Liquid Nitrogen Treatment Research Articles

Laboratory investigation on cryogenic fracturing of hot dry rock under triaxial-confining stresses

Hot dry rock (HDR) has substantial potential as a thermal energy source. It can be exploited by hydraulic fracturing to extract heat and generate electricity. Because of its great depth (2–6 km) and consequent high temperature (150–650°C) and high pressure, the stimulation of HDR is difficult. The purpose of this study is to study the feasibility of cryogenic fracturing on HDR. We conducted a series of laboratory experiments on rock samples under triaxial-confining stresses. Two schemes were utilized: gas fracturing with and without liquid nitrogen (LN) (−196°C) treatment. For the LN treatment, high-temperature granitic specimens were immersed in LN for several hours before the fracturing tests. Three critical factors were considered, namely, rock specimen temperature (100–600°C), axial stress (5–20 MPa), and lateral stress (5–20 MPa). Subsequently, the fracture patterns were identified and correlations for both LN rapid-cryogenic and natural-cryogenic stimulations were developed to predict the breakdown pressure based on the experimental data. Finally, the field applications for LN cryogenic stimulation in HDR reservoir were discussed and a thermal-hydraulic coupled transient numerical model was established to provide suggestions of LN injection schedule for an effective cryogenic stimulation. The results show that the LN cryogenic stimulation could reduce the breakdown pressure level by 9–51% more than that of the untreated specimens. Particularly, LN provides superior fracturing efficiency on rock specimens above 200°C. This could be attributed to a sharp thermal gradient induced by LN, which can cause the strong local tensile stress to expand the micro-pore structures inside the high-temperature rock samples. Furthermore, with the assistance of LN, larger fracture apertures and better-connected fracture networks inside the rocks can be generated. Additionally, LN treatment, or rapid cryogenic fracturing, plays an increasingly important role when high-temperature rocks are subjected to higher anisotropic stresses. The key findings of this study could provide an efficient approach to explore HDR resources for geothermal development.

One minute from pristine carbon to an electrocatalyst for hydrogen peroxide production

An excellent electrocatalyst for producing H2O2was prepared from pristine CMK3 by microwave and subsequent liquid nitrogen treatments in 1 minute.

Enhanced Removal of Antibiotic in Wastewater Using Liquid Nitrogen-Treated Carbon Material: Material Properties and Removal Mechanisms.

Antibiotic residues in the aquatic environment have become a global problem posing a serious threat to the environment and an inherent health risk to human beings. In this study, experiments were carried to investigate the use of carbon material modified by liquid nitrogen treatment (CM1) and carbon material unmodified by liquid nitrogen treatment (CM2) as adsorbents for the removal of the antibiotic ampicillin from aqueous solutions. The properties of the CMs (CM1 and CM2) and the effects of variations of the key operating parameters on the removal process were examined, and kinetic, isothermal and thermodynamic experimental data were studied. The results showed that CM1 had larger specific surface area and pore size than CM2. The ampicillin adsorption was more effective on CM1 than that on CM2, and the maximum adsorption capacity of ampicillin onto CM1 and CM2 was 206.002 and 178.423 mg/g, respectively. The kinetic data revealed that the pesudo-second order model was more suitable for the fitting of the experimental kinetic data and the isothermal data indicated that the Langmuir model was successfully correlated with the data. The adsorption of ampicillin was a spontaneous reaction dominated by thermodynamics. In synthetic wastewater, CM1 and CM2 showed different removal rates for ampicillin: 92.31% and 86.56%, respectively. For an adsorption-based approach, carbon material obtained by the liquid nitrogen treatment method has a stronger adsorption capacity, faster adsorption, and was non-toxic, therefore, it could be a promising adsorbent, with promising prospects in environmental pollution remediation applications.

