The reducing type is dispersed acicular crystal, and the oxidizing type is petal crystal. Both are easily soluble in water and acid solution. The former is pink in water solution, and the latter is dark red.It is relatively stable to heat.The former is more stable than the latter, with a molecular weight of about 11000-13000.Clinically, it is used as an auxiliary treatment for emergency treatment of hypoxia in various tissues, such as carbon monoxide poisoning, hypnotics poisoning, cyanide poisoning, neonatal asphyxia, hypoxia in severe shock period, cerebrovascular accident, sequelae of cerebral concussion, dyspnea caused by anesthesia and pulmonary diseases, and myocardial hypoxia caused by various heart diseases.
Chinese name
Cytochrome C
Foreign name
Cytochrome C
CAS No
Enzymatic action
molecular weight
eight hundred and eighty-four point eight eight seven zero zero
This product is an electron transmitter in the process of biological oxidation.Its action principle is that it can rapidly promote the oxidation and reduction of tissues in the presence of enzymes.Generally, exogenous cytochrome C cannot enter healthy cells, but in the absence of oxygen, the permeability of cell membrane increases, and cytochrome C may enter cells and mitochondria, enhance cell oxidation, and improve oxygen utilization.
Succinate coenzyme A from the tricarboxylic acid cycle has only 104 amino acids in its peptide chain, which is abundant in the body and generally does not need external supplementation;And the exogenous supplement is very little compared with the body content.Cytochrome C is a very important electron transmitter in biological oxidation. It forms a respiratory chain with other oxidase on the mitochondrial crest and participates in the process of cell respiration.
essential information
Chinese Name: Cytochrome C
Chinese alias: myomethemoglobin;Cytochrome C;Cytoheme C;
English name: Cytochrome C
English alias: Cytomac P;Cytochromert;myohematin;cytorest;Cytor-est;CYT-C;Cytochrome C (horse heart); Cytochrome c from horse heart; cytochrome C from pigeon breast muscle; cytochrome C from chicken heart; cytochrome C from porcine heart; cytochrome C from tuna heart; cytochrome C from bovine heart; cytochrome C from sheep heart; cytochrome C from saccharomyces*cerevisiae; cytochrome C practical grade: from*bovine heart; cytochrome C from dog heart; cytochrome C peptide map control; cytochrome C from rat heart; cytochrome C from rabbit heart; cytochrome C from bison heart; Cytochrome c, Equine Heart; Cytochrome (bovine heart); Cytochrome equine heart; Cytochrome from pigeon breast muscle; CYTOCHROME-C
Appearance and property: brown red crystalline powder
Melting point: 300 º C
Boiling point: 1323.5 º C at 760 mmHg
Flash point: 754.2 º C
Stability: Oxford when exposed to air
Storage condition: - 20 º C
security information
Customs code: 3507909000
WGK Germany：3
Safety instructions: S24/25
RTECS No.: HA5365000
Dangerous goods sign: Xi;Xn[2]
synthetic method
1. Clean the fresh or frozen pig heart, remove blood clots, fat and tendons, cut into strips, and grind them with a meat grinder.Then add 1.5 times of water, adjust the pH value to about 4 with 1mol/L sulfuric acid, stir at room temperature, extract for 2h, and press filter.The filtrate is neutralized with 1mol/L ammonia to pH=6.2, and the clear solution is obtained by centrifugation.The residue is extracted again with the same amount of water, and the two extracts are combined.Use 2mol/L ammonia water to adjust the pH value of the extract to 7.5, static precipitation of miscellaneous proteins, centrifugation, and absorption of the supernatant.Then add 10g zeolite per liter of extract, fully absorb it for 40min under constant agitation, leave it still, and discard the supernatant.Use distilled water to wash the zeolite adsorbed with cytochrome C for three times, 0.2% sodium chloride for four times, and then use distilled water to clean the washing solution.Filter the zeolite, drain it, put it into the chromatographic column, elute it with 25% ammonium sulfate solution, and collect the eluent.Add solid ammonium sulfate into the eluate to make the concentration reach 45%, and the miscellaneous proteins are completely separated and filtered.Slowly add trichloroacetic acid (20%) solution into the filtrate, stir while adding, and centrifugate to collect the precipitation after cytochrome C precipitation is completely separated.Dissolve the precipitate in distilled water, and dialysis with water until there is no sulfate ion.After dialysis, the solution is absorbed by the treated Amberlite IRC 50 (NH+4) resin column, washed with water until it is clear, and then eluted with a mixture of disodium hydrogen phosphate (0.6mol/L) and sodium chloride (0.4mol/L). The elution rate is slow, and the general flow rate is 2ml/min.The eluant is then dialyzed with distilled water until it is free of chloride ions to obtain refined cytochrome C solution.The yield is 100~150mg/kg, and the content is more than 20mg/ml.
