1

The haemoglobin molecule was shown to transport oxygen as  oxyhaemoglobin by Fredrich L.hunefeld [1799-1882] see figure 2 in 1840 he was a member of German biochemistry association while viewing the  blood of earth worm eventhough the molecular mass was determined earlier by J.F Engelhard in 1825  see figure 4.3

Haemoglobin is a molecule made up of two alpha and two beta molecules held together with 4 haem molecules. The life span of  a haem molecule is as the same as the RBC 120 days where the concentration of haemoglobin is 14-16 grams per 100ml and the iron content of each haemoglobin molecule is 3.4 nanograms.

At the end of the life span of Rbc macrophages in spleen,liver or red bone marrow phagocytize rbcs  and the globulin part is broken down to aminoacids which later on can be used to synthesize other proteins and from the haem part the iron is removed attaches to plasma protein transferrin and when reaching the liver cells,muscle fibers iron detaches from transferrin and sticks to storage proteins called ferritin and haemosiderin.

Later on iron is released from storage proteins and reattaches to transferrin and is transported to the bone marrow where it is taken up by rbc [red blood cells] precursors for utilization of new haemoglobin molecules.

The Haem part has non-iron parts which is turned into biliverdin which is a green pigment then this molecule is turned into bilirubin within the liver,passed to bile as conjugated bilirubin in the process of the hepatocyte by the action of the enzyme  UDP-glucuronyl transferase which  conjugates it with glucuronyl acid which makes bilirubin water soluble.

Bile then passes to the small intestine where it  is degraded by bacteria into colorless water soluble urobilinogen.Urobilinogen further oxidize to compounds called urobilin and stercobilin.stercobilins are  excreted via feces where as some urobilinogen is absorbed to blood and converted to urobilin and excreted via urine which gives urine its

color.2

There are two types of haemoglobin and 4 types of Haem [Haem a-C49H56O6N4Fe,Haem b-C34H32O4N4Fe,Haem c-C34H36O4N4Fe,Haem o-C49H58O5N4Fe] as represented via molecular configuration.

1-Haemoglobin A- is main adult haemoglobin with 2 alpha and 2 beta chains.

2-Haemoglobin F- a type of haemoglobin present intrauterine existence and at birth with 2 alpha and 2 gamma chains.

Haemoglobin may combine with other chemicals in a life threatening conditions like for instance.

1-carboxyhaemoglobin:- combination of carbonmonoxide and haemoglobin.

2-Methaemoglobin:-combination of sulphonamides,benzocaine,amylnitrite,chloroquine,dapsone,nitrites,nitriglycerin,nitropruside,phenacentin and others in which iron is oxidized from ferrous Fe 2+ to ferric Fe 3+ state.4

Measurement methods of Haemoglobin.

1-Acid haematin method / Sahli method.

is a visual technique of haemoglobin estimation named after its developer Hermann Sahli [1856-1933] see figure 3 where haemoglobin converted to acid haematin by H2SO4 sulphuric acid and the color produced is matched to a comparator see figure 5.9

Procedure

1-Fill the graduated tube upto the mark 20 with 0.1 N Hcl.

2-Fill the haemoglobin pippette exactly upto 20 mm3.

3-wipe off the blood outside pippette with gauze.

4-empty the pippette into the acid in the tube gently.

5-Mix the acid haematin solution in the tube  and allow to stand for 5-10 minutes.

6-place tube in sahli comparator.

7-start adding 0.1 N Hcl or distilled water drop by drop till the color matches that of the standard.

8-Read the volume of solution in the graduated tube and express as grams 100ml or as percentage.

Other methods of estimation like cyanmethaemoglobin method,Alkali method,sheard-Sanford oxyheamoglobin method,sodium lauryl sulphate method, also exist according to Ramnik soods textbook 2006 edition.

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