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Background: Doppler ultrasonography is used to study ovarian vascular characteristics. However, the outcomes are reported with a considerable variability in literature. Here we review the differences in Doppler ultrasound-measured ovarian blood flow indices between women with and without ovarian dysfunction and seeks correlations between Doppler measures and ovarian markers., Methods: A literature search was conducted in electronic databases (Google Scholar, Ovid, PubMed, Science Direct, and Springer) to identify studies that used Doppler for ovarian blood flow examination and reported Doppler measures in women with and without ovarian dysfunction and/or the correlations between wDoppler indices and markers of ovarian dysfunction. After quality assessment of included studies, a meta-analysis of weighted mean differences (WMDs) between women with and without ovarian dysfunction in vascularization index (VI), flow index (FI), vascularization flow index (VFI), pulsatility index (PI) and resistance index (RI) was performed. Correlation coefficients between Doppler indices and markers of ovarian dysfunction were pooled to achieve overall estimates., Results: A total of 27 studies [2,377 women with ovarian dysfunction and 308 controls; age 27.7 years, 95% confidence interval (CI): 26.4 to 29.1] were included. These studies were of moderate quality. The VI (WMD 9.75; P<0.0001), FI (WMD 2.73; P<0.0001), and VFI (WMD 1.29; P<0.0001) were significantly higher whereas PI (WMD -1.08; P=0.001) and RI (WMD -0.26; P<0.0001) were significantly lower in women with polycystic ovarian syndrome (PCOS) than in normal women. In women undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI), antral follicle count was positively correlated with VI (r=0.24; P=0.001), FI (r=0.42; P<0.0001), and VFI (r=0.25; P=0.002). In women with PCOS, testosterone had statistically non-significant correlations with VI (r=0.40; P=0.081), and VFI (r=0.39; P=0.063) and was inversely correlated with PI (r=-0.30; P<0.0001) and RI (r=-0.48; P<0.0001). In women with PCOS, luteinizing hormone (LH) was inversely correlated with PI (r=-0.26; P=0.086) and RI (r=-0.25; P=0.007)., Conclusions: Doppler indices are found significantly different in women with and without ovarian dysfunction and have significant correlations with markers of ovarian dysfunction. These results support the use of Doppler ultrasound to examine ovarian dysfunction. High statistical heterogeneity observed herein should be studies in future investigations., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://atm.amegroups.com/article/view/10.21037/atm-22-5813/coif). The authors have no conflicts of interest to declare., (2023 Annals of Translational Medicine. All rights reserved.)

Objectives: Contrast-enhanced ultrasonography (CEUS) is a modern diagnostic method that can also be used to study microperfusion. This study compared the time to peak intensity measured by CEUS in patients with peripheral arterial disease (PAD) and healthy control participants., Methods: After a comprehensive literature search in multiple electronic databases and study selection, a random-effect meta-analysis was performed to compare the time to peak intensity measured by CEUS in patients with PAD and healthy controls, which followed meta-regression analyses for identification of factors affecting the outcomes., Results: Fourteen studies (data for 322 patients with PAD and 314 healthy individuals) were used for the meta-analysis. The age of this sample of patients with PAD was 64.92 (95% confidence interval, 62.53, 67.31) years, and that of the healthy controls was 55.32 (51.67, 58.98) years. The times to peak intensity were 18.55 (15.62, 21.48) seconds in healthy controls, 33.40 (27.65, 39.15) seconds in patients with PAD, and 76.22 (36.23, 116.22) seconds in patients with PAD and diabetes mellitus. The difference between patients with PAD and healthy controls in the time to peak intensity was statistically significant (mean difference, 24.80 [10.16, 39.44] seconds; P < .00009). The ABI was not significantly associated with the time to peak intensity in patients with PAD. Age and sex were also not significantly associated with the time to peak intensity., Conclusions: Contrast-enhanced ultrasonography is a valuable tool for the diagnosis of PAD based on its ability to differentiate the time to peak intensity between patients with PAD and healthy individuals, but little data are yet available to assess its diagnostic ability in clinical practice., (© 2017 by the American Institute of Ultrasound in Medicine.)

