SOME QUESTIONS WERE DERIVED FROM THIS IN THE JUNE 2014 BOARD EXAMS

SOURCE: LIPPINCOTT’s BIOCHEMISTRY

In this transcription, the red highlights were asked in our board exams. Good luck! :D
This is to SUPPLEMENT your BioChem notes, you still MUST read them, like all your other notes

PHARMACOGNOSY NOTES

Lactate —> aka lactic acid or alpha hydroxy propionic acid or 2-hydroxy propionic acid
Isoprene: 2-methylbuta-1,3-diene
Rubber - from Hever brasil ensue
Coined by Joseph Priestley
1,4 (X) Polymer

Latex - crude rubber that is collected from the tree as an aqueous dispersion that is washed, dried, and coagulated by
warming in air. Hardened by heating with Sulfur - a process called vulcanisation

Alpha-Amylase - hydrolyzes alpha-1,4

Lysozyme - breaks down alpha 1,4 bonds
Ptyalin —> salivary amylase

Glucose - complete enzymatic hydrolysis of starch

Maltose - major degradation product of Starch
Maltotriose - intermediate products of starch hydrolysis
Dextrin - partial starch hydrolysis
Liquid glucose - incomplete/acid hydrolysis

Glycosidic bonds of some carbohydrates

1. A-1,2
Sucrose

2. A-1,4
Maltose
Amylose
Amylopectin
Glycogen
Maltotriose
Dextrin

3. A-1,6
Dextrin, Dextran
Amylopectin q 25 to 30 units
Glycogen q 8 to 12 units
Trehalose

4, B-1,4
Cellubiose
Cellulose
Lactose

5. B-2,1
Inulin

New Family Names of Plants

Compositae - Asteracea
Cruciferaceae - Brassicaceae
Labiatae - Lamiaceae
Palmae - Arecaeae
Umbelliferae - Apiaceae
Graminae - Poaceae
Guttiferae - Clusiaceae
Leguminosae - Fabaceae sensu stricto
Papilonaceae - Fabaceae sensu lato

Rafinose —> melitose and melitriose

BIOCHEMISTRY AND LABORATORY TESTS

Presence of Halogens
A. Beilstein test
The Beilstein test is a simple chemical test used in chemistry as a qualitative test for halides. It was developed
by Friedrich Konrad Beilstein.[1]

A copper wire is cleaned and heated in a Bunsen burner flame to form a coating of copper(II) oxide. It is then dipped in
the sample to be tested and once again heated in a flame. A positive test is indicated by a green flame caused by the
formation of a copper halide. The test does not detect fluorine/fluorides.

CARBOHYDRATES
Fun Facts:
1. Galactose is most rapidly absorbed in the small intestines.
2. Concentrated dehydrating acids change monosaccharides to Uronic acids

Functions of Amino Acids

1. Converted to carbohydrates (glucogenic amino acids)
2. Tyrosine forms hormones such as T3, T4, catecholamines, and melanin.
3. Tryptophan is made into Niacin (Vitamin B3) and indole acetic acid (a plant hormone)
4. Creatine - made of Glycine, Arginine, Methionine (MR. G)
5. Glycine and cysteine, and taurine (a cysteine derivative) - bile salts (GC)
6. Glutamate, cysteine and glycine - GSH (Glutathione)
7. Histidine —> Histamine
8. Glycine is also used for HEME production for blood
9. Nucleic acid synthesis: Pyrimidines  DQ (Aspartate and Glutamine), Purines DGQS (Glycine, Aspartate, Glutamine,
and Serine)
10. Cysteine and Glycine —> Detoxicants
11. Methionine —> involves in transmethylation reaction; precursor to Carnitine,Melatonin (the natural sleep aid) and
Choline (part of the neurotransmitter, Acetylcholine).
12. Cystine and Methionine —> sources of Sulfur
13. Tranexamic acid is a lysine analog
14. Captopril is based on L-Lysine
15. Serine is coverted to ethanol-amine when CO2 is removed
16. L-carnitine is from Lysine
17. Arginine and Glutamine is a precursor to GABA (gamma-amino butyric acid)
18. Asparagine removes AMMONIA in the body
19. Asparagine is one of the two main excitatory neurotransmitters. Glutamate, made from glutamic acid, is the other.
Among their functions as neurotransmitters, of particular interest is the fact that Aspartic Acid and Asparagine have high
concentrations in the hippocampus and the hypothalamus. The hippocampus is a part of the brain that plays the main role
in short-term memory, while the hypothalamus is involved in the biology of emotion, and serves as a neurological gate
between the brain and the rest of the nervous system.
20. Glycine —> an inhibitory neurotransmitter

