Cocarboxylase can be used in decarboxylation mechanisms in biological systems,and to investigate radical reactions of thiamin pyrophosphate in 2-oxoacid oxidoreductases.
Cocarboxylase is a thiamine (T344185) derivative produced by enzyme thiamine pyrophosphatase. Thiamine pyrophosphate is a cofactor used to catalyze various biochemical reactions.
Thiamine Pyrophosphate is a thiamine (T344185) derivative produced by enzyme thiamine pyrophosphatase. Thiamine pyrophosphate is a cofactor used to catalyze various biochemical reactions.
The active coenzyme
form of vitamin B1 which functions in aldehyde
transfer reactions.
4.5 kg of aqueous 89% orthophosphoric acid are heated to 135°C, and kept
at this temperature for about 3 h while being actively stirred. Then, the
heating is discontinued and 3.5 kg of phosphorouspentoxide are added during
a period of 2.5 to 3 h, while being actively stirred. During this period, the
interior temperature rises to 165°-175°C. After completion of the addition of
phosphorouspentoxide, the stirring is continued until all phosphorouspentoxide
is dissolved. The phosphoric acid mixture thus produced is subsequently
cooled down to 130°C.
At this temperature 2.0 kg of thiamine chloride hydrochloride (vitamin B1)
were added during 2 to 3 h while being well stirred. The stirring is continued
at 130°C until the phosphorylated mixture no longer contains chlorine ions. A
phosphorylated melt is thus obtained. The thus obtained phosphorylation melt
is dissolved in 6-8 L of water (with ice) at a temperature below 10°C, while
being vigorously stirred. The aqueous solution is stirred into 100 L of 96%
alcohol and left standing overnight. The supernatant solvent is decanted from
the separated syrup; the latter is taken up in 4 L of water. The solution thus
obtained is fed, depending upon the volume of phosphoric acid contained
therein, to an exchanger column filled with anion exchanger (weak basic, for
instance Amberlite IRA 45, 20-30 L) (a polystyrene resin with primary,
secondary and quaternary amino groups). The solution is caused to seep into
the column from the top thereof and is then washed with water until the runoff at the bottom no longer shows any thiamine reaction. About 25 L of the
solution are obtained, which are concentrated to 6 L at 30°C and 12 Torr. The
concentrated residue is added to 20 to 30 L of a cationic exchanger (Amberlite
IRC 50) in order to separate the thiamine-orthophosphoric acid ester from the
thiamine-pyrophosphoric acid ester, and subsequently washed with water until
the eluate is free of thiamine.
70-80 L of a solution are obtained which are concentrated to 1.5 L in a
circulation evaporator at 30°C and 12 Torr. 7.5 L of 96% ethanol are slowly
added to a concentrate while being stirred. The cocarboxylase-tetrahydrate
separates in the form of fine needles. The yield is 530.0 g with a melting point
of 220°-225°C (dec.).
10.0 g of cocarboxylase-tetrahydrate are dissolved in 25 ml of 5% aqueous
hydrochloric acid, and 75 ml acetone are added dropwise while being stirred.
The precipitated hydrochloride of the cocarboxylase, melting point 240°C is
sucked off. The yield is 9.5 g.
Certified pharmaceutical secondary standards for application in quality control, provide pharma laboratories and manufacturers with a convenient and cost-effective alternative to the preparation of in-house working standards. Thiamine pyrophosphate is a coenzyme of transketolase that catalyzes the cleavage of ribulose-5-phosphate; thereby forming D-glyceraldehyde-3-phosphate. This reaction requires the addition of an acceptor aldehyde such as ribose-5-phosphate, glyceraldehye or glycolaldehyde.
Thiamine pyrophosphate (TPP) is a coenzyme of transketolase, that catalyzes the cleavage of ribulose-5-phosphate. This reaction leads to the formation of D-glyceraldehyde-3-phosphate. It requires the addition of an acceptor aldehyde such as ribose-5-phosphate, glyceraldehye or glycolaldehyde. TPP is capable of decarboxylating hydroxypyruvate in the presence of an ′acceptor aldehyde′.