Cervical cancer is one of the primary causes of cancer-associated mortality worldwide. Due to the increasing incidence of cervical cancer, multiple treatment options are required. Initial responses to chemotherapy and surgical interventions are generally positive, however patients often experience relapse and tumor recurrence. Currently, the effects of cucurbitacins on different types of cancer are being investigated, as they exhibit a wide variety of bioactivities. The anticancer activity of the cucurbitacin 23,24-dihydrocucurbitacin B against a panel of human cervical cancer cell lines was investigated in the current study. Cell viability was determined using an MTT assay and apoptosis was detected using DAPI staining. The proportion of apoptotic cells, cell cycle distribution, mitochondrial membrane potential (ΔΨ m ) and reactive oxygen species (ROS) levels were estimated using flow cytometry. Protein expression was determined using western blot analysis. The results of the current study indicated that 23,24-dihydrocucurbitacin B inhibited the viability of human cervical cancer cell lines and had an IC 50 of 40-60 µM. However, its cytotoxic effects were much less pronounced in normal epithelial fr2 and HerEpiC cells, where it had an IC 50 of 125 µM. The underlying mechanisms of this were further studied and the results demonstrated that 23,24-dihydrocucurbitacin B induced apoptosis in HeLa cells and caused ROS-mediated shifts in the ΔΨ m . Additionally, it caused the cell cycle arrest of HeLa cells at the G 2 /M checkpoint. The phosphoinositide 3 kinase/protein kinase B/mechanistic target of rampamycin (PI3K/AKT/mTOR) cascade may serve an important role in cancer tumorigenesis, progression and resistance to chemotherapy. The results indicated that 23,24-dihydrocucurbitacin B significantly decreased the expression of important proteins in the PI3K/Akt/mTOR cascade. Taken together, these results suggest that 23,24-dihydrocucurbitacin B may be novel method of treating cervical cancer.

A clinical validation of the bone scan lesion area (BSLA) as a quantitative imaging biomarker was performed in metastatic castration-resistant prostate cancer (mCRPC). BSLA was computed from whole-body bone scintigraphy at baseline and week 12 posttreatment in a cohort of 198 mCRPC subjects (127 treated and 71 placebo) from a clinical trial involving a different drug from the initial biomarker development. BSLA computation involved automated image normalization, lesion segmentation, and summation of the total area of segmented lesions on bone scan AP and PA views as a measure of tumor burden. As a predictive biomarker, treated subjects with baseline BSLA [Formula: see text] had longer survival than those with higher BSLA ([Formula: see text] and [Formula: see text]). As a surrogate outcome biomarker, subjects were categorized as progressive disease (PD) if the BSLA increased by a prespecified 30% or more from baseline to week 12 and non-PD otherwise. Overall survival rates between PD and non-PD groups were statistically different ([Formula: see text] and [Formula: see text]). Subjects without PD at week 12 had longer survival than subjects with PD: median 398 days versus 280 days. BSLA has now been demonstrated to be an early surrogate outcome for overall survival in different prostate cancer drug treatments.

The healing of Bone tissue consists of a complex process. Hence, we designed our study to evaluate chondrial diseases, which are as they have a very low healing capacity. Seventy two elderly osteoarthritis (OA) and 54-paediatric juvenile idiopathic arthritis (JIA) patients were included. The group was divided as 24 OA patients and 18 JIA patients in each group. Group I received Hyualuronic acid and glucocorticoides. Group II received platelet rich plasma and fibrin glue. Group III received PRP, fibrin glue, and MSC. 40 control patients received only PRP treatment. Out of 72 OA patients 35 (48.6%) male and 37 (51.4%) female with mean age of 48 ± 6.5 years. 64 (88.9%) Patients had pain and swelling. 52 (72.2%) lacked flexibility. 42 (58.3%) had hypertrophy. 28 (38.9%) had less cartilage thickness. 34 (47.2%) were in grade 3, grade 2 has 28 (38.9%) and grade 1 has 10 (13.9%) patients respectively. Among 54 JIA patients 28 (51.9%) male and 26 (48.1%) female patients with mean, age 4.6 ± 3.8 years. 39 (72.2%) had pain and swelling. 32 (59.3%) lacked flexibility. 29 (53.7%) children's had functional disability. Group I patients showed 30% improvement with no statistical significance (P < 0.21). Group II showed 45% improvement with statistical significance (P < 0.01). In Group III 80%, improvement was observed with statistical significance (P < 0.001). In 40 control patients, 60% improvement was observed. In conclusion, use of these MSC, PRP, and PPP are safe and less cost effective for treating OA and JIA.

