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The core-tube method is a common method to measure the coal seam gas content (CSGC). However, cutting heat and friction heat will be generated in the core-tube coring process, which will increase the coal core temperature and the coal core gas loss, thus resulting in a large error in the determination of the gas content. The accuracy of the gas content determination is closely related to the temperature variation of coal core during core-taking. Based on this, the team developed the "thermal effect simulation device of coal core in the core-taking process" and carried out the temperature change test experiment of the coal core in the core-taking process under different conditions. The results show that the temperature variation of the coal core during the core-taking process shows four stages: constant temperature, rapid temperature rise, slow temperature rise, and temperature drop. The temperature rise rate, temperature rise duration, and temperature rise peak of the coal core increase with the increase in rotate speed, coal strength, friction area, and frictional load. In the axial direction, the closer to the upper end of the core pipe, the higher the core temperature. In the radial direction, the closer the core is to the wall of the core pipe, the higher the core temperature is. Under the influence of cutting heat and friction heat in the process of core-taking, the maximum heating rate of the core-taking tube wall within 8 min is 20 °C/min, the peak temperature is 158.4 °C, the average temperature of the wall is above 100 °C, and the average temperature rise of the coal core reaches 55.7 °C. Within 60 min, the average temperature of the coal core remained above 50 °C. The order of influence of coal core temperature from large to small is as follows: rotate speed, frictional load, friction area, and coal strength. It can provide a reference for accurately determining CSGC using the core-tube method or designing a coring device to eliminate or reduce the thermal effect during coring.

Is thyroid autoimmunity (TAI) associated with the decline of ovarian reserve in euthyroid women?Case-control study. Data from 4302 euthyroid women with normal ovarian reserve (NOR) and low ovarian reserve (LOR), including biochemical premature ovarian insufficiency (POI) and overt POI, were retrospectively analysed. The prevalence and effect of TAI on ovarian reserve was evaluated between women with NOR and LOR. Status of ovarian insufficiency and TSH levels was further stratified for analysis. The correlation between anti-thyroid peroxidase antibody (TPOAb), anti-thyroglobulin antibody (TgAb) titres and ovarian reserve markers was also determined.The prevalence of positive TAI and TgAb was equally distributed between women with NOR and LOR (P = 0.080, P = 0.172); the prevalence of TPOAb positivity was higher in the LOR group (P = 0.005). After stratifying ovarian reserve and TSH, positive TAI, TPOAb and TGAb were significantly associated with overt POI when TSH was2.5 µIU/ml (all P0.001); no association was observed with biochemical POI or overt POI when TSH was ≤2.5 µIU/ml. No correlation was found between TPOAb, TGAb titres and AMH (P = 0.218, P = 0.368, respectively), and bilateral AFC (P = 0.184, P = 0.315, respectively) in patients with LOR; only TPOAb titre was positively correlated with FSH (P = 0.039).Among the whole population of euthyroid women, TAI was not associated with low ovarian reserve but was significantly associated with overt POI in women with TSH2.5 µIU/ml. Further basic studies on underlying mechanisms are needed.