Liver Enzymes

1. Aspartate Transaminase (SGOT or Serum Glutmate Oxaloacetate Transaminase)

-Catalyzes the transfer of amino group of aspartic acid to ketoglutarate forming glutamate and oxaloacetate
Reaction:
Aspartate + Ketoglutarate — nitrogen transfer —> glutamate + oxaloacetate

2. Alanine Transaminase (SGPT or Serum Glutamate Pyruvate Transaminase)

-Transfer the amino group of alanine to ketoglutarate, forming glutamate and pyruvate
Reaction:
Alanine + Ketoglutarate — nitrogen transder —> glutamate + pyruvate

NOTE: CHECK YOUR MANOR HANDBOOK (ung horizontal/landscape layout) 

Optionally Essential Amino Acids

C, Y, P, Q, R, S

Essential Amino Acids

K, I, L - V, H, F - T, M, W

SUCROSE —> NOT A REDUCING SUGAR

REDUCING SUGAR - All monosaccharides, lactose, maltose, cellobiose, and gentiobiose

1. Molisch’s Test/A-napthol reaction

Purpose: Detects the presence of Carbohydrates
Reagent: Alpha-napthol and Conc. sulfuric acid
Result: Purple color at the junction.

2. Iodine Reaction
Results: Red color —> Glycogen (unbranched) ; Purple color —> Starch or Glycogen (branched)
Use: ID test for Glycogen and starch

3. Benedict’s Test
Purpose: Detection of Reducing Sugar
Uses: Sodium citrate as sequestering agent, Sodium carbonate, and Copper (II) Sulfate pentahydrate
Note: Alkali condition converts ketoses to aldoses
Results: Negative is solution remains blue or clear; Green (0.5%) Yellow (1%), orange (1.5%), Red ppt (2%) depending
on reducing sugar concentration

4. Barfoed’s Test
Purpose: Differentiates monosaccharides from polysaccharides; Ketoses due to acidity will not isomerise to form aldoses.
Reagent: Curpric ions in acidic medium (Benedict’s and Fehling’s = basic solution)
Positive result: Monosaccharides produces a ppt much faster, some dissacharides will produce ppt but the time difference
is significant.

5. Seliwanoff’s Test/Resorcinol Test)

Principle: Dehydration of furanose rings (of ketasaccharides) to hydroxymethyl furfural
Purpose: Detects ketoses
Reagent: Resorcinol in 6M HCl
Positve result: Rose color (Fructose, honey - natural fructose source, hydrolysed inulin - poly fructose)

6. Bial’s-Orcinol Test (also Benzidine Test —> for pentoses)

Reagent: Orcinol (5-methylresorcinol) in concentrated HCl with a small amount of Feric chloride.
Purpose: Test for Pentoses
Results: Pentoses undergo dehydration into fufural that condenses with orcinol to create a blue-green solution; hexoses -
yellow green; disaccharides - yellow.

7. Mucic Acid Test

Reagents: concentrate HNO3 (oxidizing agent)
Purpose: Detection of galactose
Note: Lactose will also hydrolyse to form galactose to yield insoluble dicarboxylic acid (mucic acid)
Results: Formation of crystals with sugars or solutions containing galactose

8. Phenyl Hydrazone Test/Osazone Test/Kowarsky Test

Reagents: Phenylhydrazine, Na acetate in acetic acid.
Results: Formation of Osazone crystals (yellow colour) through microscopic examination.
Glucose and Fructose has same crystals in microscope, but differentiated by melting point. Sucrose does not form an
oxazone. MANNOSE readily forms an osazone (the only insoluble osazone crystals)

Note: Hydrolysis of osazones produces ozones.

9. Moore’s Test
Results: Detection of Monosaccharides
When a solution of reducing sugar is heated with an alkali (NaOH), it turns yellow to orange and finally dark brown,
liberating the odor of caramel. This is due to the liberation of aldehyde which subsequent polymerizes to form a
resinous substance, caramel. Glucose, fructose or mannose when allowed to stand in the presence of weak alkali (BaOH)
is converted into a mixture containing all the three sugars and whichever one of them is used the same proportion of the
three sugars is always reached at equilibrium.

10. Fehling’s Reagent

Purpose: Detection of reducing sugars (like Benedict’s Test and Barfoed’s Test)
Reagents:
Fehling’s A: Copper (II) Sulfate
Fehling’s B: Rochelle salt + NaOH
Use equal volume in testing
Positive Result: Brick Red ppt of Copper (I) oxide

11. Tollen’s Test

Purpose: Detection of reducing substances (monosaccharides, citrate, and tartrate)
Reagents: Tollen’s Reagent (contains ammoniacal silver nitrate)
Result: Silver mirror due to reduction of silver ions into silver metal
Note: This test can differentiate citrate from tartrate because citrate required boiling to oxidise.

