CN105784831A - Method for detecting trace elements in human body whole blood through inductively coupled plasma mass spectrometry method - Google Patents
Method for detecting trace elements in human body whole blood through inductively coupled plasma mass spectrometry method Download PDFInfo
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Abstract
The invention provides a method for detecting trace elements in human body whole blood through an inductively coupled plasma mass spectrometry method and belongs to the technical field of biotechnical detection. The method comprises the following steps: (1) pre-treating a whole blood sample; (2) detecting a mixed element standard solution and a to-be-detected sample solution by an inductively coupled plasma mass spectrometer; and (3) performing quantitative analysis on a detection result and calculating the content of to-be-detected elements in the human body whole blood sample. The method provided by the invention can be used for simultaneously detecting the content of trace elements, such as magnesium, manganese, calcium, iron, copper, zinc and lead, in the human body whole blood; the sample dosage is less; the pretreatment is simple; the cost is low; the whole detection process is short in time and high in efficiency; the detection sensitivity and precision andthe detection stability are high.
Description
Technical field
The invention belongs to technical field of biological, relate to the method that inductively coupled plasma mass spectrometry measures human whole blood medium trace element.
Background technology
The change of body trace element particularly Trace Elements in Blood content and physiological pathology of human body situation have and directly contact.Trace Elements in Blood magnesium, manganese, calcium, ferrum, copper and zinc are the important component forming the mcroorganism molecules such as enzyme, hormone, vitamin, important biochemical functions is played at human body, and it can affect human immune system, causal connection is had with multiple disease, as calcium deficiency can cause chondroma, osteoporosis and rickets etc., and calcium excess intake can suppress the absorption of zinc, easily cause the diseases such as cholelithiasis；Ferrum is one of important composition of hemoglobin, and iron deficiency is easily caused iron deficiency anemia；Hepatocarcinoma, diabetes, gestation and have abnormal labor history manganese element content in the patient to reduce, and in acute hepatitis patient, gangrenosum acne liver cirrhosis, heart infarction, manganese poisoning patient, tumor patient visible volume, manganese element content is slight or moderate increases.Metallic pollution in environment and life style can metabolism and accumulation in vivo by all means, thus affecting the healthy of people, and lead is relatively conventional one in life.Trace Elements in Blood Toxicity of Lead can involve nerve, blood, hemopoietic, digestion, cardiovascular and urinary system, and child intelligence is produced irreversible impact.
The detection of whole blood medium trace element magnesium, manganese, calcium, ferrum, copper, zinc and lead content can reflect element concentration in human body, can as health care, auxiliary diagnostic elements shortage or toxic disease and the reliable basis observing curative effect.At present, in whole blood, the various constituent content method for measuring of trace has atomic absorption spectrography (AAS) (AAS), atomic fluorescence spectrometry (AFS) and inductively coupled plasma emission spectrography (ICP-AES) etc..AAS and AFS is long due to analytical cycle, once can only detect a kind of element, and efficiency is low, and accuracy and the range of linearity can not contain all of clinical concentration of element and be vulnerable to the reason such as impact of body interference, limits its application clinically；And ICP-AES can measure many elements simultaneously, easy to operate, cost is low, good stability, it it is the detection method commonly used clinically, but needing to use corresponding matrix modifier in its mensuration process, sample pretreatment process is loaded down with trivial details, its detection limit also cannot meet the requirement that in blood, contents of heavy metal elements measures simultaneously.
It is wide and can carry out the features such as multielement detects simultaneously, sample expense is few that inductively coupled plasma mass spectrometry (ICP-MS) detection possesses highly sensitive, the range of linearity, suitable in the mensuration of batch samples, now have been widely used for the fields such as geoscience, environmental analysis, Food Science, petroleum industry and metallurgical analysis.The paper of " blood and 34 kinds of method for detecting elements and applied research thereof in urine " by name that Zhang Sujing delivers discloses 34 kinds of element ICP-MS detection methods in a kind of blood, its sample pre-treatments adopts electric hot plate heating resolution method, reagent dosage is big, easily cause the loss polluted with volatile components, and the sample size of single treatment is few, use duration.