Aim: To investigate human epidermal growth factor receptor 2 (HER2) amplification and protein expression in mixed gastric carcinoma., Methods: Fluorescence in situ hybridization and immunohistochemistry were used to detect HER2 amplification and protein expression in 277 cases of mixed gastric carcinoma. Protein staining intensity was rate as 1+, 2+, or 3+., Results: Of the 277 cases, 114 (41.2%) expressed HER2 protein. HER2 3+ staining was observed in 28/277 (10.1%) cases, 2+ in 37/277 (13.4%) cases, and 1+ in 49/277 (17.7%) cases. A HER2 amplification rate of 17% was detected, of which 25/28 (89.3%) were observed in the HER2 3+ staining group, 17/37 (45.9%) in 2+, and 5/49 (10.2%) in 1+. Of the 47 patients with HER2 amplification who received chemotherapy plus trastuzumab, 22 demonstrated median progression-free and overall survivals of 9.1 mo and 16.7 mo, respectively, which were significantly better than those achieved with chemotherapy alone (5.6 mo and 12.1 mo, respectively) in 19 previously treated patients (Ps < 0.05)., Conclusion: HER2 detection in mixed gastric carcinoma displays high heterogeneity. Relatively quantitative parameters are needed for assessing the level of HER2 amplification and protein expression.

Alkaloids are plant secondary metabolites that are widely distributed in Nicotiana species and contribute greatly to the quality of tobacco leaves. Some alkaloids, such as nornicotine and myosmine, have adverse effects on human health. To reduce the content of harmful alkaloids in tobacco leaves through conventional breeding, a genetic study of the alkaloid variation among different genotypes is required. In this study, alkaloid profiles in leaves of five Nicotiana tabacum cultivars and Nicotiana tomentosiformis were investigated. Six alkaloids were identified from all six genotypes via gas chromatograph-mass spectrometry (GC-MS). Significant differences in alkaloid content were observed both among different leaf positions and among cultivars. The contents of nornicotine and myosmine were positively and significantly correlated (R(2)=0.881), and were also separated from those of other alkaloids by clustering. Thus, the genotype plays a major role in alkaloid accumulation, indicating a high potential for manipulation of alkaloid content through traditional breeding.

Esophageal squamous cell carcinoma (ESCC) is one of the most common primary malignant tumor of digestive tract. However, the early diagnosis and molecular mechanisms that underlie tumor formation and progression have been progressed less. To identify new biomarkers for ESCC, we performed a comparative proteomic research. Isobaric tags for relative and absolute quantitation-based proteomic method was used to screen biomarkers between ESCC and normal. 802 non-redundant proteins were identified, 39 of which were differentially expressed with 1.5-fold difference (29 up-regulated and 10 down-regulated). Through Swiss-Prot and GO database, the location and function of differential proteins were analyzed, which are related to the biological processes of binding, cell structure, signal transduction, cell adhesion, etc. Among the differentially expressed proteins, TP-alpha, collagen alpha-1(VI) chain and S100A9 were verified to be upregulated in 77.19%, 75.44% and 59.65% of ESCC by immunohistochemistry and western-blot. Diagnostic value of these three proteins was validated. These results provide new insights into ESCC biology and potential diagnostic and therapeutic biomarkers, which suggest that TP-alpha, collagen alpha-1(VI) chain and S100A9 are potential biomarkers of ESCC, and may play an important role in tumorigenesis and development of ESCC., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Objective: The development and evaluation of a computer-aided bone scan analysis technique to quantify changes in tumor burden and assess treatment effects in prostate cancer clinical trials., Methods: We have developed and report on a commercial fully automated computer-aided detection (CAD) system. Using this system, scan images were intensity normalized, and then lesions were identified and segmented by anatomic region-specific intensity thresholding. Detected lesions were compared against expert markings to assess the accuracy of the CAD system. The metrics Bone Scan Lesion Area, Bone Scan Lesion Intensity, and Bone Scan Lesion Count were calculated from identified lesions, and their utility in assessing treatment effects was evaluated by analyzing before and after scans from metastatic castration-resistant prostate cancer patients: 10 treated and 10 untreated. In this study, patients were treated with cabozantinib, a MET/vascular endothelial growth factor inhibitor resulting in high rates of resolution of bone scan abnormalities., Results: Our automated CAD system identified bone lesion pixels with 94% sensitivity, 89% specificity, and 89% accuracy. Significant differences in changes from baseline were found between treated and untreated groups in all assessed measurements derived by our system. The most significant measure, Bone Scan Lesion Area, showed a median (interquartile range) change from baseline at week 6 of 7.13% (27.61) in the untreated group compared with -73.76% (45.38) in the cabozantinib-treated group (P=0.0003)., Conclusion: Our system accurately and objectively identified and quantified metastases in bone scans, allowing for interpatient and intrapatient comparison. It demonstrates potential as an objective measurement of treatment effects, laying the foundation for validation against other clinically relevant outcome measures.