12. Anethole Test

Principle: Concentrated sulfuric acid hydrolyses bound sugars (breaking of glycosidic
bonds)→ dehydrates the monosaccharides to form furfural (if a pentose) or hydroxymethylfurfural (if a hexose)→
the furfural derivatives reacts with anthrone
Positive results: Blue-green colour
13. Tauber’s Test (Aminoguanidine Test)
Reagent: aminoguanidine and sulphuric acid
Use: Detection of Ketoses
Result: Ketohexoses (fructose and sorbose) or compounds which yield keto-hexoses (sucrose, raffinose, and inulin) give
rise to a bright reddish purple colour. The color is stable for several hours. Equivalent amounts of aldohexoses produce
no color in 24 hours.

Furfural gives a yellow color and furfuryl alcohol a brown color. Acetone, methylethyl ketone, and levulinic acid produce a
very slight yellow color.

The following pentoses, aldohexoses, and aldohexose-yielding compounds produce no color under similar conditions:
arabinose, xylose, ribose, glucose, galactose, mannose, lactose, maltose, melibiose, dextrin, starch, and glycogen. 

No color is given by formaldehyde or by the pure proteins, crystalline pepsin, crystalline trypsin, and crystalline
chymotrypsin. 

14. Benzidine Test (for ketoses)

15. Tollen’s phloroglucinol test

Use: Detection of pentose sugars, and ketohexoses (also detects Cellobiose —> a non pentose sugar)
Positive: Reddish brown colour (galactose and glucoronic acid)

16. Reaction with Alkaline Bismuth Reagent

Use: Detect the presence of reducing sugars

17. Glucose Oxidase Test (Test Tape)

Use: Detects sugar in diabetics; More specific than benedict’s test since glucose is not the only reducing sugar, and may
be interfered by antioxidants such as Vitamin C

PROTEINS
1. Biuret Test
Use: Detects the presence of peptide bonds/Positive for Histidine
Reagents: 2.5 M NaOH, 0.1 CuSO4
Positive Result: Violet solution

2. Ninhydrin Tests (2,2-Dihydroxyindane-1,3-dione)

Use: Detects the presence of amino acids
Reagents: 0.1% Ninhydrin Solution
Positive Result: Polar AA - Blue; Non-Polar AA - Blue VIolet; Proline (imino-acid) - Yellow

3. Xanthoproteic Tests
Use: Detects the presence of aromatic AA (Tyrosine, Phenyalanine, Tryptophan)
Reagents: conc. HNO3
Positive Result: Yellow ppt, which also turns orange on addition with alkali

4. Millon’s Test
Use: Specific for TYROSINE
Reagents: 10% Mercury (I) Iodine, NaNO2
Positive Results: Old rose colour (pink-red)

5. Sakaguchi Test
Use: Specific for Arginine
Reagents: A-napthol, 1 gtt of NaBrO
Positive Result: Red colour due to Guanidine group

6. Hopkin’s Cole Reaction (Glyoxylic acid reaction)

Use: Specific for Tryptophan
Reagents: Hopkins-Cole Reagent, Sulfuric Acid
Positive Result: Violet color (Collagen and gelatin is negative because it does not contain Tryptophan)

7. Nitroprusside Reaction
Use: Detection of cysteine
Reagents: Sodium nitroprusside in ammoniacal solution.
Positive result: Reddish color (This test is not positive for the sulfur of methionine)

8. Fohl’s Test (Lead acetate test)

Use: Specific for sulfur containing amino acids
Reagents: Goulard’s powder. 30% NaOH
Positive Results: Black ppt due to Lead (II) Sulfate
9. Pauly Diazo Test Test
Use: Detect Histidine, Tyrosine
Reagents: 1% Sulfanilic acid with NaNO2
Positive Results: Red

Proteins in Egg White

1. Ovalbumin
2. Ovomucin
3. Albumin
4. Vitelin

Casein —> isolated from milk through isoelectric precipitation

Tests for Kidney Efficiency
1. Phenylsulfophthelein test
2. Urea clearance Test
3. Water output test

LIPIDS
1. Acrolein Test
Use: Detection of glycerol
Reagents:
Positive Results: Silver mirror due to the reduction to the aldehyde (acrolein aka prop-2-enal)

2. Rosenheim Test
Use: Detects Choline

3. Iodine Absorption Test

Reagents: Hubl’s solution
Use: Determines degree of unsaturation

4. Lassaigne’s Test
Lassaigne’s test is used to detect the presence of elements: Nitrogen (N), Sulfur (S), Chlorine (Cl), Bromine (Br) and
Iodine (I). This test involves following two steps. 
i) preparation of sodium fusion extract (SFE). 
ii) Detection of elements using SFE.

