Urine Analyzes
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Hyperbilirubinemia
Porphyrins Porphyrins (Porphins) are cyclic tetrapyrol compounds formed by the linkage )). of four pyrrole rings through methenyl bridges (( HC In the reduced porphyrins (Porphyrinogens) the linkage of four pyrrole rings (tetrapyrol) through methylene bridges (( CH2 )) The characteristic property of porphyrins is the formation of complexes with the metal ion bound to nitrogen atoms of the pyrrole rings. e.g. Heme (iron porphyrin). Proteins which contain heme ((hemoproteins)) are widely distributed e.g. Hemoglobin, Myoglobin, Cytochromes, Catalase & Tryptophan pyrrolase. Natural porphyrins have substituent side chains on the eight hydrogen atoms numbered on the pyrrole rings. These side chains are: CH 1-Methyl-group (M)… (( 3 )) 2-Acetate-group (A)… (( CH2COOH )) 3-Propionate-group (P)… (( CH2CH2COOH )) 4-Vinyl-group (V)… (( CH CH2 )) Porphyrins with asymmetric arrangement of the side chains are classified as type III porphyrins while those with symmetric arrangement of the side chains are classified as type I porphyrins. Only types I & III are present in nature & type III series is more important because it includes heme. 1 Heme Biosynthesis Heme biosynthesis occurs through the following steps: 1-The starting reaction is the condensation between succinyl-CoA ((derived from citric acid cycle in the mitochondria)) & glycine, this reaction is a rate limiting reaction in the hepatic heme synthesis, it occurs in the mitochondria & is catalyzed by ALA synthase (Aminolevulinate synthase) enzyme in the presence of pyridoxal phosphate as a cofactor. The product of this reaction is α-amino-β-ketoadipate which is rapidly decarboxylated to form δ-aminolevulinate (ALA). 2-In the cytoplasm condensation reaction between two molecules of ALA is catalyzed by ALA dehydratase enzyme to form two molecules of water & one 2 molecule of porphobilinogen (PBG) which is a precursor of pyrrole. -
Characterisation of Bilirubin Metabolic Pathway in Hepatic Mitochondria Siti Nur Fadzilah Muhsain M.Sc
Characterisation of Bilirubin Metabolic Pathway in Hepatic Mitochondria Siti Nur Fadzilah Muhsain M.Sc. (Medical Research) 2005 Universiti Sains Malaysia Postgrad. Dip. (Toxicology) 2003 University of Surrey B.Sc.(Biomed. Sc.) 2000 Universiti Putra Malaysia A thesis submitted for the degree of Doctor of Philosophy at The University of Queensland in 2014 School of Medicine ABSTRACT Bilirubin (BR), a toxic waste product of degraded haem, is a potent antioxidant at physiological concentrations. To achieve the maximum benefit of BR, its intracellular level needs to be carefully regulated. A system comprising of two enzymes, haem oxygenase-1 (HMOX1) and cytochrome P450 2A5 (CYP2A5) exists in the endoplasmic reticulum (ER), responsible for regulating BR homeostasis. This system is induced in response to oxidative stress. In this thesis, oxidative stress caused accumulation of these enzymes in mitochondria — major producers and targets of reactive oxygen species (ROS) — is demonstrated. To understand the significance of this intracellular targeting, properties of microsomal and mitochondrial BR metabolising enzymes were compared and the capacity of mitochondrial CYP2A5 to oxidise BR in response to oxidative stress is reported. Microsomes and mitochondrial fractions were isolated from liver homogenates of DBA/2J mice, administered with sub-toxic dose of pyrazole, an oxidant stressor. The purity of extracted organelles was determined by analysing the expressions and activities of their respective marker enzymes. HMOX1 and CYP2A5 were significantly increased in microsomes and even more so in mitochondria in response to pyrazole-induced oxidative stress. By contrast, the treatment did not increase either microsomes or mitochondrial Uridine-diphosphate-glucuronosyltransferase 1A1 (UGT1A1), the sole enzyme that catalyses BR elimination through glucuronidation. -