STUDY QUESTION Are mutations in MOS (MOS proto-oncogene, serine/threonine kinase) involved in early embryonic arrest in infertile women? SUMMARY ANSWER We identified mutations in MOS that may cause human female infertility characterized by preimplantation embryonic arrest (PREMBA), and the effects of the mutations in human embryonic kidney 293T (HEK293T cells) and mouse oocytes provided evidence for a causal relation between MOS and female infertility. WHAT IS KNOWN ALREADY MOS, an activator of mitogen-activated protein kinase, mediates germinal vesicle breakdown and metaphase II arrest. Female MOS knockout mice are viable but sterile. Thus, MOS seems to be an important part of the mammalian cell cycle mechanism that regulates female meiosis. STUDY DESIGN, SIZE, DURATION Whole-exome sequencing, bioinformatics filtering analysis and genetic analysis were performed to identify two different biallelic mutations in MOS in two independent families. The infertile patients presenting with early embryonic arrest were recruited from October 2018 to June 2020. PARTICIPANTS/MATERIALS, SETTING, METHODS The female patients diagnosed with primary infertility were recruited from the reproduction centres of local hospitals. Genomic DNA from the affected individuals, their family members and healthy controls was extracted from peripheral blood. We performed whole-exome sequencing in patients diagnosed with PREMBA. Functional effects of the mutations were investigated in HEK293T cells by western blotting and in mouse oocytes by microinjection and immunofluorescence. MAIN RESULTS AND THE ROLE OF CHANCE We identified the homozygous missense mutation c.285C>A (p.(Asn95Lys)) and the compound heterozygous mutations c.467delG (p.(Gly156Alafs*18)) and c.956G>A (p.(Arg319His)) in MOS in two independent patients. The mutations c.285C>A (p.(Asn95Lys)) and c.467delG (p.(Gly156Alafs*18)) reduced the protein level of MOS, and all mutations reduced the ability of MOS to phosphorylate its downstream target, extracellular signal-regulated kinase1/2. In addition, the identified mutations reduced the capacity of exogenous human MOS to rescue the metaphase II exit phenotype, and the F-actin cytoskeleton of mouse oocytes was affected by the patient-derived mutations. LIMITATIONS, REASONS FOR CAUTION Owing to the lack of in vivo data from patient oocytes, the exact molecular mechanism affected by MOS mutations and leading to PREMBA is still unknown and should be further investigated using knock-out or knock-in mice. WIDER IMPLICATIONS OF THE FINDINGS We identified recessive mutations in MOS in two independent patients with the PREMBA phenotype. Our findings reveal the important role of MOS during human oocyte meiosis and embryonic development and suggest that mutations in MOS may be precise diagnostic markers for clinical genetic counselling. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the National Natural Science Foundation of China (81725006, 81822019, 81771581, 81971450, 81971382,82001538 and 82071642), the project supported by the Shanghai Municipal Science and Technology Major Project (2017SHZDZX01), the Project of the Shanghai Municipal Science and Technology Commission (19JC1411001), the Natural Science Foundation of Shanghai (19ZR1444500 and 21ZR1404800), the Shuguang Program of the Shanghai Education Development Foundation and the Shanghai Municipal Education Commission (18SG03), the Foundation of the Shanghai Health and Family Planning Commission (20154Y0162), the Capacity Building Planning Program for Shanghai Women and Children’s Health Service and the collaborative innovation centre project construction for Shanghai Women and Children’s Health. The authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER N/A.

To investigate the impact of cryopreservation (CP) duration on pregnancy outcomes of vitrified-warmed blastocysts transfers using an open-device liquid-nitrogen (LNThis retrospective cohort study was conducted on 6327 first vitrified-warmed single blastocyst transfer cycles with autologous oocytes from January 2015 to December 2020. The CP duration was initially divided into six groups: Group I: 0-3 months (n = 4309); Group II: 4-6 months (n = 1061); Group III: 7-12 months (n = 304); Group IV: 13-24 months (n = 113); Group V: 25-72 months (n = 466); Group VI: 73-120 months (n = 74). Multivariate logistic regression was performed to evaluate the independent effect of CP duration on pregnancy outcomes. To further examine the time limit of vitrification, propensity score matching (PSM) was applied to compare pregnancy outcome of patients with storage duration of 25-120 months to those of 0-24 months. After that, pregnancy outcomes were compared among the subgroups of Group I': 0-24 months, Group II': 25-48 months, Group III': 49-72 months, Group IV': 73-120 months. Stratification analysis based on embryo quality was also performed. Primary outcomes were clinical pregnancy rate and live birth rate. Secondary outcomes were implantation, biochemical pregnancy rate, ongoing pregnancy rate and early miscarriage rate.Logistic regression demonstrated that the odds of pregnancy outcomes were similar across Group I to IV. However, the implantation rate, chances of biochemical pregnancy, clinical pregnancy, ongoing pregnancy, and live birth significantly decreased as the storage duration increased up to 25 months, while miscarriage rate did not significantly differ between groups. Subgroup analysis confirmed a dramatical decrease of clinical pregnancy and live birth rate when cryopreserved for more than 24 months. After that, the slope was relatively steady between 25 and 72 months, then steeply decreased again as CP reached 73-120 months. In addition, there was a more remarkable decline of pregnancy outcomes in the average quality embryo transfers than in the high quality embryo transfers as cryopreservation storage increased.Prolonged cryopreservation of vitrified blastocysts in an open-device LN