A lead optimization campaign in our previously reported sulfamoyl benzamide class of CB(2) agonists was conducted to improve the in vitro metabolic stability profile in this series while retaining high potency and selectivity for the CB(2) receptor. From this study, compound 14, N-(3,4-dimethyl-5-(morpholinosulfonyl)phenyl)-2,2-dimethylbutanamide, was identified as a potent and selective CB(2) agonist exhibiting moderate in vitro metabolic stability and oral bioavailability. Compound 14 demonstrated in vivo efficacy in a rat model of post-surgical pain., (Copyright 2009 Elsevier Ltd. All rights reserved.)

A series of imidazopyrimidine derivatives with the general formula I was synthesized and identified as potent inhibitors of iNOS dimer formation, a prerequisite for proper functioning of the enzyme. Stille and Negishi coupling reactions were used as key steps to form the carbon-carbon bond connecting the imidazopyrimidine core to the central cycloalkenyl, cycloalkyl and phenyl ring templates.

Replacement of the phenyl ring in our previous (morpholinomethyl)aniline carboxamide cannabinoid receptor ligands with a pyridine ring led to the discovery of a novel chemical series of CB2 ligands. Compound 3, that is, 2,2-dimethyl-N-(5-methyl-4-(morpholinomethyl)pyridin-2-yl)butanamide was identified as a potent and selective CB2 agonist exhibiting in vivo efficacy after oral administration in a rat model of neuropathic pain.

Selective, nonpeptidic delta opioid receptor agonists have been the subject of great interest as potential novel analgesic agents. The discoveries of BW373U86 (1) and SNC80 (2) contributed to the rapid expansion of research in this field. However, poor drug-like properties and low therapeutic indices have prevented clinical evaluation of these agents. Doses of 1 and 2 similar to those required for analgesic activity produce convulsions in rodents and nonhuman primates. Recently, we described a novel series of potent, selective, and orally bioavailable delta opioid receptor agonists. The lead derivative, ADL5859 (4), is currently in phase II proof-of-concept studies for the management of pain. Further structure activity relationship exploration has led to the discovery of ADL5747 (36), which is approximately 50-fold more potent than 4 in an animal model of inflammatory pain. On the basis of its favorable efficacy, safety, and pharmacokinetic profile, 36 was selected as a clinical candidate for the treatment of pain.

Selective delta opioid receptor agonists are promising potential therapeutic agents for the treatment of various types of pain conditions. A spirocyclic derivative was identified as a promising hit through screening. Subsequent lead optimization identified compound 20 (ADL5859) as a potent, selective, and orally bioavailable delta agonist. Compound 20 was selected as a clinical candidate for the treatment of pain.

Nitric oxide (NO), a mediator of various physiological and pathophysiological processes, is synthesized by three isozymes of nitric oxide synthase (NOS). Potential candidate clinical drugs should be devoid of inhibitory activity against endothelial NOS (eNOS), since eNOS plays an important role in maintaining normal blood pressure and flow. A new series of aminopiperidines as potent inhibitors of iNOS were identified from a HTS lead. From this study, we identified compound 33 as a potent iNOS inhibitor, with >25-fold selectivity over eNOS and 16-fold selectivity over nNOS.

A novel series of malonamide derivatives was synthesized. These amides were shown to be potent and selective kappa opioid receptor agonists.

A novel series of phenylamino acetamide derivatives was synthesized. These amides were shown to be potent and selective kappa opioid receptor agonists.

Two novel chemical classes of kappa opioid receptor agonists, chroman-2-carboxamide derivatives and 2,3-dihydrobenzofuran-2-carboxamide derivatives, were synthesized. These agents exhibited high and selective affinity for the kappa opioid receptor.

We report the synthesis and radioligand binding analysis of a series of naphthalenic melatonin receptor ligands, N-[2-(7-alkoxy-2-methoxy-1-naphthyl)ethyl]propionamide. This series of ligands exhibits subpicomolar binding affinity to both MT1 and MT2 melatonin receptors expressed in chinese hamster ovary (CHO) cells.

We report the development of (E)- and (Z)-4-hydroxytamoxifen sulfamates as estrone sulfatase inhibitors, potential therapeutic agents for the treatment of breast cancer. Both compounds competitively inhibit estrone sulfatase isolated from rat liver with apparent Ki of 35.9 microM for (E)-4-hydroxytamoxifen sulfamate and an apparent Ki of > 500 microM for the (Z) isomer.