Changes of soluble sugars and ATP content during DMSO droplet freezing and PVS3 droplet vitrification of potato shoot tips

The potato's great genetic diversity needs to be maintained for future agricultural applications and can be preserved at ultra-low temperatures. To decipher detailed physiological processes, the aim of the study was to analyze the regrowth in 28 gene bank accessions and to reveal metabolite changes in a subset of four accessions that showed pronounced differences after shoot tip cryopreservation using DMSO droplet freezing and PVS3 droplet vitrification. Regrowth varied in all 28 genotypes ranging from 5% (‘Kagiri’) to 100% (‘Karakter’) and was higher after PVS3 droplet vitrification (71 ± 19%) than after cryopreservation using DMSO (54 ± 17%). Sucrose, glucose, and fructose were analyzed and showed significant increases after pre-culture in combination with PVS3 or DMSO and liquid nitrogen treatment and were reduced during regeneration. In contrast, adenosine triphosphate (ATP) reached its minimum concentration after cryoprotection and liquid nitrogen treatment and recovered most quickly after PVS3 droplet vitrification. A shortening of the explant pre-culture period reduced dramatically the regrowth after PVS3 vitrification. However, correlations between the shoot tip regrowth and sugar concentration were absent and significant at a low extent with ATP (r = 0.4, P < 0.01). Interestingly, several sub-cultivations of the donor plants from the previous stock affected negatively the regrowth. In conclusion, the cryopreservation protocol, genotypes, pre-culture period and number of sub-cultures affect the regrowth ability of explants, which was best estimated by the ATP concentration after low-temperature treatment. Due to the superior performance of PVS3, the routine potato cryopreservation at the Gatersleben gene bank was changed to PVS3 droplet vitrification.

Improving Impregnation and Penetration Properties of Refractory Woods Through Cryogenic Treatment

Cryogenic treatment via liquid nitrogen (LN) was evaluated as a means to improve the capability of impregnation and penetration in fir and spruce wood. There are a few vital problems that occur during the impregnation process of wood protection. One problem is that refractory woods have different anatomical features that make the impregnation process difficult. For the specimens that were conditioned with the air and oven, the retention changes were determined with an image analysis, scanning electron microscopy (SEM), and energy dispersive X-ray analysis (EDX). In addition, the density and equilibrium moisture content (EMC) values were measured. The LN treatment resulted in degradation in the bordered pits. Then, the margo and torus bonds were torn, the bordered pits were opened, and the liquid transportation of wood was facilitated. It was determined that the LN treatment and impregnation increased the depth and dispersion capability of the penetration. The LN treatment helped facilitate the flows from pores to pores by hindering the aspirations of the bordered pits, and thus the flow of chemicals was facilitated. Obtaining deeper penetration in refractory wood species will result in an improved impregnation process.

The effects of liquid nitrogen treatment on the some physical and mechanical properties of scots pine and oriental spruce wood

The wood materials widely used from wood construction sector, urban furniture and interior design to furniture sector are renewable resource. Wood has some disadvantages regarding durability to combustion and biological factors. To increase service life especially in urban furniture are coated and/or impregnated against to fire and biological factors. Thus, the wood materials used in construction sector should be treated with fire-retardant chemicals. In this study; Scots pine and Oriental spruce wood materials were exposed to liquid nitrogen for 30 minutes before impregnated with Firetex by using pressure-vacuum method. According to the results, amount of retention in Oriental spruce and Scots pine samples have been increased with liquid nitrogen treatment. Additionally, it was seen that the liquid nitrogen operation had not significant adverse effect on the mechanical properties. Therefore, the liquid nitrogen treatment can apply effectively as a method to improve the impregnability of the wood.

Hierarchical porous nitrogen-doped carbon beads derived from biosourced chitosan polymer

Carbon microbeads with diameters of a few hundreds of microns are produced by dropping chitosan acetate (CA) solution in liquid nitrogen, freeze drying and thermal treatment in inert atmosphere at 700 °C. The structure of the beads is characterized by hierarchical porosity at the three length scales: (i) macropores originating from the phase separation during the water ice crystal grow inside the droplets, immersed in liquid nitrogen, (ii) mesoporores arising in course of the beads thermal treatment, leading to the removal of the porogen molecules (Pluronic F127) added to the CA solution and (iii) micropores induced by the carbonization of chitosan resin. The carbon beads have a rigid skin which present wrinkles with well-defined structure.The carbon bead pore size and volume could be tuned with glyoxal and glyoxylic acid which are able to cross-link in a different manner with chitosan as showed by 13C NMR. The beads are produced by a green process employing entirely biosourced and renewable materials that make them attractive for the large scale applications requiring nanoporous carbons.