To explore the optimal cycle regimen for frozen-thawed embryo transfer (FET) in women with previous intrauterine adhesions (IUAs).In this retrospective cohort study, a total of 1,002 FET cycles for patients with previous IUAs from January 2015 to December 2020 were included. Among them, 294 conventional hormone replacement therapy (HRT) cycles were matched with 155 HRT with gonadotropin releasing hormone agonist pretreatment (HRT+GnRH-a) cycles using propensity score matching. Multivariate logistic regression analysis was performed to further investigate the impact of cycle regimen on pregnancy outcomes.After propensity score matching, baseline characteristics were consistent between HRT and HRT+GnRH-a group. Logistic regression analysis revealed that there was a significant superiority of HRT+GnRH-a over the conventional HRT group regarding the incidences of live birth (aOR=1.966, 95%CI: 1.212-3.188, P=0.006) and ongoing pregnancy (aOR=1.710, 95%CI: 1.057-2.767, P=0.029). HRT+GnRHa also had a higher odd of clinical pregnancy (aOR=1.414, 95%CI: 0.903-2.216, P=0.130), and lower odd of early miscarriage (aOR=0.511, 95%CI: 0.219-1.195, P=0.121) compared to HRT, yet not reached statistical significance.HRT with GnRH-a pretreatment improves pregnancy outcomes in women with previous IUAs. GnRH-a may restore the endometrial receptivity in the FET cycles of IUAs.HRT with GnRH-a pretreatment may promote pregnancy prognosis of FET in women with history of IUAs by restoring endometrial receptivity.

Endometriosis, defined as the presence of endometrial tissues outside the uterus, affects approximately 10% of women of reproductive age. Assisted reproductive technology (ART) is considered the most effective technique to treat infertility in women, although the impact of endometriosis on ART outcomes remains ambiguous. In this study, 433 patients with endometriosis and 1299 infertile patients with tubal factors receiving in vitro fertilization (IVF) treatment were retrospectively enrolled to determine whether a history of endometriosis affects pregnancy outcomes. Patients with endometriosis had markedly fewer retrievable oocytes, a lower oocyte maturity rate, and decreased numbers of available and high-quality embryos (all p 0.001) compared to those with tuber factors. The rates of clinical pregnancy and live birth in the endometriosis group were lower in the frozen-thawed embryo transfer cycles (p = 0.028 and p = 0.008, respectively), and a decreased cumulative live birth rate (CLBR) (p = 0.001) was observed. Logistic regression analysis revealed a negative association between endometriosis and CLBR (p = 0.002). The number of macrophages in the follicular fluid (FF) of patients with ovarian endometriosis was significantly higher than that of patients without ovarian endometriosis (p 0.001). The levels of interleukin (IL)- 1α, IL-1β, tumor necrosis factor-α, IL-6, IL-13, and IL-10 in FF were also elevated in the endometrioma group than in the control group (p 0.05). These results indicate that endometriosis is negatively associated with CLBR in IVF, which may be caused by the follicular immune microenvironment that affects the development and fertilization of oocytes.

PURPOSE: We aimed to identify novel variants in TUBB8 and corresponding new abnormal phenotypes in oocytes/fertilization/ embryonic development responsible for female infertility. METHODS: Sanger sequencing of TUBB8 was performed in infertile women with abnormalities in oocyte maturation or embryonic development. The effects of the variants were evaluated in patients’ oocytes by morphological observations and immunofluorescence. RESULTS: We identified 34 novel variants of TUBB8 in 51 patients who were diagnosed with abnormalities in oocyte maturation or early embryonic development. We found a novel phenotype in which large polar bodies were present in three independent patients possibly associated with a recurrent variant. Moreover, we identified a novel type of TUBB8 variant consisting of an in-frame deletion-insertion, which has not been previously reported. CONCLUSIONS: Our present study identified 34 novel variants in TUBB8 in 51 patients. These patients show oocyte maturation arrest, oocytes with large polar body, fertilization failure, early embryonic arrest or embryonic implantation failure. These results expand the kinds of variants and phenotypic spectrum of TUBB8 variants with regard to female infertility. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10815-020-01830-6) contains supplementary material, which is available to authorized users.