Therefore, those skilled in the art are simple in the urgent need to a kind of pre-treatment, good stability, the detection time is short, efficiency is high, detection precision is high, the with low cost method measuring human whole blood medium trace element.
Summary of the invention
In order to solve problems of the prior art, the present invention provides a kind of method that inductively coupled plasma mass spectrometry measures human whole blood medium trace element, can be used for the content detection of detection human whole blood medium trace element magnesium, manganese, calcium, ferrum, copper, zinc and lead simultaneously.The amount of samples of human whole blood medium trace element detection method provided by the invention is few, and pre-treatment is simple, with low cost, and whole detection process time is short, efficiency is high, detection sensitivity and precision is high, detection good stability.
Technical scheme is as follows:
Inductively coupled plasma mass spectrometry measures the method for human whole blood medium trace element, comprises the steps:
(1) pre-treatment of whole blood sample: take whole blood sample, fully after mixing, uses the mixed liquor of nitric acid and TritonX to be diluted as diluent, fully shakes even, obtain sample detection liquid；
(2) complex element standard solution and described sample liquid icp ms ICP-MS to be measured is detected: after icp ms being carried out instrument optimization with tuning liquid, adopt full quantitative model, spectrogram scan mode, isotope ion scan mode, in online addition, mark working solution, is carried out detection by quantitative to complex element standard solution and sample liquid to be measured successively by ICP-MS；；
Wherein, target quota ion includes elemental isotope to be measured, and its mass number and condition include: magnesium mass number is 23 or 24；Calcium mass number is 44；Weight of iron number is 57 or 58；Manganese mass number is 55；Copper mass number is 63 or 65；Zinc mass number is 66 or 68；Plumbous mass number is 206,207 and 208；
Wherein, described icp ms running parameter includes: high frequency generator power 1550W；Flow rate of carrier gas 1.0L/min；Assisted gas flow velocity 1.0L/min；Carrier gas flow velocity 0.0L/min；Compensation current speed 1.0L/min；Cooling gas velocity 15.0L/min；Aerochamber temperature: 2 DEG C；Peristaltic pump rotating speed: 0.2r/s；Sampling depth 6-8mm；Lifting time 30s；The time of integration 0.1s；Reading duration 8s；Number of repetition 2 times；
Collision/reaction tank condition includes: CellEntrance:-40V；CellExit:-60V；Deflect:0V；PlateBias:-60V；OctPRF:180V；OctPBias:-18V；QPBias:-15V；Gasflow (He): 4.0～5.0mL/min；
(3) quantitative analysis of testing result, in ratio according to elemental signals to be measured in complex element standard solution and interior mark signal and complex element standard solution, the concentration drawing standard curve of element to be measured, utilizes the content of element to be measured in the quantitative sample of ratio of elemental signals to be measured and interior mark signal in sample.
Preferably, the mass concentration of described diluent is 0.01～0.1%.
Preferably, described whole blood sample extension rate is 1: 10～1: 40.
Preferably, the nitric acid that described tuning liquid is 0.2% using volume fraction obtains as diluted preparation, and concentration is 5.0-40.0 μ g/L, and described tuning liquid includes lithium, cobalt, yttrium, thallium, cerium, indium and barium element.
Preferably, the nitric acid that described interior mark working solution is 0.2% using volume fraction obtains as diluted preparation, and concentration is 20-100 μ g/L, and described internal standard element is scandium, germanium, yttrium, rhodium, indium, terbium and bismuth.
Preferably, described complex element standard solution is the complex element standard solution containing magnesium, manganese, calcium, ferrum, copper, zinc and plumbous seven kinds of elements.
Compared with prior art, the invention has the beneficial effects as follows: detection method provided by the invention can realize the content detection of whole blood sample medium trace element magnesium, manganese, calcium, ferrum, copper, zinc and lead simultaneously, there is amount of samples few, pre-treatment is simple, the feature such as with low cost, and whole detection process time is short, efficiency is high, detection sensitivity and precision are high, detection good stability.