Modulation of anthraquinones and phloroglucinols biosynthesis in Hypericum spp. by cryogenic treatment

Beside the high post-cryogenic recovery rate, a reinstated scale of secondary metabolites in recovered plant tissues represents another inevitable aspect of an effective cryopreservation protocol for medicinal plants. The current study was aimed at evaluation of the elicitation potential of cryogenic treatment on secondary metabolism of some Hypericum species. In agreement with our assumption, the cold stimuli applied during the pre-cryogenic phase increased the tolerance to low temperatures (−196°C) in H. perforatum, H. rumeliacum and H. tetrapterum reaching a maximum of 46% recovery rate in St. John’s wort plants. The effect of cryogenic treatment-associated stressors on the spectrum of the profiling secondary metabolites, naphthodianthrones and phloroglucinols, was ambiguous. The content of hypericins in both pre-cultured H. tetrapterum donor plants and H. perforatum shoots regenerated from cryopreserved meristems increased more than 3-times. The highest 38-fold enhancement of phloroglucinols was observed in H. rumeliacum shoots recovered after cryostorage. Our findings indicate that modulated biosynthesis of secondary metabolites represented by naphtodianthrones and phloroglucinols can be considered as a part of overall plant adaptations to stress conditions associated with liquid nitrogen (LN) treatment.

Freezing Nitrogen Ethanol Composite May be a Viable Approach for Cryotherapy of Human Giant Cell Tumor of Bone.

Liquid nitrogen has been used as adjuvant cryotherapy for treating giant cell tumor (GCT) of bone. However, the liquid phase and ultrafreezing (-196°C) properties increase the risk of damage to the adjacent tissues and may lead to perioperative complications. A novel semisolid cryogen, freezing nitrogen ethanol composite, might mitigate these shortcomings because of less-extreme freezing. We therefore wished to evaluate freezing nitrogen ethanol composite as a coolant to determine its properties in tumor cryoablation. (1) Is freezing nitrogen ethanol composite-mediated freezing effective for tumor cryoablation in an ex vivo model, and if yes, is apoptosis involved in the tumor-killing mechanism? (2) Does freezing nitrogen ethanol composite treatment block neovascularization and neoplastic progression of the grafted GCTs and is it comparable to that of liquid nitrogen in an in vivo chicken model? (3) Can use of freezing nitrogen ethanol composite as an adjuvant to curettage result in successful short-term treatment, defined as absence of GCT recurrence at a minimum of 1year in a small proof-of-concept clinical series? The cryogenic effect on bone tissue mediated by freezing nitrogen ethanol composite and liquid nitrogen was verified by thermal measurement in a time-course manner. Cryoablation on human GCT tissue was examined ex vivo for effect on morphologic features (cell shrinkage) and DNA fragmentation (apoptosis). The presumed mechanism was investigated by molecular analysis of apoptosis regulatory proteins including caspases 3, 8, and 9 and Bax/Bcl-2. Chicken chorioallantoic membrane was used as an in vivo model to evaluate the effects of freezing nitrogen ethanol composite and liquid nitrogen treatment on GCT-derived neovascularization and tumor neoplasm. A small group of patients with GCT of bone was treated by curettage and adjuvant freezing nitrogen ethanol composite cryotherapy in a proof-of-concept study. Tumor recurrence and perioperative complications were evaluated at a minimum of 19months followup (mean, 24months; range, 19-30months). Freshly prepared freezing nitrogen ethanol composite froze to -136°C and achieved -122°C isotherm across a piece of 10±0.50-mm-thick bone with a freezing rate of -34°C per minute, a temperature expected to meet clinical tumor-killing requirements. Human GCT tissues revealed histologic changes including shrinkage in morphologic features of multinucleated giant cells in the liquid nitrogen (202±45μm; p=0.006) and freezing nitrogen ethanol composite groups (169±27.4μm; p<0.001), and a decreased nucleated area of neoplastic stromal cells for the 30-second treatment. Enhanced counts of terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)-positive cells verified the involvement of DNA fragmentation in cryoablated GCT tissues. Western blotting analysis on the expression of apoptosis regulatory proteins showed enhancement of proteocleavage-activated caspases 3, 8, and 9 and higher ratios of Bax/Bcl2 in the liquid nitrogen- and freezing nitrogen ethanol composite-treated samples. Numbers of blood vessels and human origin tumor cells also were decreased by freezing nitrogen ethanol composite and liquid nitrogen treatment in the GCT-grafted chicken chorioallantoic membrane model. Seven patients with GCT treated by curettage and adjuvant cryotherapy by use of freezing nitrogen ethanol composite preparation had no intra- or postoperative complications related to the freezing, and no recurrences during the study surveillance period. These preliminary in vitro and clinical findings suggest that freezing nitrogen ethanol composite may be an effective cryogen showing ex vivo and in vivo tumor cryoablation comparable to liquid nitrogen. The semisolid phase and proper thermal conduction might avoid some of the disadvantages of liquid nitrogen in cryotherapy, but a larger clinical study is needed to confirm these findings. Level IV, therapeutic study.

