Porphyrins HPLC Assay Kit


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Porphyrins HPLC Assay kit: 
For Research Use Only
Size:  100 Tests
Incubation Time:  5 minutes
Sample Type:  Urine
Sample Size: 1 mL

Product Developed and Manufactured in Germany.
Product Support in the USA.

Intended Use
The Eagle Biosciences Porphyrins HPLC Assay Kit is intended for the quantitative determination of porphyrins in urine. This Porphyrins HPLC Assay Kit is for research use only and is not for use in diagnostic procedures.

Assay Principle
For the determination of the porphyrins a very easy sample preparation is performed first.  The pH value of the sample, calibrator and control is adjusted below 2.5 by addition of some drops (approx. 20 µl) of a stabilization solution. After centrifugation the supernatant is injected into the HPLC system.  The separation via HPLC follows an gradient method at 30°C, using a “reversed phase” column; one run lasts 25 minutes. The quantification is performed with the delivered calibrator; the concentration is calculated via integration of the peak areas or peak heights.

  1. Pipette into 1.5 ml reaction tubes: 1000 µl sample, CAL or CTRL + 20 µl HCL
  2. Mix well (pH value should be less than 2.5). Centrifuge at 10.000g for 5 minutes.
  3. Inject 100 µl of the supernatant for chromatography into the HPLC-system

Typical Chromatogram

Assay Background
Hemoglobin, myoglobin and the cytochromes contain the heme structure as active site. The porphyrins are precursors of the heme molecules. They play an important role in the oxygen metabolism. The heme synthesis is located mainly in the erythrocyte cells. Glycin and succinyl-CoA are condensed to δ-aminolävulinic acid. Two molecules of δ-aminolävulinic are then condensed to porphobilinogen. The porphyrin ring is built up by four porphobilinogen molecules.

An increase of porphyrins caused by a defect in the synthesis is described as porphyria. The different porphyries are determined by the characteristic pattern of porphyrins in urine.   In autosomal dominant acute porphyrin disorders uro-, copro-, penta- und tricarboxyporphyrin are increased. In chronic porphyries including porphyria cutanea tarda the concentration of uro- and heptaporphyrin are increased. In chronic lead intoxication the coproporhyrins are slightly increased, whereas an acute lead intoxication shows extremely high excretion of porphyrins in urine.

The Eagle Biosciences Porphyrins HPLC Assay Kit makes it possible to determine them in an easy, fast and precise way. The Porphyrins HPLC Assay Kit includes all reagents in ready to use form for preparation and separation of the samples with exception of the columns (IC2601rp) and the controls (IC2601ko).   Both can be supplied by Eagle Biosciences.  Beside the complete test kit it is possible to order all components separately. Please request our single component price list.

Limitations of the Method
Blood samples should not be analyzed with the Porphyrins HPLC Assay kit.