2. Extraction method using fresh yeast as raw material
purpose
1. Cell respiration activators.It has a rapid enzymatic effect on the oxidation and reduction of cells in the tissue.It is used for tissue hypoxia caused by various reasons in emergency or auxiliary treatment.The leukopenia caused by anticancer drugs, limb circulation disorders, liver diseases, nephritis also have a certain therapeutic effect.
2. It is suitable for treating a series of symptoms caused by cerebral hypoxia, myocardial hypoxia and other tissue hypoxia.This product can also promote the regeneration of damaged liver cells, repair of bone marrow hematopoietic function, and significantly reduce leukopenia caused by radiotherapy.[3]
Pharmacopoeia standard of cytochrome C solution
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Source content
This strain is an aqueous solution of cytochrome C extracted from pig or cattle heart.The content of cytochrome C in every 1ml shall not be less than 15mg.
Preparation requirements
This product should be extracted from the hearts of pigs or cattle that have passed quarantine inspection, and the production process should meet the requirements of the current version of Good Manufacturing Practice for Drugs.
character
This product is a dark red clear liquid.
identify
(1) Take 1ml of the test solution under the item of iron content, and drop 20% trichloroacetic acid solution to generate brown or brownish red coagulated precipitates, and the red color of the solution disappears.The precipitation can be dissolved in water, and the solution is brownish red.
(2) Take 1ml of the test solution under the item of iron content, place it in a 50ml volumetric flask, dilute it to the scale with phosphate buffer solution (take 1.38g of sodium dihydrogen phosphate and 31.2g of disodium hydrogen phosphate, add appropriate amount of water to dissolve it into 1000ml, adjust the pH value to 7.3), add about 15mg of sodium hydrosulfite, shake it up, and determine it by ultraviolet visible spectrophotometry (Appendix IVA, Volume II, Pharmacopoeia 2010),There is maximum absorption at 520nm and 550nm, and minimum absorption at 535nm.
inspect
Iron content -- preparation of standard iron solution
Take 50g of ammonium ferric sulfate, add 300ml of water and 6ml of sulfuric acid to dissolve it, add appropriate amount of water to make it 1000ml, and shake it up.Precisely measure 25ml, put it into an iodine bottle, add 5ml of hydrochloric acid, mix it, add 12ml of potassium iodide test solution, close it, leave it for 10 minutes, titrate it with sodium thiosulfate titrant (0.1mol/L), when it is near the end point, add 1ml of starch indicator solution, continue titrating until the blue disappears.Every 1ml of sodium thiosulfate titrant (0.1mol/L) is equivalent to 5.585mg of Fe.Calculate the content of Fe (mg) per 1ml according to the consumption (ml) of sodium thiosulfate titrant (0.1mol/L).Precisely measure an appropriate amount, add sulfuric acid diluent (2ml diluted sulfuric acid, diluted to 500ml with water) to make 23 μ g Fe per 1ml.
Preparation of test solution
Precisely measure an appropriate amount of the product (about 100mg of cytochrome C), place it in a 10ml measuring flask, and dilute it with water to the scale.