Bilirubin Metabolism
Bilirubin Metabolism By Aseel .j.abdullah Introduction • Bilirubin is the orange-yellow pigment derived from senescent red blood cells. • It is a toxic waste product in the body. • It is extracted and biotransformed mainly in the liver, and excreted in bile and urine. • It is a bile pigment • Elevations in serum and urine bilirubin levels are normally associated with Jaundice. Erythrocytes become “old” as they lose their flexibility and increasingly rigid and fragile,they easily destruct during passage through tight circulation spots, especially in spleen, where the intra-capillary space is about 3 micron as compared to 8 micron of cell size RBCs useful life span is 100 to 120 days,After which they become trapped and fragment in smaller circulatory channels, particularly in those of the spleen. For this reason, the spleen is sometimes called the “red blood cell graveyard.” Dying erythrocytes are engulfed and destroyed by macrophages. Formation of Bilirubin • Primary site of synthesis:- SPLEEN : The Graveyard of Red Blood Cells • Secondary site of synthesis:- LIVER & BONE MARROW Pathophysiology RBCs Breakdown Hemoglobin Produces & Breakdown Heme Heme Oxygenase Biliverdin Biliverdin Reductase Bilirubin In Blood Unconjugated bilirubin • The bilirubin synthesized in – Lipid soluble spleen, liver & bone marrow – : limits excretion is unconjugated bilirubin. – 1 gm albumin binds 8.5 mg bilirubin • It is hydrophobic in nature so – Fatty acids & drugs can it is transported to the liver displace bilirubin as a complex with the – Indirect positive reaction plasma protein, albumin. in van den Bergh test • Most of the reabsorbed urobilinogen is taken up by the liver & is re-excreted in the bile. -
16Th March 2020 Blood Revised
Blood is the fluid circulating in a closed system of blood vessels and the chambers of the heart It is the medium which transports substances from one part of the body to the other Blood is composed of Plasma Platelets Cells WBCs RBCs (Erythrocytes) Hemoglobin (Hb) is red , oxygen carrying pigment present exclusively in erythrocytes HEMOGLOBIN A conjugated protein containing Globin Protein part ( 4 polypeptide chains- ) 96% of the total Hb mass Varies from species to species( species specificity) Heme Non protein (prosthetic group) Red colour Iron containing tetrapyrrole porphyrin derivative 4% of the total Hb mass Reversibly binds Oxygen Structure of Heme An Iron –porphyrin (Protoporphrin IX) compound with tetrapyrrole structure Protoporphyrin IX consists of 4 pyrrole rings combined through — CH= bridges (methyne bridges) The methyne bridges are referred as α,β,γ, and δ. The 2 Hydrogen atoms in the –NH groups pyrrole rings (II & IV) are replaced by Ferrous( Fe++) . The four pyrrole rings present in the porphyrin molecule are designated as I,II,III & IV . Each of these four rings has 2 groups attached to them M = Methyl –CH3 V = Vinyl – CH=CH2 P = Propionyl - CH2 - CH2 - COOH . The Fe++ can form 2 additional bonds .One of these position is linked internally (5th linkage ) to nitrogen of imidazole ring of Histidine of the Globin polypeptide chains . Other position is available to bind Oxygen Heme is the most prevalent metalloporphyrin in humans Common prosthetic group in Hemoglobin — Transport of O2 in blood Myoglobin — Storage of O2 in muscles Cytochromes — Part of electron transport chain Catalase — Degradation of H2O2 Tryptophan pyrolase — Oxidation of Tryptophan Cytochrome P450 — Hydroxylation of Xenobiotics HEME SYNTHESIS Major sites Liver Erythrocyte producing cells of bone marrow Rate of heme synthesis in liver is highly variable & depends upon size of heme pool while it is relatively constant in in bone marrow is relatively constant Mature RBC lack mitochondria and are unable to synthesize heme. -