Estrogen levels in breast tumors of postmenopausal women are as much as 10 times higher than estrogen levels in plasma, presumably due to in situ formation of estrogen. The major source of estrogen in breast cancer cells may be conversion of estrone sulfate to estrone by the enzyme estrone sulfatase. Thus, inhibitors of estrone sulfatase are potential agents for treatment of estrogen-dependent breast cancer. Several steroidal compounds have been developed that are potent estrone sulfatase inhibitors, most notably estrone-3-O-sulfamate. However, these compounds and their metabolites may have undesired effects, including estrogenicity. To avoid the problems associated with a potentially active steroid nucleus, we designed and synthesized a series of nonsteroidal estrone sulfatase inhibitors, the (p-O-sulfamoyl)-N-alkanoyl phenylalkyl amines. The compounds synthesized vary in the length of their alkanoyl chain and in the number of carbons separating the phenyl ring and the carbonyl carbon. The ability of these compounds to inhibit estrone sulfatase activity was tested using human placental microsomes and intact cultured human breast cancer cells. Estrogenicity was also evaluated, using growth of estrogen-dependent human breast cancer cells. All of the test compounds inhibited estrone sulfatase activity of human placental microsomes to some extent, with the most effective compound having an IC50 value of 72 nM. In general, compounds with longer alkanoyl chains (12-14 carbons) were more effective than those with shorter chains. The test compounds also inhibited estrone sulfatase activity in intact cultures of MDA-MB-231 human breast cancer cells. Again, the longer chain compounds were more effective. In both the placental and breast cancer cell sulfatase assays, the optimal distance between the phenyl ring and the carbonyl carbon was 1-2 carbons. The MCF-7 cell proliferation assay revealed that estrone and estrone-3-O-sulfamate were both estrogenic, but the (p-O-sulfamoyl)-N-alkanoyl phenylalkyl amines were not. Our data indicate the utility of (p-O-sulfamoyl)-N-alkanoyl phenyl alkylamines for inhibition of estrone sulfatase activity. Furthermore, our data support the concept that nonsteroidal estrone sulfatase inhibitors may be useful as therapeutic agents for estrogen-dependent breast cancers.

Estrogen levels in breast tumors of post-menopausal women are as much as 10 times higher than estrogen levels in plasma, presumably due to in situ formation of estrogen. The major source of estrogen in breast cancer cells may be conversion of estrone sulfate to estrone by the enzyme estrone sulfatase. Thus, inhibitors of estrone sulfatase have potential for the treatment of estrogen-dependent breast cancers. Several steroidal agents have been developed that are potent estrone sulfatase inhibitors, most notably estrone-3-O-sulfamate. These compounds may have undesired actions, especially estrogenicity. Recently, non-steroidal estrone sulfatase inhibitors have been designed that avoid the problems associated with an active steroid nucleus; however, these have not achieved the potency of estrone-3-O sulfamate. We have designed and synthesized a series of compounds, 17 beta-(N-alkylcarbamoyl)-estra-1,3,5(10)-trien-3-O-sulfamates (6a-d) and 17 beta-(N-alkanoyl)-estra-1,3,5(10)-trien-3-O-sulfamates (11a-d) that combine the structural features of the steroidal estrone sulfatase inhibitors with a membrane insertion region that should increase the affinity for the sulfatase enzyme and decrease the estrogenicity of the steroid. We tested the compounds for estrone sulfatase inhibition by measuring estrone sulfatase activity in intact cultures of human breast cancer cells (MDA-MB-231). We tested for estrogenicity by measuring growth of estrogen-dependent MCF-7 human breast cancer cells. All of the test compounds (10 nM) substantially inhibited estrogen sulfatase activity of intact MDA-MB-231 cells. Dose-response analysis indicated an IC50 of approximately 0.5 nM for two of the compounds (6a and 11a). In the test for estrogenicity, estrone and estrone-3-O-sulfamate significantly stimulated MCF-7 cell growth. In contrast, neither the 17 beta-(N-alkylcarbamoyl)-estra-1,3,5,(10)-trien-3-O-sulfamates++ + nor the 17 beta-(N)-alkanoyl)-estra-1,3,5,(10)-trien-3-O-sulfamates stimulated growth of MCF-7 cells at a concentration of 1 microM, indicating that they are not estrogenic at levels 2000 times greater than their IC50 for estrone sulfatase. Our data indicate the utility of the new compounds for inhibition of breast cancer cell estrone sulfatase activity. Further, our data support the concept that estrone sulfatase inhibitors may be useful as therapeutic agents for estrogen-dependent breast cancers.