Accompanying drawing explanation
Fig. 1 inductively coupled plasma mass spectrometry measures the standard curve of magnesium (Mg) in human whole blood trace element method.
Fig. 2 inductively coupled plasma mass spectrometry measures the standard curve of ferrum (Fe) in human whole blood trace element method.
Fig. 3 inductively coupled plasma mass spectrometry measures the standard curve of copper (Cu) in human whole blood trace element method.
Fig. 4 inductively coupled plasma mass spectrometry measures the standard curve of manganese (Mn) in human whole blood trace element method.
Fig. 5 inductively coupled plasma mass spectrometry measures the standard curve of zinc (Zn) in human whole blood trace element method.
Fig. 6 inductively coupled plasma mass spectrometry measures the standard curve of plumbous (Pb) in human whole blood trace element method.
Fig. 7 inductively coupled plasma mass spectrometry measures the standard curve of calcium (Ca) in human whole blood trace element method.
Detailed description of the invention
Below in conjunction with drawings and Examples, the present invention is described in further details.
The detection of embodiment one human whole blood medium trace element magnesium, manganese, calcium, ferrum, copper, zinc and lead
1, instrument: icp ms
2, solution preparation:
Diluent preparing: the ratio of 1: 999 adds nitric acid and the TritonX mixed liquor that deionized water preparation mass concentration is 0.1% by volume, is diluent.
Tuning liquid preparation: the salpeter solution that tuning storing solution volumetric concentration is 0.2% being configured to the tuning liquid that mass concentration is 10 μ g/L standby, tuning element is lithium, cobalt, yttrium, thallium, cerium, indium and barium.
Interior mark working solution is prepared: the salpeter solution that interior mark storing solution volumetric concentration is 0.2% being configured to the interior mark working solution that mass concentration is 100 μ g/L standby, internal standard element is scandium, germanium, yttrium, rhodium, indium, terbium and bismuth.
Complex element standard solution is prepared: take each elemental standards solution of lead, manganese, copper, ferrum, zinc, magnesium and calcium, with the complex element standard solution of salpeter solution that volumetric concentration is 0.2% 5 kinds of variable concentrations of preparation, six kinds of trace element concentration in every kind of standard solution is as shown in table 1.
The concentration of six kinds of trace element in table 1 standard solution
P/A tunes the preparation of liquid (detector double mode calibration solution): formulated by the salpeter solution that volumetric concentration is 0.2%, its concentration is according to the instrument standard curve that the same day, each element to be measured was set up, its concentration value corresponding to 100-200 ten thousand response signal corresponding.
3, sample detection
(1) pre-treatment of whole blood sample: extract whole blood 2mL with anticoagulant heparin pipe, the mixing whole blood fully drawing 0.25mL after mixing joins in the diluent prepared, and in the ratio of 1: 30, whole blood is diluted, fully shakes even, obtains sample detection liquid.Diluent contains the nitric acid of low concentration, can the hemocyte in blood be decomposed, and makes the Element release contained in hemocyte enter in diluent；TritonX in diluent can ensure the uniformity of the solution after fully mixing simultaneously.
(2) with tuning liquid, ICP-MS being carried out instrument optimization, 0.1s, tuned requirement and reached following standard: 1 sweep time) NoGasMode sensitivity: Li (7) >=15000；Y(89)≥50000；Tl(205)≥30000；2) resolution (at10%): 0.65～0.80；3) mass axes: ± 0.05amu；4) oxide: CeO/Ce≤3.0%；5) double charge: Ce2+/Ce≤3.0%；6) RSD≤5.0%；7) P/A tunes coefficient is 0.75～0.2.