About Wales' Pathology Handbook
Aneurin Bevan University Health Board About Wales' Pathology Handbook Wales Pathology Handbook (WPH) is a knowledge service within the Welsh national information architecture. It provides national and local catalogues of requestable tests, rules and guidance for electronic requesting (via the Welsh Clinical Portal (WCP) and GP systems) and a web-based knowledge resource for all clinical staff. The introduction of the new national Laboratory Information Management System (LIMS) for Wales means that the operational WPH needs to be reviewed and updated to ensure that it is fit for purpose in the future environment. Page 1 of 697 Date Created - Monday, September 27, 2021 Aneurin Bevan University Health Board Departments Andrology Blood Sciences Cytology Histopathology Microbiology Mortuary Services Transfusion Page 2 of 697 Date Created - Monday, September 27, 2021 Aneurin Bevan University Health Board Andrology Department Information PLEASE NOTE: THE PATHOLOGY HANDBOOK INFORMATION BELOW IS ONLY VALID AT THE TIME OF ACCESS, PRINTED COPIES MUST NOT BE USED Andrology services available are: Semen analysis for fertility investigations. Semen analysis following a vasectomy. Retrograde Ejaculation Investigation. Andrology work is carried out within the Andrology Department, Pathology at Nevill Hall Hospital. Lead Andrologist: Mrs Karen Llewelyn (01873 733062). Deputy Lead: Mrs Ruth Lancaster (01873 733062). Department Manager: Mr Julian Bendle (01633 234502). Please Scroll or Page Down for additional information relating to telephone numbers, sample requesting and collection. ROUTINE OPENING TIMES Nevill Hall site: Andrology Service Monday to Friday: 9:00am - 5:00pm Semen samples for Fertility analysis, Vasectomy analysis or Retrograde Ejaculation Investigation are processed by appointment only. Any sample booked without an appointment will not be tested. -
OPNF Hyperbilirubinemia Presentation
OPNF Hyperbilirubinemia Presentation 1) Definitions: a) Jaundice: _____________ discoloration of the skin and sclera caused by an elevated _________ level. b) Bilirubin: Metabolic end product of ____ breakdown. c) Always assess for jaundice in good lighting. Jaundice is not a reliable indicator for bilirubin level. 2) Bilirubin Physiology: a) Synthesis: Red blood cell breakdown Heme Biliverdin Bilirubin b) Bilirubin is _____ soluble. c) Transport: Bilirubin Free floating OR binds to Albumin and transports directly to the liver Taken up into the hepatocytes d) Metabolism (conjugation): Bilirubin is converted and becomes _____ soluble and secreted into bile and excreted then into the digestive track. e) Clearance/Excretion: i) Conjugated bilirubin changes into urobilinogen and excretes in urine. ii) Urobilinogen converts into stercobilinogen and excreted in the stool f) Enterohepatic circulation: Beta-glucuronidase from the lining of the small intestine deconjugates the bilirubin allowing it to be reabsorbed into circulation 3) Basics of Hyperbilirubinemia: a) Hyperbilirubinemia: _________ ___________ _________________ b) Physiologic Pathologic Jaundice after 24-48hr Jaundice within the first 24hr Requires no treatment Requires evaluation & treatment Peak: Day 3 in term, Day 5-6 in preterm Bili increases > 5mg/dL each day Resolved by 14 days of life Jaundice lasting longer than 14 days Normal infant appearance Can be anemic, discolored stools/urine Normal physiology of increased RBC Cause varies, any process that is destruction, reduced hepatic uptake, exaggerated enterohepatic reabsorption & decreased clearance c) Causes of Hyperbilirubinemia: i) Increased bilirubin production ii) Increased enterohepatic circulation iii) Decreased clearance of unconjugated bilirubin iv) Metabolic conditions v) Inborn errors of metabolism 4) Diagnosis a) Gold standard is the ___________________ which is the plasma level of bilirubin bound to albumin. -