Recently, we reported the synthesis and biomedical studies of a series of (p-O-sulfamoyl)-N-alkanoyl tyramines as nonsteroidal estrone sulfatase inhibitors. One of the most potent inhibitors in this series is (p-O-sulfamoyl)-N-tridecanoyl tyramine 1 with an 1C50 value of 61.3 nM. In this study, we synthesized four analogs of 1 (compounds 2-5) to investigate the structure-activity relationships of the amide functionality in (p-O-sulfamoyl)-N-tridecanoyl tyramine. Replacement of the amide functionality in 1 with an ethylene moiety to form the alkyl analog 5 resulted in complete loss of sulfatase inhibitory activity (IC50 of 61.3 nM vs. > 20 microM). The keto, hydroxy, and ester analogs (inhibitors 2-4) are 8-15 times less in affinity to the sulfatase than inhibitor 1. However, their inhibitory activities are significantly higher than the alkyl analog 5. The results suggest that the amide functionality is favorable for sulfatase inhibitory activity and that there may be a hydrogen bonding component to the enzyme interaction in this region.

The synthesis of sodium androst-5-ene-17-one-3 beta-methylene sulfonate 2, a stable analog of memory-enhancing neurosteroid dehydroepiandrosterone sulfate, is described. The synthesis of compound 2 is carried out in six steps from dehydroepiandrosterone.

The presence of transforming growth factor-beta (TGF-beta) at the site of acute injury, its ability to attract inflammatory and connective tissue cells, and its stimulatory effect on the deposition of connective tissue matrix combine to suggest that it may play a key role in the response to injury. The effect of exogenous TGF-beta form 2 on dermal wounds healing by secondary intent was investigated using a sponge composed of collagen and heparin as a delivery vehicle. Longitudinal lenticular-shaped wounds on the dorsum of adult guinea pigs were treated at the time of wounding with delivery vehicle containing 0.5, 1, or 5 micrograms of purified, bovine bone-derived TGF-beta 2, and were compared with wounds that received vehicle only or were untreated. At days 8 and 14 the amount of connective tissue in the wounds and the extent of epithelialization were determined by histomorphometric methods, and wound breaking strength was determined. At day 8, but not at day 14, wounds treated with 1 or 5 micrograms of TGF-beta 2 contained a significantly higher proportion of connective tissue than did wounds treated with vehicle only, and they also exhibited higher wound strength. No effect on wound size or re-epithelialization was detected. The observations provide evidence that a single treatment with exogenous TGF-beta 2 delivered in collagen/heparin sponge vehicle can accelerate repair in guinea pig dermal wounds allowed to heal by secondary intent.

Collagen has been prepared from steer glomerular basement membrane by controlled pepsin solubilization. Four collagen polypeptides of potential alpha chain size have been isolated following denaturation and reduction with mercaptoethanol. Purification was obtained through sequential chromatography by gel filtration on agarose and by ion exchange on carboxymethyl- and diethylaminoethylcellulose and confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Fractions A and B resemble interstitial alpha chains in apparent molecular weight by gel filtration (93,000). Fractions C and D were released from a single high molecular weight fraction (II) by reduction with mercaptoethanol and had a larger apparent molecular weight by gel filtration (140,000). Amino acid composition of all fractions demonstrated that they are closely interrelated by generally distinctive from interstitial collagens. Cysteine was present in all fractions except Fraction A. Prior to reduction, all mercaptan groups were inaccessible to iodoacetate and p-chloromercuribenzoate but became completely titrable after treatment with mercaptoethanol. The starting material and all fractions contained large amounts of hexose. Glucose and galactose predominated; but mannose, glucosamine, and galactosamine were also present in substantial amounts. Small amounts of fucose and sialic acids were found in starting material only.

The white muscle of the Sacramento blackfish (Orthodon microlepidotus) was processed by freezing, dehydration, and cooking. Myosin was extracted immediately afterwards or following a period of storage in order to examine evidence for denaturation. The tests used were the solubility of whole muscle protein and the intrinsic viscosity, isoelectric point, ATPase activity, ultra-violet absorption spectrum, and optical rotatory dispersion of purified myosin extract. Almost all measures used showed that denaturation increased in the order: fresh < frozen < frozen-stored < dehydrated < dehydrated-stored < cooked.