Lab Anthraquinones PDF

## Exp.No. 2 ### **[Lab.3] Anthraquinone Glycosides** *Laxative* Anthraquinone and related glycosides, are stimulant cathartics, and exert their action by increasing the tone of the smooth muscle in the wall of colon and stimulate the secretion of water and electrolytes into the large intestine. A...

## Exp.No. 2 ### **[Lab.3] Anthraquinone Glycosides** *Laxative* Anthraquinone and related glycosides, are stimulant cathartics, and exert their action by increasing the tone of the smooth muscle in the wall of colon and stimulate the secretion of water and electrolytes into the large intestine. After the oral administration, the anthraquinone glycosides are hydrolyzed in the colon by the action of enzymes of the micro flora, to the pharmacologically active free aglycones which usually produce their effect in 8-12 hrs. After administration, these agents are indicated for constipation in patient who do not respond to milder drugs and for bowel evacuation before investigational procedure or surgery. Stimulant laxative are habit forming so the long-term use may result in laxative dependence and loss of normal bowel function. The glycosides of anthranols and anthroneselicit a more drastic reaction than do corresponding anthraquinone glycosides and cause discomforting and gripping action. The drugs mostly used are cascara, frangula, hypericum and Senna. Aloe and Rhubarb are not recommended due to their irritating actions which increase the chance for gripping effect. The anthraquinone hydrolyzed to give aglyconewhicharedi, tri, or tetra - hydroxyanthraquinone. Also there are antherone, dianthrones and oxanthrones. ### **Isolation and Identification of the Anthraquinone Glycosides:** #### **1. Extraction:** *Aim: To isolate the anthraquinone glycosides.* #### **Equipments:** * Large beaker & two medium size beakers. * Two conical flasks. * Centrifuge & Centrifuge tubes. * Separatory funnel. * Water bath. * Round bottom flask. * Filter paper. * Reagent bottle. #### **Reagents:** * Conc.HCl acid. * Chloroform. * 60% w/v ferric chloride solution. #### **Procedure:** * **Method of extraction:** Decoction. * **Plant used:** Senna *Cassia acutifolia*, *Cassia angustifolia* * **Family:** Leguminosaea. * **Part used:** Dry leaves. **Senna** Place 0.5 gm of powdered dry leaves of Senna in 50 ml of water *Boiling (15 min)* *Cool & filter* Place the filtrate in separatory funnel and extract by shaking with **[10 ml of Chloroform]** two times. * **Upper layer (Aqueous layer) [Free aglycone (dianthrone)]** * **Combine lower layer (Chloroform layer) Fraction B** The upper aqueous layer will be divided into two parts. * **(Whole Glycosides) Fraction A [put in reagent bottle]** * **Other part of the aq. Layer (10 ml)** * **Reflux (20 min)** Add (1) 3.5 ml of Ferric Chloride sol. (60 %w/v). (2) 2 ml of Conc.HCl acid. * **Cool** Place in a separatory funnel and extract with **[10 ml of Chloroform]** two times. * **Aqueous layer Glycone part** * **Chloroform layer Aglycone part (monoanthrone) Fraction C** #### **2. Results:** * **Fraction A:** Contain the whole glycosides. * **Fraction B:** Contain the aglycone (dianthrone). * **Fraction C:** Contain the aglycone part (monoanthrone). ### **[Lab.4] The Chemical Tests** #### **A. General Reaction:** For the following tests, boil 1 gm of the crude drugs (Aloe) given with 100 ml of water, add a little of kieselghur, filter and use the solution for the following tests: | | Compound | |---|---| | | | | | | #### **1. Schontetens Reaction (Borax test):** *Aim: Identification of the anthraquinone glycosides in general.* #### **Equipments & Reagents:** * Test tube. * Small beaker. * Water bath. * Borax. #### **Procedure:** To 2 ml of the Aloe extract, add 0.1 gm of Borax and heat until dissolved. Pour a few drops of the liquid into test tube nearly full of water. #### **Results:* A green fluorescence is produced. #### **Discussion:** This green fluorescence is due to aloe- emodinanthranols liberated from barbaloin by hydrolysis with Borax giving this reaction. #### **2. Bromine Test for Aloin:** *Aim: Identification of the anthraquinone glycosides in general.* #### **Equipments & Reagents:** * Test tube. * Bromine solution. #### **Procedure:** Take 2 ml of the Aloe extract, add an equal volume or an excess of freshly prepared solution of bromine. Record the color. #### **B. Specífic Reaction:** **Borntrager's test:** *الكاشف* *Aim: Identity test for aglycone part of anthraquinone glycosides.* #### **Equipments & Reagents:** * Separatory funnel. * Test tube. * Dilute HCl. * Benzene. * Dilute ammonia (10%). #### **Procedure:** To 5 ml of the Senna extract (fraction A), add 5 ml dilute HCl, then place the mixture in a separatory funnel and partitioing with 5 ml of benzene for 1 min. Take the upper benzene layer (free aglycone) and shake it with dilute ammonia (10%). Check the intensity of the color. #### **Results:** Pink color will be produced which is very clear with monoanthrones than dianthrones. #### **Discussion:** The benzene extracts the aglycone, and with ammonia, forms anthraqunone salts, which have pink color. ### **The Identification of Anthraquinone Glycosides By Chromatography:** *By the use of thin layer chromatography (T.L.C)* * The stationary phase = Silica gel G. * The mobile phase = n-propanol: Ethyl acetate: Water (60:30:30). * The standard compound = Sennoside. * The spray reagent = Alcoholic KOH 5%w/v. * Mechanism of separation = Adsorption. * Developing = Ascending. *NOTE/ for the best result spray first with 25%w/v nitric acid then heat in the oven after that spray with KOH reagent. This step is done to intensity the color of the spot.*