Fate of Rbc's
HEMOGLOBIN AND HEMOGLOBINOPATHIES LECTURE 01 FATE OF RBCS AND JAUNDICE LECTURE 02 & 03 HEMOGLOBIN FIRST YEAR MBBS 2020 Features of a Mature RBC • Biconcave disc • Mean Diameter 7.8 um • Can deform easily. • Bag of fluid with dissolved substances and hemoglobin • No sub cellular particles • Metabolism – Anaerobic respiration- Glycolysis – Pentose phosphate pathway. RBC Count • Remains remarkably constant although there are some variations. • MALE : 5.2 ± 0.3 x 106 /uL. • FEMALE : 4.7 ± 0.3 x 106 /uL. • Life span : 120 Days HEMOGLOBIN HEME-CONTAINING PROTEINS Hemoglobin Myoglobin Cytochromes Catalase Some peroxidases HEMOGLOBIN • Metallo-conjugate protein • Molecular weight is 64,500 • One hemoglobin molecule is composed of four heme groups (subunits) attached with globin (having four polypeptide chains) • One hemoglobin molecule binds with it four oxygen molecules (eight oxygen atoms) • 1 gm hemoglobin carries 1.34 ml of oxygen • Heme synthesis occurs in the mitochondria of bone marrow erythroblasts HEMOGLOBIN FORMATION 2 succinyl-CoA + 2 glycine Pyrrole 4 pyrrole Protoporphyrin IX Protoporphyrin IX + Fe++ Heme Heme + Polypeptide Hemoglobin chain (α β) 2 α chains + 2 β chains Hemoglobin A Types of Hemoglobin • Variations in Hb subunit chains – W.r.t amino acid composition of polypeptide portion – Alpha, beta, gamma, delta Type of Hb Chain Fraction of total composition Hb HbA α2 β2 90% HbF α2 γ2 < 2% HbA2 α2 δ2 2-5 % HbA1C α2 β2 - Glucose 3-9% Embryonic Hemoglobins • Gower 1 Two zeta & two epsilon chains • Gower 2 Two alpha & two epsilon chains • Portland Two zeta & two gamma chains Embryonic/Minor Hemoglobins Organization of Hemoglobin Genes Developmental changes in Hb Hemoglobin F • Blood of the human fetus normally contains fetal hemoglobin • Its structure is similar to that of hemoglobin A except that the β chains are replaced by γ chains • hemoglobin F is α2γ2. -
Porphyrins and Bile Pigments: Metabolism and Disorders Dr
Porphyrins and bile pigments: metabolism and disorders Dr. Jaya Chaturvedi Porphyrins • Porphyrins are cyclic compounds formed by the linkage of four pyrrole rings through methyne (ÓHC—) bridges.In the naturally occurring porphyrins, various side chains replace the eight numbered hydrogen atoms of the pyrroles. • Porphyrins have had different structures depend on side chains that are attached to each of the four pyrrole rings. For example; Uroporphyrin, coporporphyyrin and protoporphyrin IX (heme). • The most prevalent metalloporphyrin in humans is heme, which consists of one ferrous (Fe2+) iron ion coordinated at the center of the tetrapyrrole ring of protoporphyrin IX. What is bilirubin? •Bilirubin is a yellowish pigment found in bile, a fluid made by the liver. •The breakdown product of Hgb from injured RBCs and other heme containing proteins. •Produced by reticuloendothelial system •Released to plasma bound to albumin •Hepatocytes conjugate it and excrete through bile channels into small intestine. Bilirubin di-glucoronid Structure of heme: • Heme structure: a porphyrin ring coordinated with an atom of iron side chains: methyl, vinyl, propionyl • Heme is complexed with proteins to form: • Hemoglobin, myoglobin and cytochromes Pathway of Heme Biosynthesis. Heme biosynthesis begins in the mitochondria from glycine and succinyl- CoA, continues in the cytosol, and ultimately is completed within the mitochondria. The heme that it produced by this biosynthetic pathway is identified as heme b. PBG: porphobilinogen; ALA: δ- aminolevulinic -