Objective: This retrospective study aimed to explore the optimal endometrial preparation method in women with intrauterine adhesions (IUAs).Method: A total of 882 frozen-thawed embryo transfer (FET) cycles from patients with history of IUAs were categorized into three groups based on endometrial preparation methods: hormone replacing therapy cycle (HRT, n=636), natural cycle (NC n=174), and HRT with GnRH-a pretreatment (HRT+GnRH-a, n=72. Logistic regression was performed to investigate the association between cycle regimens and pregnancy outcomes. Subgroup analysis of IUAs combined with thin endometrium (≤7mm) was also performed.Results: HRT with GnRH-a pretreatment was associated with higher incidences of clinical pregnancy, ongoing pregnancy, and live birth, but lower early miscarriage compared with either HRT or NC. Logistic regression indicated that after controlling for potential confounders, the incidences of live birth (HRT+GnRH-a as reference; NC: aOR=0.577, 95%CI 0.304-1.093; HRT: aOR=0.434, 95%CI 0.247-0.765) and ongoing pregnancy (NC: aOR=0.614, 95%CI 0.324-1.165; HRT: aOR=0.470, 95%CI 0.267-0.829) remained significantly higher in HRT+GnRH-a compared to those in HRT, but comparable to those in NC. While there was no significant difference with respect to the clinical pregnancy rate (NC: aOR=0.695, 95%CI 0.374-1.291; HRT: aOR=0.650, 95%CI 0.374-1.127) and early miscarriage rate (NC: aOR=1.734, 95%CI 0.417-7.175; HRT: aOR=2.594, 95%CI 0.718-9.378) between groups. Subgroup analysis suggested there was no superiority of endometrial preparation method in IUAs combined with thin endometrium.Conclusion: HRT with GnRH-a pretreatment improves pregnancy outcomes in women with history of IUAs. GnRH-a may restore the endometrial receptivity in the FET cycles in IUAs.

Objective: This retrospective study aimed to explore the optimal endometrial preparation method in women with intrauterine adhesions (IUAs).Method: A total of 882 frozen-thawed embryo transfer (FET) cycles were categorized into three groups based on endometrial preparation methods: hormone replacing therapy cycle (HRT, n=636), natural cycle (NC n=174), and HRT with GnRH-a pretreatment (HRT+GnRH-a, n=72. Logistic regression was performed to investigate the association between cycle regimens and pregnancy outcomes. Subgroup analysis of IUAs combined with thin endometrium (≤7mm) was also performed.Results: HRT with GnRH-a pretreatment was associated with higher incidences of clinical pregnancy, ongoing pregnancy, and live birth, but lower early miscarriage compared with either HRT or NC. Logistic regression indicated that after controlling for potential confounders, the incidences of live birth (HRT+GnRH-a as reference; NC: aOR=0.577, 95%CI 0.304-1.093; HRT: aOR=0.434, 95%CI 0.247-0.765) and ongoing pregnancy (NC: aOR=0.614, 95%CI 0.324-1.165; HRT: aOR=0.470, 95%CI 0.267-0.829) remained significantly higher in HRT+GnRH-a compared to those in HRT, but comparable to those in NC. While there was no significant difference with respect to the clinical pregnancy rate (NC: aOR=0.695, 95%CI 0.374-1.291; HRT: aOR=0.650, 95%CI 0.374-1.127) and early miscarriage rate (NC: aOR=1.734, 95%CI 0.417-7.175; HRT: aOR=2.594, 95%CI 0.718-9.378) between groups. Subgroup analysis suggested there was no priority of endometrial preparation method in IUAs combined with thin endometrium.Conclusion: HRT with GnRH-a pretreatment improves pregnancy outcomes in women with history of IUAs. GnRH-a may restore the endometrial receptivity in the FET cycles in IUAs.

Objective To report a very rare case of live birth with unexpectedly low serum hCG level on day 11 after blastocyst embryo transfer. Design Case report. Setting Private infertility center. Patient(s) A 30-year-old nulliparous woman presented with PCOS and 1 year of infertility. Interventions(s) Conventional IVF was scheduled and a long-acting agonist protocol was selected. Main Outcome Measure(s) Maternal serum hCG levels and transvaginal ultrasound exams for the embryo’s well-being. Result(s) The hCG level was 11.6 IU/L on day 11 after the transfer of two blastocyst embryos, which was considered as either failing or extrauterine pregnancy. After blood titration, there were delayed hCG increases. A series of transvaginal ultrasounds also indicated a delayed but normal-appearing intrauterine pregnancy. A healthy baby boy was delivered at term by means of cesarean section. Conclusion(s) A low initial serum hCG level may be associated with certain maternal or fetal characteristics and IVF treatment variables. Close conservative observation is warranted before undertaking any therapeutic intervention.