Novel assembly of homogeneous reduced graphene oxide-doped mesoporous TiO2 hybrids for elimination of Rhodamine-B dye under visible light irradiation

Mesoporous TiO2 nanocrystals were hydrothermally synthesized and then dispersed in graphene oxide (GO) solution through ultrasonicated treatment. Visible light photocatalysts incorporating TiO2 nanocrystals and reduced GO (TiO2-rGO) were obtained by subsequent rapid solidification with liquid nitrogen, freeze-drying and heat treatment in flowing nitrogen. The resulting samples were characterized by thermogravimetric analysis (TGA), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), transmission election microscopy (TEM), Fourier transform infrared (FT-IR) spectroscopy, Raman spectroscopy, diffuse reflectance ultraviolet–visible (DR UV–Vis) spectroscopy and nitrogen adsorption-desorption. Their adsorption capacity and visible light photocatalytic degradation activity in the elimination of rhodamine B (RhB) were evaluated at room temperature. The results showed that the mesoporous TiO2 nanoparticles were well dispersed onto rGO, leading to the enhanced adsorption of RhB and absorption of visible light. Consequently, the resulting TiO2-rGO hybrids showed outstanding efficiency for the elimination of RhB. Furthermore, the content of rGO had substantial influences on the photocatalytic performance of hybrids, which was the best for the hybrid with a content of rGO of 20% [TiO2-rGO(20)]. The adsorptive efficiency in the elimination of RhB dye with an initial concentration of 20.0 mg L−1 was as high as 66.7% and the photo-degradation efficiency may reach up to 81% under visible light irradiation over 10.0 mg of TiO2-rGO(20).

Changes in transcript expression patterns as a result of cryoprotectant treatment and liquid nitrogen exposure in Arabidopsis shoot tips.

Transcripts related to abiotic stress, oxidation, and wounding were differentially expressed in Arabidopsis shoot tips in response to cryoprotectant and liquid nitrogen treatment. Cryopreservation methods have been implemented in genebanks as a strategy to back-up plant genetic resource collections that are vegetatively propagated. Cryopreservation is frequently performed using vitrification methods, whereby shoot tips are treated with cryoprotectant solutions, such as Plant Vitrification Solution 2 (PVS2) or Plant Vitrification Solution 3 (PVS3); these solutions remove and/or replace freezable water within the meristem cells. We used the model system Arabidopsis thaliana to identify suites of transcripts that are up- or downregulated in response to PVS2 and PVS3 treatment and liquid nitrogen (LN) exposure. Our results suggest that there are many changes in transcript expression in shoot tips as a result of cryoprotection and that these changes exceed the number detected as a result of LN exposure. In total, 180 transcripts showed significant changes in expression level unique to treatment with either the cryoprotectant or cryopreservation followed by recovery. Of these 180 transcripts, 67 were related to stress, defense, wounding, lipid, carbohydrate, abscisic acid, oxidation, temperature (cold/heat), or osmoregulation. The responses of five transcripts were confirmed using qPCR methods. The transcripts responding to PVS2+LN suggest an oxidative response to this treatment, whereas the PVS3+LN treatment invoked a more general metabolic response. This work shows that the choice of cryoprotectant can have a major influence on the patterns of transcript expression, presumably due to the level and extent of stress experienced by the shoot tip. As a result, there may be divergent responses of study systems to PVS2 and PVS3 treatments.