After instrument optimization completes, adopting full quantitative model, spectrogram scan mode, isotope (mass-to-charge ratio) ion scan mode, in online addition, mark working solution, is carried out detection by quantitative to complex element standard solution and sample liquid to be measured successively by ICP-MS；
Wherein, target quota ion mass number and condition include: magnesium mass number is 23 or 24；Calcium mass number is 44；Weight of iron number is 57 or 58；Manganese mass number is 55；Copper mass number is 63 or 65；Zinc mass number is 66 or 68；Plumbous mass number is 206,207 and 208；
Icp ms running parameter includes: high frequency generator power 1550W；Flow rate of carrier gas 1.0L/min；Assisted gas flow velocity 1.0L/min；Carrier gas flow velocity 0.0L/min；Compensation current speed 1.0L/min；Cooling gas velocity 15.0L/min；Aerochamber temperature: 2 DEG C；Peristaltic pump rotating speed: 0.2r/s；Sampling depth 7mm；Lifting time 30s；Reading duration 8s；The time of integration is 0.1s；Number of repetition 2 times；
For eliminating interference, measuring in collision/reaction tank pattern, condition includes: CellEntrance:-40V；CellExit:-60V；Deflect:0V；PlateBias:-60V；OctPRF:180V；OctPBias:-18V；QPBias:-15V；Gasflow (He): 4.0～5.0mL/min；
(3) quantitative analysis of testing result, the concentration drawing standard curve of element to be measured in ratio according to elemental signals to be measured in complex element standard solution and interior mark signal and complex element standard solution, standard curve need to meet R > 0.995, and result is shown in Fig. 1-Fig. 7.Utilize the content of element to be measured in the quantitative sample of ratio of elemental signals to be measured and interior mark signal in sample, simultaneously, for eliminating the difference of separate sources sample isotope ratio, use correction equation Pb (208)=Pb (206)+Pb (207)+Pb (208), namely lead isotope 206,207,208 three's signal value (CPS) is all added by each test, then is calculated.
The mensuration of embodiment 2 detection method detection limit
Taking 0.25mL deionized water, the ratio using 1: 30 mixes deionized water and diluent as sample blank, measures, calculate its average X and standard deviation SD in repeating sample blank to make for 20 times batch, using 3 times of standard deviation SD as detection limit.The detection limit of each element is as shown in table 2.
The each method for detecting element detection limit of table 2
| Element to be measured | Magnesium (mg/L) | Manganese (μ g/L) | Ferrum (mg/L) | Copper (μ g/L) | Zinc (mg/L) | Plumbous (μ g/L) | Calcium (mg/L) |
| Average | 0.023 | 0.383 | 0.998 | 1.400 | 0.008 | 0.632 | 0.233 |
| Standard deviation | 0.009 | 0.113 | 0.830 | 0.636 | 0.002 | 0.064 | 0.040 |
| Detection limit | 0.026 | 0.339 | 2.491 | 1.908 | 0.005 | 0.192 | 0.120 |
Embodiment 3 detection method precision measures
1, quality-control product preparation:
Quality-control product one is SeronomTMTraceElementsWholeBloodL-2 (purchased from this nation of Australia), by specification requirement pipettor accurately draws 3mL ultra-pure water in bottle, stands 30min, then it is inverted bottle, after placing 1 hour, shakes up subpackage gently, it is stored in-20 DEG C of refrigerators, uses in stable phase.
Quality-control product two is autogamy cattle whole blood.
Quality-control product adds each elemental standards solution to be measured as sample to be tested, the concentration of each element in quality-control product is made to be embodied within the scope of whole blood medical science decision level, wherein, lead is 100.0 μ g/L, and manganese is 6.6 μ g/L, copper is 720.0 μ g/L, zinc is 3.7mg/L, and ferrum is 557.2mg/L, magnesium is 50.4mg/L, and calcium is 84.0mg/L.
2, test method
(1) withinrun precision: using quality-control product one as detecting sample, replication 20 times.Calculate its average X, standard deviation SD and coefficient of variation CV；
(2) betweenrun precision: measure the data of 20 different batches samples.Calculate its average X, standard deviation SD and coefficient of variation CV.
3, Precision test result, in Table 3.
Table 3 precision test coefficient of variation CV result
| Project | Magnesium | Manganese | Ferrum | Copper | Zinc | Plumbous | Calcium |
| Withinrun precision | 1.0% | 3.5% | 2.0% | 2.0% | 1.0% | 2.0% | 1.0% |
| Betweenrun precision | 2.5% | 4.0% | 3.5% | 3.0% | 3.5% | 5.0% | 4.0% |
Result of the test shows, detection method withinrun precision ranges for 1.0-3.5%, and betweenrun precision ranges for 2.0-5.0%, and assay method of the present invention detection precision is high.
