Recombinant PBP2a of methicillin-resistant S. aureus formulation in Alum and Montanide ISA266 adjuvants induced cellular and humoral immune responses with protection in Balb/C mice.

Staphylococcus aureus is an important cause of both hospital and community acquired infections worldwide. S.aureus can develop multidrug resistance; thus, immunotherapy can be a rational alternative. High level ß-lactam resistance of S. aureus has been attributed to the penicillin binding protein 2a (PBP2a). In this study, we assessed the immunogenicity and protectivity of PBP2a formulated in Montanide ISA266 and Alum adjuvants. Recombinant PBP2a with a molecular weight of approximately 13 kDa was expressed and purified by nickel-nitrilotriacetic acid (NI-NTA) affinity chromatography and characterized by SDS-PAGE and Western blot. To investigate the immunogenicity and protective effects of recombinant protein, 20 µg of r-PBP2a in various formulations were subcutaneously injected in different groups. Two booster vaccinations were carried out in two-week intervals and blood samples were collected two weeks after each injection. To determine the type of induced immune response, sera and splenocytes were analyzed by ELISA for total IgG and isotypes (IgG1 and IgG2a) and cytokine secretion (IFN-γ, IL-4, IL-17 and TNF-α), respectively. Three weeks following the last immunization, experimental mice were challenged with 5 × 108 CFU of bacteria intraperitoneally and mortality rate and bacterial load were assessed. Interestingly, analysis of humoral immune responses revealed that administration of r-PBP2a with Montanide ISA266 significantly increased specific IgG responses and also IgG1 isotype compared to alum-adjuvanted vaccine group. Also, r-PBP2a formulation with alum and MontanideISA266 adjuvants raised IFN-γ, IL-4, IL-17 cytokines secretion, and protectivity following experimental challenge. The results of the present study provide evidences for immunogenicity and protectivity of PBP2a protein as a vaccine candidate.

A comparison of alum sludge with peat for aqueous glyphosate removal for maximizing their value for practical use.

This study compares and contrasts the glyphosate removal efficiency of alum sludge (waterworks residue) and Irish peat in aqueous solution. Organic phosphonate of glyphosate aqueous solution was removed in pot tests separately filled with peat and alum sludge, while effluent samples were taken from each pot to analyse the concentration of phosphorus (P) and COD (chemical oxygen demand); physical and chemical analysis for both media before and after use was carried out subsequently. The results show that the P removal capacity of alum sludge was significant (>99%), while the removal capacity of peat was considerably less than 10% after 10 weeks. Both materials significantly reduced the levels of COD, but it was noted that peat had a marginally greater initial P removal capacity (68 ± 22%) and did perform better than alum sludge (57 ± 12%). Moreover, pre-treatment is a crucial step to harness the full potential of peat. Overall, this study provides a scientific clue for sorbents selection when considering alum sludge and peat to maximize their value in practice.

Determination of quality markers of Xuezhiling tablet for hyperlipidemia treatment.

BACKGROUND: The massive number of ingredients in traditional Chinese medicines (TCMs) makes quality control very difficult. The concept of quality markers (Q-marker) was recently proposed to evaluate the quality of TCMs. Xuezhiling tablets (XZL) are widely used for the treatment of hyperlipidemia in China owing to its noticeable effectiveness and mild adverse effects, but there are no proper Q-markers for this Chinese patent medicine. PURPOSE: The aim of the present study was to determine the Q-markers of XZL against hyperlipidemia through an integration of investigations on its lipid-lowering effect, metabolomics, content determination and pharmacokinetics. METHODS: XZL was prepared in accordance with the method described in the Chinese pharmacopoeia (Ch.P.). Hyperlipidemia was induced in rats through the administration of a high-fat diet (HFD). The hypolipidemic effect of XZL was investigated through the detection of the blood levels of total glyceride (TG), total cholesterol (TC), and low density lipoprotein cholesterol (LDL-C). A metabolomics study was conducted to analyze the overall effects of XZL on the regulation of lipid metabolism. The main bioactive compounds of XZL were identified and determined in the XZL preparation and the medicated plasma of hyperlipidemic rats. RESULTS: XZL lowered the levels of TG, TC, and LDL-C through alterations of metabolic patterns. 2,3,5,4'-Tetrahydroxystilbene-2-O-ß-D-glucopyranoside (THSG), chrysophanol-1-O-ß-glucopyranosyl-(1→3)-O-ß-D-glucopyranosy1-(1→6)-O-ß-D-glucopyranosyl-(1→6)-O-ß-D-glucopyranoside (SHJ), cassiaside, rubrofusarin gentiobioside, aurantio-obtusin, chryso-obtusin, and obtusinfolin were identified and determined both in the preparation and the blood of hyperlipidemic rats. CONCLUSION: SHJ, obtusinfolin, THSG, rubrofusarin gentiobioside, and aurantio-obtusin, which are more abundant in the preparation, leading to greater exposure in vivo, were suitable Q-markers to guarantee the medicinal quality of XZL and ensure the clinical effectiveness on hyperlipidemia.