Validated method for quality assessment of henna (Lawsonia inermis L.) leaves after postharvest blanching and its cosmetic application

Henna (Lawsonia inermis L., Family- Lythraceae) has been used for number of cosmetic purposes, including body painting, palm colouring and dyeing of hair. In spite of huge demand for cosmetic applications, no validated analytical method is available for the quality assessment of henna. Therefore, present validated method demonstrates the simultaneous quantification of eight marker compounds. Chemical markers, namely 1) gallic acid, 2) quercetin, 3) fraxetin, 4) luteolin-7-O- glucoside, 5) p-coumaric acid, 6) lawsone, 7) luteolin and 8) apigenin were chosen for the quality assessment of henna. The method was also applied to assess the effects of postharvest treatments (PHT) on the quality of henna leaves as well its derived product i.e. oil mixed paste. Both the chemical and thermal blanching treatments severely (p<0.5) changed the content of targeted secondary metabolites (1–8). The contents of fraxetin and lawsone were found to increase on deep freezing, remaining six phytochemicals reduced significantly on blanching. Luteolin-7-O- glucoside in henna was most susceptible to both salt and liquid nitrogen treatment. Additionally, the effects of mixing of oils (olive, castor oil, and mentha) on modulation of chemical markers and color intensity over palm were also evaluated. We observed a significant increase in the color intensity attributed to mixing of castor>>menthol>olive oil. The apigenin content was about 2.4 times higher in olive oil mixed henna paste than control, while, fraxetin content reduced to half. Mixing of castor oil in henna paste has produced the most intense color; while the mentha oil facilitated the persistence action when applied for palm ornamentation.

A combined electrochemical route to fabricate large-area and free-standing inverse opaline film

A combined electrochemical route involving electrophoresis and electrodeposition is developed to fabricate a large-area unsupported nickel inverse opaline film with thickness less than 10μm. A vertical electrophoresis allows for the packing of 495nm polystyrene microspheres on an ITO-coated glass to form a colloidal crystal with significantly reduced defects. Subsequently, a nickel electrodeposition is employed to fill the interstitial voids among the close-packed polystyrene microspheres, followed by the removal of the colloidal template in a two-stage process, leaving a robust nickel skeleton with hexagonally arranged pores and interconnected pore channels. This nickel skeleton is then detached from the ITO-coated glass via a liquid nitrogen treatment, rendering a free-standing nickel inverse opaline film in 2×2cm2. Porometer measurements indicate a narrow pore size distribution consistent with images from scanning electron microscope. We determine that the intensity ratio of (111)/(100) diffraction peak is an indicator for the strength of the Ni inverse opals, and thus affects its structural integrity upon detachment from the ITO-coated glass.

The Effect of Liquid Nitrogen on Bone Graft Survival.

Liquid nitrogen is used in medicine for cancer treatment and tissue preservation; however, bone viability after its application is controversial. This study aims to evaluate both the tissue viability and the clinical and histopathologic findings following liquid nitrogen application with different thawing techniques in rats. Mandibular bone grafts were taken from 45 Wistar rats and freezed in liquid nitrogen for 20 minutes. In the rapid-thawing technique (Rapid Thawing-1, Rapid Thawing-2), the grafts were held for 20 minutes in room temperature; in the slow-thawing technique (Slow Thawing-1, Slow Thawing-2), 20 minutes in -20°C, 20 minutes in +4°C, and 20 minutes in room temperature, respectively. In Rapid Thawing-2 and Slow Thawing-2 groups, autografts were implanted to their origin for 3 weeks and bone staining with India ink was performed and samples taken for histologic examination. The amount of cells and blood vessels and the density of bone canaliculi were significantly reduced in Rapid Thawing-1 and Slow Thawing-1 groups comparing to the Control group. However, the reduction rate was more significant in the Slow Thawing-1 group. Histomorphometric evaluation of the healing autografts after 3 weeks revealed that the decreased amounts of canaliculi were not changed in Slow Thawing-2 group. The study results demonstrated that bone tissue survives after liquid nitrogen treatment regardless of the performed thawing technique; however, slow thawing causes more tissue damage and metabolism impairment.

Elimination of chrysanthemum stunt viroid and chrysanthemum chlorotic mottle viroid from infected chrysanthemum by cryopreservation