Assay
Accurately measure 5ml of the test solution, place it in a crucible that has been ignited to constant weight, evaporate it to dryness, dry it at 105 ℃ to constant weight, accurately weigh W1, slowly ignite it to complete carbonization, and then continue to ignite it at 500~600 ℃ to complete ashing and constant weight, precisely weigh W2.In addition, accurately measure 1ml of the test solution, place it in a 25ml measuring flask, add 0.7ml of 30% hydrogen peroxide solution and 0.5ml of dilute sulfuric acid, place it in a water bath, heat it for 30 minutes, take it out, cool it down, precisely add 2ml of bipyridine test solution, place it in a cold water bath, slowly add 5ml of sodium sulfite test solution, shake it as you add it, place it in a 60~70 ℃ water bath, heat it for 30 minutes, take it out, cool it down to room temperature, dilute it to the scale with water,Shake well, measure the absorbance A1 at the wavelength of 522nm according to the ultraviolet visible spectrophotometry (Appendix Ⅳ A of Pharmacopoeia Volume II, 2010 Edition), and precisely measure 2ml of standard iron solution, place it in a 25ml measuring flask. According to the above method, operate according to the law from "adding 0.7ml of 30% hydrogen peroxide solution", measure the absorbance A2, and calculate the iron content as per the following formula, which should be 0.40%~0.46%.
Bacterial endotoxin
Take this product and check it according to the law (Appendix XI E, Part II, Pharmacopoeia 2010 Edition). The amount of endotoxin in each 1mg of cytochrome C should be less than 5.0EU.
Anaphylactic reaction
Take an appropriate amount of this product and add water for injection to dilute it into a solution containing 7.5mg of cytochrome C per 1ml, as the test solution, and check according to the law (Appendix XI K of Part II of Pharmacopoeia 2010 Edition), which should meet the requirements.
vitality
It shall not be less than 95.0% according to the cytochrome C activity determination method (Appendix XII B, Part II, Pharmacopoeia 2010).
Assay
Accurately measure 1ml of the test solution under the item of iron content, place it in a 50ml volumetric flask, dilute it to the scale with the phosphate buffer under the item of identification (2), add about 15mg of sodium hydrosulfite, shake it up, and use the ultraviolet visible spectrophotometry (Appendix IVA, Part II, Pharmacopoeia 2010) to find the maximum absorption wavelength at the wavelength of about 550nm with an interval of 0.5nm to determine the absorbance,Calculated as the absorption coefficient (E) of cytochrome C is 23.0.
preparation
(1) Cytochrome C injection (2) Cytochrome C for injection[4]
Method name:
Cytochrome C Injection Determination of Cytochrome C Spectrophotometry
Scope of application:
This method uses spectrophotometry to determine the content of cytochrome C in cytochrome C injection.
This method is applicable to cytochrome C injection.
Method and principle:
Take an appropriate amount of this product, add phosphate buffer solution to dissolve it and fix the volume, then add sodium hydrosulfite, measure the absorbance with ultraviolet visible spectrophotometry, and calculate according to the absorption coefficient (E1 m) of cytochrome C as 23.0.
Reagent:
1. Sodium hydrosulfite
2. Phosphate buffer
Instrument and equipment:
Visible spectrophotometer
Sample preparation:
Preparation of test solution
Accurately measure 1mL of this product as the test solution.
Note: "Precise weighing" means that the weighing shall be accurate to one thousandth of the weighing."Precision measurement" means that the accuracy of volume measurement shall meet the accuracy requirements of the national standard for the volume pipette.[5]
pharmacological action
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Cytochrome C is extracted from pig or cow heart and is an electron transmitter in the process of biological oxidation.The action is similar to coenzyme.When the tissue is hypoxic, the cell permeability increases. After injection, cytochrome C can enter the cell to correct cell respiration and promote material metabolism.
This product is a cell respiration activating enzyme, which can be used as an emergency medicine and auxiliary medicine for the treatment of tissue hypoxia, such as carbon monoxide poisoning, hypnotics poisoning, newborn suspended animation, irreversible shock hypoxia, and treatment before anesthesia;Pneumonia, lung cancer, silicosis, emphysema, dyspnea caused by bronchiectasis, altitude sickness hypoxia, cerebrovascular disorders, brain trauma, cerebral hemorrhage, cerebral arteriosclerosis, concussion sequelae, Japanese encephalitis sequelae, pertussis encephalopathy, etc;Toxic myocardial damage, myocarditis, myocardial insufficiency, angina pectoris, diphtheria myocarditis, myocardial infarction, etc;The decrease of white blood cells, circulatory disorders of limbs, nerve paralysis, liver disease and nephritis caused by anti-cancer drugs can also be improved.[6]
Induce apoptosis
Research on apoptosis shows that cytochrome C is related to apoptosis, and cytochrome C leaked from mitochondria can induce apoptosis.