The aim of this study was to explore specific factors that predispose to monozygotic twinning (MZT) at the blastocyst stage. This was a retrospective observational study of a cohort of 2863 pregnancies after single blastocyst transfer (SBT) between January 2011 and June 2019 in our hospital. MZT pregnancy was identified as the number of fetuses exceeded the number of gestational sacs (GSs) by transvaginal ultrasound at 6–7 gestational weeks. The incidences of MZT regarding the maternal age at oocyte retrieval, paternal age, ovarian stimulation protocol, fertilization method, endometrium preparation protocol, vitrified day, and the Gardner grading of the blastocyst were calculated. The serum estrogen (E2), progesterone (P) levels, endometrium thickness and serum hCG levels on day 11 after embryo transfer (ET) were compared between the MZT and singleton pregnancies. Statistical analyses were used appropriately. Fifty-one MZT pregnancies (1.78%) were identified. The only significant differences observed between MZT and singleton pregnancies were the proportion of TE grade (P = 0.022) and the hCG levels on day 11 after ET (P = 0.003). Multivariate logistic regression revealed that trophectoderm (TE) grade was an independent factor affecting MZT, the adjusted odds ratios (aORs) of grade A and B TE were 5.46 [95% confidential interval (CI) 1.48–20.16, P = 0.011) and 3.96 (95% CI 1.17–13.40, P = 0.027) compared to grade C respectively. There were no significant associations between the parental age, fertilization method, ovarian stimulation protocol, endometrium preparation protocol, vitrified day, expansion stage, inner cell mass (ICM) grade and MZT. TE grade is associated with MZT at the blastocyst stage, potentially mediated via increased secretion of hCG from more well developed TE. Increased hCG secretion in turn may prolong the implantation window to support the embryo splitting.

Background: It is paramount to consider the appropriate preparation of the endometrium to receive the transferred embryos as the amount of frozen embryo transfer (FET) cycles is increasing worldwide. However, there remains lack of evidence about what is the most optimal protocol of endometrial preparation regarding pregnancy outcomes in different subgroup of infertile women. This retrospective cohort study was aim to explore the best endometrial preparation protocols among different maternal age groups.Methods: A total of 16870 FET cycles were categorized into three groups based on endometrial preparation protocols: Natural cycle (NC n=3893), artificial cycles (AC, n=11459) and AC with pretreatment with GnRH-a (AC+GnRH-a, n=1518). Logistic regression was performed to investigate the independent effect of endometrial preparation protocols on IVF pregnancy outcomes. Subgroup analyses were conducted to evaluate the most optimal endometrial preparation protocols for different maternal age groups.Results: In overall populations, after controlling for potential confounders, the incidence of live birth (NC as reference; AC: adjusted odds ratio (aOR) =0.840, 95%CI 0.774-0.912; AC+GnRHa: aOR=0.907, 95%CI 0.795-1.034) in NC was significantly higher than that of AC, while comparable to that of AC+GnRH-a. The early miscarriage rate (AC: aOR=1.413, 95%CI 1.220-1.638; AC+GnRHa: aOR=1.537, 95%CI 1.232-1.919) was significantly lower in NC compared to either AC group. In younger women, the live birth rates (AC: aOR=0.894, 95%CI 0.799-1.001; AC+GnRHa: aOR=1.111, 95%CI 0.923-1.337) were comparable between the three groups, with a slightly higher in AC+GnRH-a. Early miscarriage rate was only significantly lower in NC compared to that of AC without GnRH-a (aOR=1.452, 95%CI 1.159-1.820). While in older women, the incidence of live birth (AC: aOR=0.811, 95%CI 0.718-0.916; AC+GnRHa: aOR=0.760, 95%CI 0.626-0.923) was significantly higher, and early miscarriage (AC: aOR=1.358, 95%CI 1.114-1.655; AC+GnRHa: aOR=1.717, 95%CI 1.279-2.305) was significantly lower in NC compared to those of two AC groups.Conclusions: NC protocol is associated with lower early miscarriage late in overall IVF population. There is a mild favor of AC+GnRH-a in younger women, while the priority of NC is remarkable in older women. Maternal age should be a considerable factor when determine endometrial preparation method for FET.