Above content is in conjunction with concrete preferred implementation further description made for the present invention, it is impossible to assert that specific embodiment of the invention is confined to these explanations.For general technical staff of the technical field of the invention, without departing from the inventive concept of the premise, it is also possible to make some simple deduction or replace, protection scope of the present invention all should be considered as belonging to.
Claims (6)
1. the method that inductively coupled plasma mass spectrometry measures human whole blood medium trace element, it is characterised in that: comprise the steps:
(1) pre-treatment of whole blood sample: take whole blood sample, fully after mixing, uses the mixed liquor of nitric acid and TritonX to be diluted as diluent, fully shakes even, obtain sample detection liquid；
(2) complex element standard solution and described sample liquid icp ms ICP-MS to be measured is detected: after icp ms being carried out instrument optimization with tuning liquid, adopt full quantitative model, spectrogram scan mode, isotope ion scan mode, in online addition, mark working solution, is carried out detection by quantitative to complex element standard solution and sample liquid to be measured successively by ICP-MS；
Wherein, target quota ion includes elemental isotope to be measured, and its mass number and condition include: magnesium mass number is 23 or 24；Calcium mass number is 44；Weight of iron number is 57 or 58；Manganese mass number is 55；Copper mass number is 63 or 65；Zinc mass number is 66 or 68；Plumbous mass number is 206,207 and 208；
Wherein, described icp ms running parameter includes: high frequency generator power 1550W；Flow rate of carrier gas 1.0L/min；Assisted gas flow velocity 1.0L/min；Carrier gas flow velocity 0.0L/min；Compensation current speed 1.0L/min；Cooling gas velocity 15.0L/min；Aerochamber temperature: 2 DEG C；Peristaltic pump rotating speed: 0.2r/s；Sampling depth 6-8mm；Lifting time 30s；The time of integration 0.1s；Reading duration 8s；Number of repetition 2 times；
Collision/reaction tank condition includes: CellEntrance:-40V；CellExit:-60V；Deflect:0V；PlateBias:-60V；OctPRF:180V；OctPBias:-18V；QPBias:-15V；Gasflow (He): 4.0～5.0mL/min；
(3) quantitative analysis of testing result, in ratio according to elemental signals to be measured in complex element standard solution and interior mark signal and complex element standard solution, the concentration drawing standard curve of element to be measured, utilizes the content of element to be measured in the quantitative sample of ratio of elemental signals to be measured and interior mark signal in sample.
2. the method that inductively coupled plasma mass spectrometry according to claim 1 measures human whole blood medium trace element, it is characterised in that: the mass concentration of described diluent is 0.01～0.1%.
3. the method that inductively coupled plasma mass spectrometry according to claim 1 measures human whole blood medium trace element, it is characterised in that: described whole blood sample extension rate is 1: 10～1: 40.
4. the method that inductively coupled plasma mass spectrometry according to claim 1 measures human whole blood medium trace element, it is characterized in that: the nitric acid that described tuning liquid is 0.2% using volume fraction obtains as diluted preparation, concentration is 5.0-40.0 μ g/L, and described tuning liquid includes lithium, cobalt, yttrium, thallium, cerium, indium and barium element.
5. the method that inductively coupled plasma mass spectrometry according to claim 1 measures human whole blood medium trace element, it is characterized in that: the nitric acid that described interior mark working solution is 0.2% using volume fraction obtains as diluted preparation, concentration is 20-100 μ g/L, and described internal standard element is scandium, germanium, yttrium, rhodium, indium, terbium and bismuth.
6. the method that inductively coupled plasma mass spectrometry according to claim 1 measures human whole blood medium trace element, it is characterised in that: described complex element standard solution is the complex element standard solution containing magnesium, manganese, calcium, ferrum, copper, zinc and plumbous seven kinds of elements.
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