Pharmacokinetics
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After injection of cytochrome C, most of it is used by hypoxic tissues.A small part is distributed in heart tissue, gradually reduced by liver and kidney, and discharged by urine.[6]
indication
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Cytochrome C is an activator of cell respiration, which is used for first-aid or auxiliary treatment of various tissue hypoxia, such as carbon monoxide poisoning, central depressant (hypnotic) poisoning, neonatal asphyxia, hypoxia in severe shock period, dyspnea caused by anesthesia and pulmonary diseases, alpine hypoxia, cerebral hypoxia and hypoxia caused by heart disease, but the effect is sometimes not significant.In foreign countries, cytochrome C is a component of some ophthalmic drugs for cataract treatment, but its role is unclear.[6]
Contraindication
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1. People who are allergic to cytochrome C.
2. Allergy test shall be conducted before medication, and it is forbidden for those with positive reaction.[6]
Dosage and usage
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1. Intramuscular injection: 15mg each time for adults, once a day, 30mg each time for severely ill patients, twice a day, and more serious patients can be increased as appropriate.The dosage for children should be reduced as appropriate.
2. Intravenous injection: 15~30mg each time, 1~2 times a day, add 20ml of 25% glucose solution, mix well, and slowly inject;It can also be diluted with 5%~10% glucose solution or isotonic saline and then drip.
3. For powder injection (freeze-dried type), 20 ml of 25% glucose solution or 5% glucose solution or sterilized osmotic salt solution are used for infusion.[6]
matters needing attention
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1. May cause allergic reaction, and allergy test shall be conducted before use.
2. Allergy test shall be conducted again when it is used again after being stopped, and it shall not be used for those who are positive.
3. In the treatment, if the drug is stopped, shock is likely to occur when the drug is used again. If shock occurs, it should be treated with vasopressors, cardiotonic drugs, antihistamines, and adrenocortical hormone
4. Comprehensive measures should be taken for the treatment of hypoxia.[6]
Adverse reactions
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1. May cause allergic reaction, and allergy test shall be conducted before use.
2. Local spasm, rash, fever, thirst, temporary shock and other reactions.[6]
Expert comments
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Cytochrome C is used for first-aid or auxiliary treatment of various tissue hypoxia, such as carbon monoxide and hypnotic poisoning, neonatal asphyxia, brain trauma and other causes of cerebral hypoxia.The leukopenia caused by anticancer drugs, limb circulation disorders, liver diseases and nephritis also have some improvement effects.[6]
Cytochrome
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There are many kinds of cytochromes, which can be distinguished by spectroscopy, the precise structure of heme groups, the sensitivity of inhibitors and the size of reduction potential.
According to the different cofactors, cytochrome can be roughly divided into three categories:
Classification auxiliary basis
Cytochrome a Heme a (proheme)
Cytochrome b Heme b (formyl heme)
Derivatives of cytochrome d dihydroporphyrin [1]
Cytochrome c is not named according to the cofactor;Different from the other three types of non covalently bound hemes, cytochrome c covalently binds to heme through the cysteine sulfhydryl group in the molecule.[2] There is no cytochrome e, but there is cytochrome f (one of cytochrome c).[3] In addition, cytochrome d only exists in bacteria.
In mitochondria and chloroplasts, cytochrome is often bound to complex enzymes of electron transport and related metabolic pathways.
Relational judgement
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The closer the relatives are, the more similar or identical their cytochrome C.For example, people and chimpanzees.However, if there is a great difference in the evolutionary history, the difference in cytochrome C is also greater, for example, between human and yeast.
In view of the indispensable position of cytochrome C in the respiratory electron transport chain, its molecular structure is relatively conservative.However, with the evolution of organisms, their molecular composition and structure will also change. However, compared with other proteins, many variations of cytochrome C molecules may lead to problems in the electronic transmission of organisms and lead to death. Therefore, their molecular evolution should be slow and conservative.