Volumetric modeling of two sludge piles from water treatment plants in a Brazilian reservoir.

Water treatment plants are designed to continuously produce drinkable water, meeting defined criteria of potability. However, besides potable water, these plants produce sludges that are disposed of in the environment. The present work aimed to evaluate the sludges generated in two water treatment plants and disposed of in the margin of the Juturnaíba dam. Since alum has been used as a flocculating agent in these two plants, the concentrations of aluminum were measured in the sludges and in surface sediments. The generated piles are extremely soft to walk on and difficult to measure, so indirect modeling procedures had to be applied. The calculated mass of the sludge piles at each plant are similar and respectively 60,370 and 61,479 tons. The aluminum content of the residues, calculated according to its dosage, was 33.2 and 32.6 g kg-1 in the piles from the two plants. The amount of alum dosed to the water corresponds almost to the excess of aluminum in the sludge, compared to the sediments. It was concluded that regardless of the fact that residues are disposed of in very restricted areas, they are directly in contact with the water and may constitute a threat for the environment and humans' health.

Characterization of water treatment sludge and its reuse as coagulant.

Coagulation-flocculation process results in the generation of large volume of waste or residue, known as water treatment sludge (WTS), in the purification of surface water for potable supplies. Sustainable management of the inevitable waste requires careful attention from the plant operators and sludge managers. In this study, WTS produced with the optimum alum dose of 30 ml/L at the laboratory scale has been treated with sulphuric acid to bring forth a product known as sludge reagent product (SRP). The performance of SRP is evaluated for its efficiency in removing the colloidal suspensions from the Yamuna river water over wide pH range of 2-13. 1% sludge acidified with sulphuric acid of normality 2.5 at the rate of 0.05 ml/ml sludge has been observed as the optimum condition for preparing SRP from WTS. The percentage turbidity removal is greater at higher pH value and increases with increasing the dosage of SRP. The optimum SRP dosage of 8 ml/L in the pH range of 6-8 performed well in removing the colloidal suspension and other impurities from the Yamuna water. The quality of treated water met the prescribed standards for most of the quality parameters. Thus, SRP has the potential to substitute the conventional coagulants partially or completely in the water treatment process, depending on the quality needed at the users end.

A parametric study of alum recovery from water treatment sludge.

Alum recovery from water treatment sludge is a promising technique applied to decrease usage of fresh coagulants in the water treatment industry. In addition, alum recovery reduces sludge volume for easy handling. The undertaken work investigated the parametric conditions for alum recovery procedure by acidification. The results show that alum recovery reaches up to 69.03%, and the reduction of sludge volume reaches its highest level at 90%. Moreover, results of the parametric investigation reveal that the mixing time of 60 minutes and mixing intensity of 150 rpm are the optimum conditions of mixing for alum recovery from water treatment sludge. The optimum pH level is 1.50 for alum recovery as indicated by maximum aluminum releasing, maximum reduction of sludge volume, and reasonable dosages of added sulfuric acid.

Stormwater runoff to an impaired lake: impacts and solutions.

Stormwater runoff can represent a significant source of pollutants to downstream water bodies. An integrated assessment was conducted for the Spring Lake watershed in western Michigan because of concerns that increased impervious land cover in the watershed, especially in sub-basins adjacent to Spring Lake, would result in greater stormwater runoff and pollutant loads. Spring Lake has a history of high total phosphorus (TP) concentrations and cyanobacterial blooms. An alum treatment, paid for by an assessment, was applied to Spring Lake in 2005 to control internal phosphorus loading; hence, there was an economic incentive for stakeholders to limit new phosphorus from entering the lake, which otherwise would reduce the long-term efficacy of the alum treatment. This study provides a novel six-step process that identifies priority areas and optimally reduces nonpoint sources of pollution. We identified a suite of best management practices to be placed in the watershed, assessed their optimal locations based on pollutant sources, and modeled the degree to which their implementation would reduce TP and total suspended solids. Application of the modeled best management practices (BMPs) resulted in a 15 % reduction in TP load and a 17 % reduction in total suspended solid load. Reductions were not uniform throughout the watershed, with the greatest reductions closest to Spring Lake. We also developed a flow chart for BMP selection, which may be transferable to other watersheds with similar issues.

