BP-506P INDUSTRIAL PHARMACY-I

AHNIK SANJIB KASHYAP

BPH/10032/19

EXPERIMENT- 12

AIM:

To prepare and evaluate 5 ampoules each containing 10 mL of Calcium Gluconate IP.

REFERENCE:

The Theory and practice of Industrial Pharmacy, Leon Lachman, Herbert A. Lieberman, Joseph
L. Kanig, 3rd edition.

THEORY:

Calcium Gluconate is slowly soluble in 30 parts of water and soluble in 5 parts of boiling water.
Calcium in the form of Calcium Gluconate is used in the strength of 10% w/v. Any temperature
fluctuation during storage yields crystals of Calcium Gluconate. Thus, IP prescribes to replace a part of
Calcium Gluconate (not more than 5% of the total) by an equal amount of more soluble calcium salt such
as Calcium D-Saccharate or Calcium Lactobionate. These soluble calcium salts are called Stabilizing
Agents. The pH of the Calcium Gluconate injection can be adjusted from 6.0 to 8.2 using sodium
hydroxide (NaOH) or hydrochloric acid (HCl) solution. This is prepared in a single dose container in
Type-I glass container.

Calcium Gluconate is administered usually intravenously as 10% solution, slowly by direct

intravenous injection. Maximum administration rate that has been recommended for direct intravenous
(I.V.) injection is 1.5 mL/min and 2 mL/min. By intermittent infusion a maximum of 200 mg/min has
been suggested. Calcium Gluconate is compatible with excipients like 5% dextrose, 0.9% w/v sodium
chloride (NaCl), ascorbic acid, etc. Also, it is compatible with drugs like furosemide, heparin sodium,
chloramphenicol sodium succinate and penicillin G sodium. But it is incompatible with Indomethacin
sodium trihydrate, citrate of Indomethacin, soluble carbonate of Indomethacin, phosphates and
sulphates of Indomethacin.
Formula:
Calcium gluconate 9.65 g

Calcium D-saccharate 0.35 g

Water for injection (q.s.) 100 mL

PROCEDURE:

PREPARATION OF CALCIUM GLUCONATE SOLUTION:-

1. Required quantity of Calcium Gluconate and Calcium D- Saccharate was weighed accurately.

2. Because of the solubility problem, Calcium Gluconate is first dissolved in water for injection in
a beaker with the aid of heat application. Once it is ensured that Calcium Gluconate has
solubilised Calcium D-Saccharate is to be dissolved in the Calcium Gluconate solution.

3. The solution is allowed to cool down and after it is to be filtered through G-4 sintered glass filter
or by membrane filter of size 0.22 micrometre.

FILLING OF AMPOULES:-
1. Before filling, the ampoules are thoroughly washed and are dried properly to prevent dilution
of drug.

2. Using a hypodermic needle some amount of the solution is pipetted out from the prepared bulk
solution.

3. The needle should be inserted deep into the neck of the ampoules to prevent sticking of drug to
the neck and is blowed slowly under low pressure to prevent any bubble formation and
entrapment of oxygen and microbes.

STERILIZATION:-
Sterilization of the Calcium Gluconate is done preferably by Moist Heat Sterilization using Autoclave for
30 minutes at 121 ℃ at a pressure of 15 lb/psi.
USES:

It is used as calcium replenisher in osteoporosis patient.

EVALUATION:

1. LEAKAGE TEST:

Place the sealed ampoules in a tub or vessel containing methylene blue. Vacuum and pressure is applied
from the bottom of the vessel. In case of any kind of crack or improper sealing, methylene blue will enter
into the solution. The ampoules are then taken out from the vessel and checked if any of them is/are blue
in colour. Such blue colored ampoules are rejected, considering the presence of crack or improper
sealing.

2. CLARITY TEST:

A specific wooden background is taken, having two parts, black and white in colour. Over the
background, there is a hood installed with a tube light. The whole set-up is switched on. The ampule is
placed in front of the white background. If no particulate matter can be seen in front of the white
background, it is placed in front of the black background. White background will expose dark colored
particles in the product whereas black background will expose white coloured particles in the product.

3. STERILITY TEST:

A fluid glycolate medium or any other microbiological media is prepared. The product is
incubated in that media for 24 hours. After 24 hours, if there is no growth of any microbes in that media,
the product can be said to be completely sterile. If there is a growth of microbes in the media, it is said
that microbes have contaminated the product. Such type of product needs to be discarded. Pyrogen test:

This is a test to determine the presence of pyrogen and microbes inside the formulation, which
if enter in the glass container somehow, then will lead to an increase in temperature.

Pyrogens are metabolites of gram-negative bacteria. Gram positive bacteria alone releases
pyrogen but the lethality or the pathogenicity of gram-negative bacteria is much higher in comparison
to gram positive bacteria. Chemically pyrogens are mixture of polysaccharides conjugated with lipid
materials. In the pyrogen test, the product has to be sterilized. That’s why sterilization is done. In case of
drugs which are thermolabile, i.e. at high temperature there is a possibility of degradation, the removal
of pathogens or pyrogens is carried out by keeping the product in a UV radiation chamber or a gamma
radiation chamber, where the radiation will be given at higher intensity for smaller duration, because of
which most of the radiation will be able to enter and kill all the spores (if any) and will be able to
breakdown the complex of polysaccharide and lipid, thus destroying all the pyrogens in the product.
Gamma radiation is much more effective than UV radiation because the intensity of UV radiation is lesser
than that of gamma radiation. To detect whether a product is contaminated with pyrogens: Previously
animals (smaller mammals) were used. Rats and mice weren’t used as it is difficult to measure their body
temperature. Guinea pigs are preferred here.

They are injected with the product through the tail vein. After a period of one hour, a
thermometer is inserted into the anus of the guinea pig, and the temperature is checked. If there is no
increase in the temperature, then it is pyrogen free.

Later ethical issues arised due to difficulty in capturing these mammals. In order to avoid these
situations, Limulus Amoebocyte Lysate (LAL) test is exclusively done for detection of pyrogen in the
product. Limulus amoebocyte lysate is obtained from the blood of horseshoe crab (a sea animal). The
product is incubated with Limulus amoebocyte lysate, for one hour time period. If Limulus amoebocyte
lysate starts gelling, then it can be said that the product is contaminated with pyrogens. This is a very
quick estimation and it is followed throughout all pharmaceutical industries which are preparing
intravenous or parenteral preparations.

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