In Vitro DNA-Damaging Effects of Intestinal and Related Tetrapyrroles in Human Cancer Cells
View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector EXPERIMENTAL CELL RESEARCH 319 (2013) 536–545 Available online at www.sciencedirect.com journal homepage: www.elsevier.com/locate/yexcr Research Article In vitro DNA-damaging effects of intestinal and related tetrapyrroles in human cancer cells Christine Mo¨lzera,Ã, Barbara Pflegera, Elisabeth Putza, Antonia Roßmanna, Ursula Schwarza, Marlies Wallnera, Andrew C. Bulmerb, Karl-Heinz Wagnera,b aDepartment of Nutritional Sciences, Emerging Field Oxidative Stress and DNA Stability, Faculty of Life Sciences, University of Vienna, Althanstraße 14, 1090 Vienna, Austria bHeart Foundation Research Centre, Griffith Health Institute, Griffith University (Gold Coast Campus), Queensland 4222, Australia article information abstract Article Chronology: Epidemiological studies report a negative association between circulating bilirubin concentra- Received 4 September 2012 tions and the risk for cancer and cardiovascular disease. Structurally related tetrapyrroles also Received in revised form possess in vitro anti-genotoxic activity and may prevent mutation prior to malignancy. 3 December 2012 Furthermore, few data suggest that tetrapyrroles exert anti-carcinogenic effects via induction Accepted 4 December 2012 of cell cycle arrest and apoptosis. To further investigate whether tetrapyrroles provoke DNA- Available online 13 December 2012 damage in human cancer cells, they were tested in the single cell gel electrophoresis assay Keywords: (SCGE). Eight tetrapyrroles (unconjugated bilirubin, bilirubin ditaurate, biliverdin, biliverdin-/ Stercobilin bilirubin dimethyl ester, urobilin, stercobilin and protoporphyrin) were added to cultured Caco2 Urobilin and HepG2 cells and their effects on comet formation (% tail DNA) were assessed. Flow Protoporphyrin cytometric assessment (apoptosis/necrosis, cell cycle, intracellular radical species generation) SCGE assisted in revealing underlying mechanisms of intracellular action. -
Conversion of Amino Acids to Specialized Products
Conversion of Amino Acids to Specialized Products First Lecture Second lecture Fourth Lecture Third Lecture Structure of porphyrins Porphyrins are cyclic compounds that readily bind metal ions (metalloporphyrins)—usually Fe2+ or Fe3+. Porphyrins vary in the nature of the side chains that are attached to each of the four pyrrole rings. - - Uroporphyrin contains acetate (–CH2–COO )and propionate (–CH2–CH2–COO ) side chains; Coproporphyrin contains methyl (–CH3) and propionate groups; Protoporphyrin IX (and heme) contains vinyl (–CH=CH2), methyl, and propionate groups Structure of Porphyrins Distribution of side chains: The side chains of porphyrins can be ordered around the tetrapyrrole nucleus in four different ways, designated by Roman numerals I to IV. Only Type III porphyrins, which contain an asymmetric substitution on ring D are physiologically important in humans. Physiologically important In humans Heme methyl vinyl 1) Four Pyrrole rings linked together with methenyle bridges; 2) Three types of side chains are attached to the rings; arrangement of these side chains determines the activity; 3) Asymmetric molecule 3) Porphyrins bind metal ions to form metalloporphyrins. propionyl Heme is a prosthetic group for hemoglobin, myoglobin, the cytochromes, catalase and trptophan pyrrolase Biosynthesis of Heme The major sites of heme biosynthesis are the liver, which synthesizes a number of heme proteins (particularly cytochrome P450), and the erythrocyte-producing cells of the bone marrow, which are active in hemoglobin synthesis. Heme synthesis occurs in all cells due to the requirement for heme as a prosthetic group on enzymes and electron transport chain. The initial reaction and the last three steps in the formation of porphyrins occur in mitochondria, whereas the intermediate steps of the biosynthetic pathway occur in the cytosol. -