A review on alum sludge reuse with special reference to agricultural applications and future challenges.

Alum salts are commonly used in the water industry to promote coagulation in the production of clean drinking water, which results in the generation and accumulation of 'waste' by-product 'alum sludge' in large volumes. Effective and efficient management of alum sludge in an economically and environmentally sustainable manner remains a significant social and environmental concern with ever increasing demand for potable water as a result of rapidly escalating world population and urban expansion. Various intensive practices have been employed to reuse the alum sludge in an attempt to figure out how to fill the gap between successful drinking water treatment process and environmentally friendly alum sludge management for over the years. This paper primarily aimed at comprehensive review of the existing literature on alum sludge characteristics, its environmental concerns and their potential utilization, especially in agricultural and horticultural sectors leading to update our recent state of knowledge and formulate a compendium of present and past developments. Different types of alum sludge utilizations in various fields were recognized and examined. The strengths, weaknesses, opportunities and potential risks of alum sludge reuse options with particular reference to agriculture were highlighted and knowledge gaps were identified. Research priorities and future challenges that will support in the development of effective alumsludgemanagement practices in agriculture with multi-prong strategies were discussed.

Sorption and desorption of diverse contaminants of varying polarity in wastewater sludge with and without alum.

Sewage sludge sorption and desorption measurements were conducted for nine diverse contaminants of varying polarity: caffeine, sulfamethoxazole, carbamazepine, atrazine, estradiol, ethinylestradiol, diclofenac, and, for the first time desethylatrazine and norethindrone. Two types of sorption behaviour were observed. Compounds with a log octanol-water partition coefficient, log Kow, below 3 showed little or no sorption over 48 hours of shaking, while compounds with log Kow over 3 showed 30 to 90% sorption within the first few minutes. After 6 hours of shaking, mass loss through suspected biotransformation became evident for some compounds. At the pH range considered (5.7-6.7), diclofenac (pKa 4.0, log Kow 4.5) was the only compound in which pH dependent sorption could be quantified. The log sewage sludge-water distribution coefficients, log Kd, ranged from 0.2 to 2.9, and, as expected, increased with increasing log Kow of the compound and organic carbon (OC) content of the sewage sludge. A sewage sludge precipitated with alum had a substantially lower Kd values, as well as lower OC content, compared to alum-free sludge. Desorption was studied by sequentially replacing supernatant water. With each water replacement, log Kd values tended to either remain similar (following a linear isotherm) or in some cases increase (following a Freundlich-type isotherm). The length of time required to restore equilibrium increased with each rinsing step. A literature review of reported Kd values compared well with the alum-free sludge data, but not the alum-sludge data. Sewage sludge Kd across the literature appear more consistent with increasing Kow.

Effect of water treatment additives on lime softening residual trace chemical composition--implications for disposal and reuse.

Drinking water treatment residues (WTR) offer potential benefits when recycled through land application. The current guidance in Florida, US allows for unrestricted land application of lime softening WTR; alum and ferric WTR require additional evaluation of total and leachable concentrations of select trace metals prior to land application. In some cases a mixed WTR is produced when lime softening is accompanied by the addition of a coagulant or other treatment chemical; applicability of the current guidance is unclear. The objective of this research was to characterize the total and leachable chemical content of WTR from Florida facilities that utilize multiple treatment chemicals. Lime and mixed lime WTR samples were collected from 18 water treatment facilities in Florida. Total and leachable concentrations of the WTR were measured. To assess the potential for disposal of mixed WTR as clean fill below the water table, leaching tests were conducted at multiple liquid to solid ratios and under reducing conditions. The results were compared to risk-based soil and groundwater contamination thresholds. Total metal concentrations of WTR were found to be below Florida soil contaminant thresholds with Fe found in the highest abundance at a concentration of 3600 mg/kg-dry. Aluminum was the only element that exceeded the Florida groundwater contaminant thresholds using SPLP (95% UCL = 0.23 mg/L; risk threshold = 0.2 mg/L). Tests under reducing conditions showed elevated concentrations of Fe and Mn, ranging from 1 to 3 orders of magnitude higher than SPLP leachates. Mixed lime WTR concentrations (total and leachable) were lower than the ferric and alum WTR concentrations, supporting that mixed WTR are appropriately represented as lime WTR. Testing of WTR under reducing conditions demonstrated the potential for release of certain trace metals (Fe, Al, Mn) above applicable regulatory thresholds; additional evaluation is needed to assess management options where reducing conditions may develop.