PROBLEM The definition of chronic endometritis (CE) differs among studies, and currently, there is no accepted consensus. This study aimed to establish the minimum number of immunohistochemical analysis of CD138+ plasma cells to identify a clinically relevant CE. METHOD OF STUDY We performed a retrospective study on 716 infertile patients who never did CE analysis and respective antibiotic treatment before. Samples were obtained by endometrial scratching in the mid-luteal phase before IVF-ET treatment. The number and distribution of CD138+ cells were analyzed by immunohistochemistry. Thirty high-power fields (HPF) were evaluated for each sample. Patients were classified in 2 main groups: (a) CD138low (

The definition of chronic endometritis (CE) differs among studies, and currently, there is no accepted consensus. This study aimed to establish the minimum number of immunohistochemical analysis of CD138We performed a retrospective study on 716 infertile patients who never did CE analysis and respective antibiotic treatment before. Samples were obtained by endometrial scratching in the mid-luteal phase before IVF-ET treatment. The number and distribution of CD138In the CD138We conclude that immunohistochemical analysis of CD138

PURPOSE: The purpose of this study was to determine the heterotopic pregnancy rate using fresh versus frozen-thawed embryo transfers and factors associated with heterotopic pregnancy (HP). Management and clinical outcomes after heterotopic pregnancy were also evaluated. METHODS: In this retrospective cohort study, we included 12,484 women who had clinical pregnancies after in vitro fertilization treatment at our fertility center between 2012 and 2017. Patients received fresh day 3 embryos (F-D3 group), fresh day 5 blastocysts (F-D5 group), frozen-thawed day 3 embryos (T-D3 group), or frozen-thawed day 5 or 6 blastocysts (T-D5/6 groups) transfers. The primary outcome measure was the occurrence of heterotopic pregnancy. Factors associated with heterotopic pregnancy were analyzed using logistic regression. RESULTS: The heterotopic pregnancy rates were 0.58% in the F-D3, 0.39% in F-D5, 0.56% in T-D3, and 0.33% in T-D5/6 groups, but no differences were found between groups. The risk factors of HP included a history of previous ectopic pregnancy (odds ratio [OR] 5.805, 95% CI 4.578–9.553, P = 0.016) and pelvic inflammation diseases (OR 1.129, 95% CI 1.021–3.178, P = 0.047). Salpingectomy was performed in 62.9% patients either through laparoscopy or through laparotomy. The early abortion rate and late abortion rate were 29.03% and 1.61%, respectively. In total, 66.13% of the patients had a live birth, either a singleton (90.24%) or twins (9.76%). CONCLUSION: No significant difference in the incidence of heterotopic pregnancy in fresh IVF cycles vs. frozen-thawed cycles could be demonstrated in a large cohort of patients. The risk factors of HP included history of ectopic pregnancy and PID. The clinical outcome after heterotopic pregnancy appears to be favorable.

The present paper reports the structure and tribological studies of MoS x films bombarded with 75 keV Ar + at a dose of 2.1 X 10 17 Ar + cm −2 . The results show that it is possible to amorphize MoS x crystalline films and to change the film composition with Ar + bombardment. After Ar + bombardment, the sliding wear life of sputter-deposited MoS x films increases by a factor of 10, while the friction coefficient increases from 0.05 to 0.16. The mechanism of sputter-deposited MoS x films modified by ion bombardment is also discussed in this paper.

TiN films were prepared on beryllium by ion implantation. The microhardnesses and wear resistances of the TiN films were measured. Experimental results show that the TiN film has a higher microhardness and a better wear resistance at an implanted dose of 3.3×10 17 N + cm -2 . The result of X-ray diffraction confirmed that a TiN film was formed.

A high performance liquid chromatographic (HPLC) method was proposed to simultaneously determine four common nonprotein nitrogen substances, including creatine (Cr), creatinine (Cn), uric acid (Ua) and pseudouridine (Pu) in urine. After proteins being removed by acetone precipitation method, freeze drying and redissolving, the urine samples were analyzed by HPLC. Chromatographic separation was performed on a Waters RP18 Column (150 mm x 4.60 mm, 3.5 microm) in gradient elution mode using 10.0 mmol/L KH2PO4 solution (pH 4.78) and acetonitrile as mobile phases at a flow rate of 0.8 mL/min. The samples were detected at 220 nm. Rapid separation was achieved within 7 min. Under the optimized conditions, good linearities of four common nonprotein nitrogen substances were obtained in the range of 0.1-250 mg/L. The detection limits were 9.31 (Cr), 26.19 (Cn), 4.70 (Ua), an 6.30 (Pu) microg/L and the recoveries were in the range of 81%-111% with the relative standar deviations of 0.23%-2.78% (n = 3). The results demonstrate that this method is simple, rapid and accurate with good reproducibility, and can provide early diagnosis and preliminary judgment for type 2 diabetes mellitus (T2DM) patients with renal damage.