Chrysanthemum morifolium 'Borami' and 'Secret Pink' showing symptoms of stunt disease caused by chrysanthemum stunt viroid (CSVd) and 'Yellow Cap' showing chlorotic mottle disease caused by chrysanthemum chlorotic mottle viroid (CChMVd) were confirmed to be infected by the respective viroids by using reverse transcription polymerase chain reaction (RT-PCR). Real-time PCR results showed that the viroid concentrations in the infected cultivars varied between the different regions of origin (Chilgok, Gumi, and Gyeongsan). We applied a cryopreservation protocol for elimination of CSVd from naturally infected 'Borami' collected from Gumi, showing the lowest concentration of CSVd, by varying several factors such as plant vitrification solutions (PVS2 and PVS3), duration of exposure to liquid nitrogen, shoot-tip size, and low-temperature treatment. The solution (PVS2) and low-temperature treatment were found to be critical factors determining the efficacy of viroid elimination. We optimized the protocol by combining of all resulted optimal factors and tested the applicability of the protocol in 'Borami' collected from Chilgok and Gyeongsan and in 'Secret Pink' from Chilgok, Gumi, and Gyeongsan, which displayed different viroid concentrations. We found that the elimination rates varied depending on the cultivar and region of origin. Similar results were observed when the protocol was applied to eliminate CChMVd from the 'Yellow Cap' collected from the same regions. Finally, we found that nested PCR is more reliable for viroid detection than RT-PCR. Overall, cryopreservation can be used to eliminate viroids from infected chrysanthemums; however, the efficacy depends on genotype and initial viroid concentration.

Synthesis and functionalization of green carbon as a Pt catalyst support for the oxygen reduction reaction

This study investigates the synthesis and characterization of a carbon derived by a simple combustion process of natural camphor. Resulted carbon was subjected to a chemical (HNO3 and KOH) and physical treatment (thermal treatment –TT, liquid Nitrogen treatment – LN2) for surface functionalization. Synthesized carbon materials were used as Pt catalyst support for the oxygen reduction reaction (ORR) in acid media. Physicochemical characterization of the carbon support was performed by multiple techniques such as, SEM, EDAX, FTIR, and Raman spectroscopy. Results conducted to spherical particles of camphor carbon (CC) with 50–60 nm in size. The chemical composition revealed a CC without sulfur content. This could be an advantage for avoiding the sulfur poisoning on the Pt surface. FTIR results shows that the oxidative chemical treatment (HNO3 and KOH) incorporate functional group of CO to CC. However, the TT eliminates the functional surface group on the CC. Apparently CC and CC-LN2 samples have the same behaviour, consequently it can be inferred that the cryogenic treatment with liquid nitrogen does not significantly modify the functional chemistry of carbon surface. Raman spectroscopy reveals the disorder in the CC (ID/IG = 1.01) which is lesser than the Vulcan Carbon (ID/IG = 1.16) defect. Furthermore, the chemically treated CC and LN2 shows fewer defects than the thermally treated CC. Electrochemical results show that materials with enhanced specific catalytic activity toward the ORR follow the tendency: Pt/CC-KOH > Pt/CC-TT > Pt/CC > Pt/CC-LN2 > Pt/Vulcan > Pt/C Etek > Pt/CC-HNO3. Our finding results open a new avenue for carbon from natural source as an effective ORR catalyst support.

Hiperplasia multiepitelial focal: tratamiento comparativo, ácido glicirricínico contra nitrógeno líquido

In pediatric population, focal multiepithelial hyperplasia (FMH) is characterized by the onset of multiple lesions in the mucosa of the mouth; it is associated to the human papillomavirus. Presently, no special treatment is preferred, the most widely used is liquid nitrogen (LN) treatment, nevertheless, this is an uncomfortable and painful treatment for the patient. This would prompt us to look for new, less invasive treatment alternatives such as use of glycyrrhizinic acid (GA). Aim: To compare GA efficiency versus LN in FMH in Mexico Children's Hospital patients (HIM) aged 5 to 13 years. To determine clinical epidemiological characteristics as well as adverse effects. Material and methods: The present was a randomized controlled clinical trial. It was divided into two groups: group A was treated with GA applications, four times a day, two atomizer shots, for one month. Group B was treated with LN once a month for three months. Results: 20 pediatric-age patients were studied, out of which 11 were male and 9 female. 12 patients had relatives afflicted with the disease. Most frequent topography was the lower lip. Efficiency of group treated with GA was 63% when compared to group treated with LN which exhibited 81% efficiency. Systemic diverse effects were only observed with use of LN. 0.075 p was obtained (7.5%). Conclusions: GA efficiency was ascertained although it was lesser than that of LN. GA can be used as an adjuvant treatment in order to decrease lesions, it elicits minimal local adverse effects.

Efficiency of cryotherapy in avoiding myringotomy closure in rats.