5. Osmic Test
Use: To detect the presence of prosthetic groups in lipids

6. Ammonium phosphomolybdate test

Use: to detect phosphorus in phospholipids
Positive results: Canary Yellow

GLYCOSIDES
1. Cardiac Glycosides
A. Keller-Killiani Test
Reagent: Acetic acid + Ferric Chloride + Sulfuric acid
Use: Test for deoxy sugars (found in Cardiac glycosides)
Result: Blue or violet coloration at junction, and reddish brown solution

B. Liberman-Burchard/Acetic-acid and Sulfuric-acid Test

Use: Test for unsaturated sterol group
Reagent: Sulfuric acid and acetic anhydride
Result: Pale yellow if saturated sterol; Green or Blue if unsaturated sterol
C. Salkowski Test/Sulfuric Acid Test
Use: Test for sterol
Reagent: conc. H2SO4
Result: Red or violet color

D. Kedde’s Test
Use: Test for unsaturated lactone
Reagent: Standard digitoxin in methanol
Result: Blue-violet color/amethyst

E. Baljet’s Test

F. Legal’s Test

2. Anthraquinone
A. Borntrager’s Test
Reagent: Sulfuric acid + Benzene/Chloroform/Ether + ammonia
Positve result: Red/Pink layer

B. Modified Borntrager Test

Reagent:  NH4OH instead of ammonia
Positive result: Red/Pink layer

3. Saponin
A. Froth Test
Result: Honeycomb froth for 10 minutes above 2 cm

B. Capillary Test
Principle: Saponins tend to lower the surface tension in water;
Result: If the level of the plant extract in the capillarytube is half or less than that in the other tube containing water,then
the presence of saponins may be inferred.

C. RBC Hemolysis Test

Reagents: Gugo solution as standard; blood agar plate
Results: (+) formation of hemolytical halo (white ring); zone of hemolysis

D. Lieberman-Burchard
Use: Differentiate steroidal saponins from triterpenoidal saponins
Result: (+) for steroidal type —> blue or green; red, purple, or violet if tritertepenoid; yellow is saturated sterol.

4. Cyanophore Glycosides
A. Guignard’s Test
Use: nonspecific test for cyanophore glycosides; other substances may interfere - hydrogen sulphide, sulfur dioxide, or
aldehye
Reagent: Sodium picrate paper
Result: (+) yellow to red; bitter almond smell; If within 15 minutes —> presence of cyanophore glycosides; if after 3 hours
—> absence of cyanophore glycosides

5. Flavonol Glycosides
A. Batesmith-Metcalf Test
Purpose: To detect leucoanthocyanidins, chalcones, aurones
Positive result: Strong red —> violet

B. Wilstatter Test (Cyanidin Test)

Purpose: to detect gamma-benzo-pyrone nucleus
Positve result: Red/Magenta/Pink color

6. Isothiocyanate glycosides:
A. Odor Test - for volatile type
Positive result: Garlic odor for Allicin
Confirmatory Test for Garlic
Reagent: Na nitroferricyanide + 10 mL water + heat + NaOh
Result: Red orange color

B. Ferric chloride Test

Positive result: Blood red solution of Fe(SCN)3

7. Alcohol glycoside
Principle: detected by ester formation
Reagent: Acetic acid
Results: Fruity odor

8. Aldehyde glycoside
A. Fehling’s Test
<See carbohydrates>

B. Nessler’s Test
Reagent: Nessler’s Reagent (Potassium mercury iodine - K2HgI4)
Positive result: Gray ppt of mercury metal

C. Tollens/Silver mirror/Ammoniacal silver nitrate test

<See carbohydrates>

D. Schiff’s Test
Positive result: magenta red colour if there is aldehyde complexation reaction

9. Phenolic glycosides
A. Millon’s Test
B. Folin ciocalteau
C. Vanillin-HCl
C. Gibb’s reagent

TANNINS
A. Goldbeater’s Test
Use: General test for tannins, classifies tannins by MW
Principle: goldbeater's skin or ox skin is dipped in hydrochloric acid and treated with one percent ferrous sulfate solution,
after washing with water, it gives a blue-black colour
Result: Positive for HIGH MW Tannines (>5000); Negative for pseudo tannins (<5000)

B. Potassium ferricyanide and ammonia

Produces deep red color

C. Bayer’s Test

D. Bromine Test
Results: (+) with Non-hydrolyzable tannins; (-) with Hydrolyzable tannins

PRESUMPTIVE TEST FOR DRUGS

1. Marquis Test —> Ecstacy
2. Froehde Test —> Opioid
Murexide Test —> for purines and uric acid

Ketones
1. Tauber’s Test/Aminoguanidine reaction