Neutralisation of an acidic pit lake by alkaline waste products.

A former open pit where black shale (alum shale) was excavated during 1942-1965 has been water filled since 1966. The water chemistry was dominated by calcium and sulphate and had a pH of 3.2-3.4 until 1997-1998, when pH was gradually increasing. This was due to the intrusion of leachates from alkaline cement waste deposited close to the lake. A stable pH of around 7.5 was obtained after 6-7 years. The chemistry of the pit lake has changed due to the neutralisation. Concentrations of some dissolved metals, notably zinc and nickel, have gone down, as a result of adsorption/co-precipitation on solid phases (most likely iron and aluminium hydroxides), while other metals, notably uranium and molybdenum, are present at elevated levels. Uranium concentration is reaching a minimum of around pH 6.5 and is increasing at higher pH, which may indicate a formation of neutral and anionic uranyl carbonate species at high pH (and total carbonate levels around 1 mM). Weathering of the water-exposed shale is still in progress.

Aluminum sulfate (alum) application interactions with coupled metal and nutrient cycling in a hypereutrophic lake ecosystem.

Many lake ecosystems worldwide experience severe eutrophication and associated harmful blooms of cyanobacteria due to high loadings of phosphorus (P). While aluminum sulfate (alum) has been used for decades as chemical treatment of eutrophic waters, the ecological effects of alum on coupled metal and nutrient cycling are not well known. The objective of our study was to investigate the effects of an in-situ alum treatment on aluminum and nutrient (P, N, and S) cycling in a hypereutrophic lake ecosystem. Our results indicate that the addition of alum along with sodium aluminate (as a buffer) increased dissolved aluminum and sulfate in the surface and pore waters, and altered nitrogen cycling by increasing nitrous oxide (N2O) concentrations in the surface water. The increase of aluminum and sulfate may potentially feedback to alter benthic community dynamics. These results enhance our understanding of the unintended ecological consequences of alum treatments in hypereutrophic freshwater ecosystems.

Regular consumption of a silicic acid-rich water prevents aluminium-induced alterations of nitrergic neurons in mouse brain: histochemical and immunohistochemical studies.

Silicon is not generally considered an essential nutrient for mammals and, to date, whether it has a biological role or beneficial effects in humans is not known. The results of a number of studies suggest that dietary silicon supplementation might have a protective effect both for limiting aluminium absorption across the gut and for the removal of systemic aluminium via the urine, hence, preventing potential accumulation of aluminium in the brain. Since our previous studies demonstrated that aluminium exposure reduces the number of nitrergic neurons, the aim of the present study was to compare the distribution and the morphology of NO-containing neurons in brain cortex of mice exposed to aluminium sulphate dissolved in silicic acid-rich or poor drinking water to assess the potential protective role of silicon against aluminium toxicity in the brain. NADPH-d histochemistry and nNOS immunohistochemistry showed that high concentrations of silicon in drinking water were able to minimize the impairment of the function of nitrergic neurons induced by aluminium administration. We found that silicon protected against aluminium-induced damage to the nitrergic system: in particular, we demonstrated that silicon maintains the number of nitrergic neurons and their expression of nitrergic enzymes at physiological levels, even after a 12 and 15 month exposure to aluminium.

Development of alum sludge-based constructed wetland: an innovative and cost effective system for wastewater treatment.

This article describes a research attempt to integrate the dewatered alum sludge, a residual by-product of drinking water treatment process, into a constructed wetland (CW) system for the purpose of enhancing the wastewater treatment performance, thus developing a so called alum sludge-based constructed wetland system. A multi-dimensional research project including the batch tests of phosphorus (P) adsorption onto alum sludge followed by the model CWs trials of single and multi-stage CWs, has been conducted since 2004. It has been successfully demonstrated that the alum sludge-based CW is capable of enhanced and simultaneous removal of P and organic matter (in terms of BOD5 and COD), particularly from medium and high strength wastewater. The sludge cakes act as the carrier for developing biofilm for organics removal and also serve as adsorbent to enhance P immobilization. Batch P-adsorption tests revealed that the alum sludge tested possesses excellent P-adsorption ability of 14.3 mg-P/g x sludge (in dry solids) at pH 7.0 with the adsorption favored at lower pH. The results obtained in a 4-stage treatment wetland system suggest that high removal efficiencies of 90.4% for COD, 88.0% for BOD5, 90.6% for SS, 76.5% for TN and 91.9% for PO4(3-)-P under hydraulic loading of 0.36 m3/m2 x d can be achieved. The field demonstration study of this pioneering development is now underway.