Myringotomy and ventilation tube placement is an office-based procedure and represents the treatment of choice for middle ear effusions and for transtympanic drug administration. The aim of our study was to study the efficiency of low-pressure spray cryotherapy on protraction of patency time of myringotomies in rats. Bilateral myringotomies were performed with cold instruments. Afterward, one ear in each rat was randomly assigned for liquid nitrogen treatment of the perforated eardrum. Myringotomy patency was recorded daily and histological analysis of the eardrums was performed. In the myringotomy group patency time was 4.75±2.93days, while in the cryotherapy group was 11.01±4.87days. The difference is statistically significant: Student t test, p<0.00001, 95% CI (-8.82; -3.67). Moreover, according to Kaplan-Meier method, myringotomy survival is significantly longer in the cryotherapy group. The healing processes of the cryotreated eardrums last longer and the patency of myringotomy was longer as compared to untreated myringotomies.

An investigation of oxidative stress and antioxidant biomarkers during Greenshell mussel (Perna canaliculus) oocyte cryopreservation

Oxidative damage to proteins and lipids, the enzymatic and nonenzymatic antioxidants' response, and the fertilization and development capability of Perna canaliculus oocytes were investigated at critical treatment steps in a previously published controlled-rate cryopreservation protocol. The cryoprotectant (CPA) from this protocol comprises 10% ethylene glycol (v:v) and 0.2 M trehalose (wt/vol) final concentration. Critical treatment steps included (1) seawater control, (2) CPA addition, (3) CPA addition followed by cooling to −6 °C, (4) CPA addition and cooling to −10 °C, and (5) CPA addition and cooling to −35 °C and immersion in liquid nitrogen (LN). The percentage of fertilized oocytes was 53.8 ± 13.3% in the seawater control but was reduced to 26.0 ± 15.6% after −35 °C + LN treatment, whereas development to D-larvae was 21.0 ± 6.4% in the seawater control reduced to 4.8 ± 2.9% after cooling to −6 °C, and was zero at all the subsequent cooling steps. All oxidative damage biomarkers, protein carbonyls (PCs) and lipid hydroperoxides (LPs), and antioxidants, superoxide dismutase (SOD), catalase, glutathione peroxidase, percent reduced glutathione (%GSH), and total glutathione (defined as glutathione; reduced [GSH] plus glutathione disulphide; derived from two molecules of GSH [GSSG]) were measured over all treatments on unfertilized oocytes over a post-treatment recovery period of 0 to 240 minutes in seawater. An ANOVA showed that both treatment and post-treatment periods had significant effects on the concentrations of all biomarkers (P < 0.05). Protein carbonyls and LPs increased very little after CPA addition and cooling treatments, when compared with the seawater control, but large increases up to sixfold occurred between 0 and 240 minutes for the −35 °C + LN treatment. Concentrations of SOD, catalase, total glutathione, and %GSH at 0 minutes decreased by −31.2%, −26.9%, −21.9%, and −25.0%, respectively, between the seawater control and the −35 °C + LN treatment. In contrast, glutathione peroxidase concentrations at 0 minutes increased by 34.3% between the seawater control and the −35 °C + LN treatment. Although most biochemical biomarkers showed an increasing trend over time (0–240 minutes), total glutathione decreased in concentration over time in all treatments, with the greatest decrease after the −35 °C + LN treatment (−41.2%). Significant correlations between biomarkers and D-larvae yield were negative for LPs and positive for SOD, total glutathione, and %GSH (P < 0.05). This is the first report of an investigation using these oxidative stress biomarkers and antioxidant responses on mussel oocytes, and the results have proved useful in characterizing cellular injury during the cryopreservation process.

Lichen planopilaris after imiquimod 5% cream for multiple BCC in basal cell naevus syndrome

Basal cell naevus syndrome is an inherited autosomal dominant genetic disorder characterised by multiple basal cell carcinomas (BCC), skeletal, neurological and opthalmological abnormalities. The treatment of choice of the often multiple and large BCC consists of a combined approach including surgery, liquid nitrogen and other topical treatment modalities. Imiquimod 5% cream is an immune-response-modifying drug with antiviral and anti-tumour activity. Recent reports have associated the immune-stimulant properties of imiquimod with the exacerbation of several autoimmune skin diseases, such as eczema, psoriasis, vitiligo and lichenoid dermatitis. Here we report a patient with basal cell naevus syndrome who developed a lichen planopilaris on the same site of the scalp, which had been previously treated with two cycles of imiquimod for multiple BCC.