Recent Studies Ofthe Urobilin Problem1
J Clin Pathol: first published as 10.1136/jcp.16.1.1 on 1 January 1963. Downloaded from J. clin. Path. (1963), 16, 1 --Recent studies of the urobilin problem1 C. J. WATSON2 From the Department of Medicine, University of Minnesota Hospital, Minneapolis, Minn. It is now almost a century since Jaffe (1868, 1869) 1961). Sjostrand (1949) demonstrated that this is described urobilin. It would manifestly be an imposi- lost as CO, an important observation not yet tion on your patience if I were to attempt any com- exploited to any extent by clinical investigators. It is prehensive treatment of the ensuing history of this intriguing that the opening of the porphyrin ring to topic, but I shall strive to bring together for you a form bile pigment involves only the ix bridge carbon. few of what seem to me the more important mile- A number of years ago Schwartz and I (Watson and stones in their relation to recent studies. For reasons Schwartz, 1942) converted the bilirubins from each that will become apparent, the term urobilin, both of a series of human fistula biles to urobilinogen, on historical and clinical chemical grounds, is best thence to a crystalline urobilin, which was shown to applied to a group of closely related substances. be the same in all instances, i.e., 9, ax in type. Gray, Under ordinary circumstances the urobilin group is Nicholson, and Nicolaus (1958) at King's College related mainly to destruction of the haemoglobin of Hospital, using a much more elegant and precise circulating red cells. Other possible sources will be method depending on oxidation to monopyrrolic referred to later. -
Nomenclature of Tetrapyrroles
Pure & Appi. Chem. Vol.51, pp.2251—2304. 0033-4545/79/1101—2251 $02.00/0 Pergamon Press Ltd. 1979. Printed in Great Britain. PROVISIONAL INTERNATIONAL UNION OF PURE AND APPLIED CHEMISTRY and INTERNATIONAL UNION OF BIOCHEMISTRY JOINT COMMISSION ON BIOCHEMICAL NOMENCLATURE*t NOMENCLATURE OF TETRAPYRROLES (Recommendations, 1978) Prepared for publication by J. E. MERRITT and K. L. LOENING Comments on these proposals should be sent within 8 months of publication to the Secretary of the Commission: Dr. H. B. F. DIXON, Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, UK. Comments from the viewpoint of languages other than English are encouraged. These may have special significance regarding the eventual publication in various countries of translations of the nomenclature finally approved by IUPAC-IUB. PROVISIONAL IUPAC—ITJB Joint Commission on Biochemical Nomenclature (JCBN), NOMENCLATUREOF TETRAPYRROLES (Recommendations 1978) CONTENTS Preface 2253 Introduction 2254 TP—O General considerations 2256 TP—l Fundamental Porphyrin Systems 1.1 Porphyrin ring system 1.2 Numbering 2257 1.3 Additional fused rings 1.4 Skeletal replacement 2258 1.5 Skeletal replacement of nitrogen atoms 2259 1.6Fused porphyrin replacement analogs 2260 1.7Systematic names for substituted porphyrins 2261 TP—2 Trivial names and locants for certain substituted porphyrins 2263 2.1 Trivial names and locants 2.2 Roman numeral type notation 2265 TP—3 Semisystematic porphyrin names 2266 3.1 Semisystematic names in substituted porphyrins 3.2 Subtractive nomenclature 2269 3.3 Combinations of substitutive and subtractive operations 3.4 Additional ring formation 2270 3.5 Skeletal replacement of substituted porphyrins 2271 TP—4 Reduced porphyrins including chlorins 4.1 Unsubstituted reduced porphyrins 4.2 Substituted reduced porphyrins.