(1.5 g),Ferric Chloride FeCl3 (1 mL 10 %).

Principle:- When pentoses are heated with Conc. HCl Furfural is formed which

condenses with Orcinol in the presence of (Fe3+) to give a blue – green color.

While hexoses give negative result because pentoses is faster than hexoses in

formation of furfural.

Procedure:- 4 drops xylose + 8 drops Bial`s reagent

3 min

blue – green

complex

* Avoid prolonged boiling, because hexoses yields hydroxymethylfurfural which

also reacts with orcinol to give brown complex (false positive test).

6Seliwanoff`s test

Purpose:- Test for ketoses sugars.

Seliwanoff`s reagent:- Hydrochloric acid HCl (3M),Resorcinol (0.05 g).

Principle:- Ketoses are dehydrated by Conc. HCl more rapidly than aldoses to

give furfural derivatives which then condenses with resorcinol to form red

complex.

Procedure:- 8 drops fructose + 16 drop seliwanoff`s reagent

7Osazones test :-Purpose:- This test used to recognize between the reducing sugar from the same

class.

For examples:-

Glucose, mannose, galactose.

All of them are reducing monosaccharide (hexoses), and they all give positive

result with:-

Molisch`s , Benedict`s, Barfoed`s tests.

So how can you distinguish among these saccarides? By using Osazone test.

Xylose, arbinose, ribose.

All of them are reducing monosaccharides (pentoses), and they all give Positive

result with :-

Molisch`s, Benedict`s, Barfoed`s, Bial`s tests. So how can you distinguish

among these saccarides? By using Osazone test.

Maltose, lactose.

Both of them are reducing disaccharides, and they give positive with:-

Molisch,Benedict`s tests.

So how can you distinguish between these saccarides? By using Osazone test.

Reagent:- Phenyl hydrazine Principle:- Yellow crystals compounds called

Osazone result from the reaction between sugars containing hydroxyl and

carbonyl group (reducing sugars) with phenyl hydrazine.The Osazone crystals have properties which assist in the identification of

reducing sugars only (monosaccharides , reducing disaccharides).That means

neither sucrose, nor polysaccharides can give Osazone crystals These properties

are:-

1-

Crystal shape by microscope. 2- Melting point. 3- Crystal formation time.

4- Osazone precipitated from hot or cold solution.

Procedure:- To 0.5 g of phenylhydrazine hydrochloride add 0.1 gram of sodium

acetate and ten drops of glacial acetic acid. Add 5 mL of test solution to this

mixture and heat under boiling water bath for about half an hour. Cool the

solution slowly and examine the crystals on slides under a microscope and record

your observations.17

Qualitative tests for Carbohydrates

8Sucrose test

Sucrose:- is only common non-reducing disaccharide, so that it does not reduce

alkaline copper solution or form osazone.

In sucrose the linkage is between carbon no. 1 in glucose, which represents

carbonyl end (reducing), and carbon no. 2 in fructose, which represents middle

reducing group, so that sucrose loses reducing ability.

Sucrose hydrolysis:-

Hydrochloric acid HCl hydrolyses the glycosidic bond of sucrose to yield

glucose and fructose 9reducing monosaccharides), and they give positive results

with benedict`s reagent.

Fructose is ketose and give positive result with Seliwanoff`s reagent. The sucrose

hydrolyses in human body by an enzyme called (sucrose).

Procedure:- In a test tube add 2.5 mL of sucrose solution, and then add 3 drops

of Conc. HCl.

Boil for 5 min in a boiling water bath. Cool the solution.

Divide it into two portions.

Perform Benedict`s test on the first portion as the following:-

Add 16 drops on this solution, and 2 drops of (5N) NaOH (Benedict`s test need an alkaline solution, heat for about 3 min in aboiling water bath, notice the

orange precipitate of Cu2O.

Perform Seliwanoff`s test on the second portion by adding 16 drops of the

reagent and heat for 3 min in aboiling water bath, notice the red complex

formation.19

Hydrolysis of starch

Starch:- is a polysaccharide consisting of glucose units joined by glycosidic

(etheric) bonds. The chains formed during the condensation reaction are either

linear or highly branched molecules. α1

6

branched molecules ofstarch are called amylopectin.while α1

4

linear both straight and helix molecules of starch is called amylase.

Natural starch from plants – consist of a mixture of amylase (10-25%) and

amylopectin (75- 90%).The structure of the helical amylase is key to the iodine

starch reaction.

Starch Conc.HCl

Glucose Steps of starch hydrolysis Starch

blue –

dark

Amylodextrine russet

Erythrodextrin

Acrodextrin

Maltose

yellow

glucose• The starch is not soluble in cold water, but soluble in hot water because of

high molecular weight.

Procedure of starch hydrolysis

- Take a test plates containing 6 wells to perform the iodine hydrolysis like this

shape.21

- Enumerate the wells as:-(0min, 3min, 6min, 9min, 12min and15min)

-Take 6 test tubes with its rach to perform Benedict`s test, enumerate also.

-Prepare a bout (5mL) of starch solution in another test tube .

- Using aplastic dropper, take one drop of starch solution and put it in the plate

well no.(0min),and another ten drops to put them in the test tube no.(0min), in

this stage the starch hydrolysis is 0 (no hydrolysis yet).

- Add 5 drops of Conc. HCl to the starch solution and start heating in a boilingwater bath for 3 minutes, then take a drop of the solution and put it in plate well

no.(3min), and another ten drops to put them in the test tube no.(3min).in this

stage the hydrolysis is 3 minutes.

- Continue heating the starch solutin in the boiling water bath for another 3

minutes,and repeat the steps above for the tube no.(6min),(9min),(12min)

and(15min).

- After 15 minutes we suppose that all starch has been hydrolysed in the presence

of Conc. HCl and heating , and the breaking of all glycosidic (etheric) bondings

between sugar units.

To continue testing the hydrolysis steps do the following:-

-

Perform the Iodine test on the 6 wells, by adding 2drops of iodine solution

oneach well of the plate, and record the colors resulting.23

-

Perform Benedict`s test on the 6 test tubes as

the following:- 1- Add 4 drops of NaOH to make the

medium basic,Why?

2-

add (1mL) of Benedict`s reagent.

3-

Heat in a boiling water bath until orange

precipitate appears in one of the 6 tubes.

* Notice the colours obtained in each tube (each

hydrolysis time) and explain the

results.

**Conc. HCl hydrolysed the glycosidic (etheric)

bonds in starch to yield the glucose.

*** The starch hydrolyses in human body by an

enzyme called (α-amylase).

Red complex

Avoid prolonged boiling, because aldoses yields

hydroxy methyl furfural which also reacts with

resorcinol to give red complex (false positive tests).24