Phosphorus removal in laboratory-scale unvegetated vertical subsurface flow constructed wetland systems using alum sludge as main substrate.

This research has two eventual goals: (1) To optimize performance of subsurface constructed wetlands for removal of phosphorus (P) (2) To demonstrate that dewatered alum sludge (a by-product), can be reused as a constructed wetland substrate. To achieve these, alum sludge from a water treatment plant was characterized and used as main substrate in four experimental vertical sub-surface flow constructed wetland systems treating dairy farm wastewater. Results show that the alum sludge has suitable hydraulic characteristics (uniformity coefficient = 3.6) for use as a substrate, and in the batch studies, up to 48.6 mg-P was removed by 1 g of the alum sludge at a P concentration of 360 mg-P/l and a dosage of 5 g/l. Results from the experimental systems highlight the significant P removal ability of the alum sludge. However, the inclusion of pea gravel at the infiltrative surface of some of the systems had a negative effect on the P removal performance. Sequential P-fractionation results show that there was no significant increase in the easily extractable P, but for total P, there was significant increase, although this was found to decrease with depth. This study shows that the novel use of dewatered alum sludge can bring about high P removal in vertical subsurface flow constructed wetland systems.

Characterization of hydration products of mineral trioxide aggregate.

OBJECTIVE: To characterize the hydration products of white mineral trioxide aggregate (MTA). METHODOLOGY: Mineral trioxide aggregate, white Portland cement and bismuth oxide were evaluated using X-ray diffraction (XRD) analysis and Rietveld XRD. The cements were tested un-hydrated and after hydration and curing for 30 days at 37 degrees C. Analysis of hydrated cement leachate was performed weekly for five consecutive weeks from mixing using inductively coupled plasma atomic emission spectroscopy after which the cements were viewed under the scanning electron microscope to evaluate the cement microstructure. Quantitative energy dispersive analysis with X-ray was performed and atomic ratios were plotted. RESULTS: Both Portland cement and MTA produced calcium silicate hydrate (C-S-H) and calcium hydroxide (CH) on hydration. The tricalcium aluminate levels were low for MTA which resulted in reduced production of ettringite and monosulphate. On hydration the bismuth level in the hydrated MTA decreased; bismuth oxide replaced the silica in the C-S-H and was leached out once the C-S-H decomposed with time. Both MTA and Portland cement released a high amount of calcium ions which decreased in amount over the 5-week period. CONCLUSIONS: The hydration mechanism of MTA is different to that of Portland cement. In MTA the bismuth oxide is bound to the C-S-H and is leached out from the cement with time as the C-S-H decomposes. MTA produces a high proportion of calcium ions from CH a by-product of hydration and also by decomposition of C-S-H. The release of calcium ions reduces with time.

Behavior of aluminum in aluminum welders and manufacturers of aluminum sulfate--impact on biological monitoring.

OBJECTIVES: The suitability of determining aluminum in serum or urine as a form of biological monitoring was critically assessed. METHODS: Airborne and internal aluminum exposure was assessed for 12 aluminum welders in a shipyard and 5 manufacturers of aluminum sulfate. Particles were characterized with X-ray diffraction and scanning electron microscopy. Aluminum in air and biological samples was analyzed using electrothermal atomic absorption spectrometry. Basic toxicokinetic features were inferred from the data. RESULTS: The mean 8-hour time-weighted average concentration of aluminum was 1.1 (range 0.008-6.1) mg/m(3) for the shipyard and 0.13 (range 0.02-0.5) mg/m(3) for the aluminum sulfate plant. Welding fume contained aluminum oxide particles <0.1 microm in diameter and their agglomerates, whereas bauxite and aluminum sulfate particles ranged from 1 to 10 microm in diameter. The shipyard welders' mean postshift serum and urinary concentrations of aluminum (S-Al and U-Al, respectively) were 0.22 and 3.4 micromol/l, respectively, and the aluminum sulfate workers' corresponding values were 0.13 and 0.58 micromol/l. Between two shifts, the welders' S-Al concentration decreased by about 50% (P<0.01), but their U-Al concentration did not change (P=0.64). No corresponding temporal changes occurred among the aluminum sulfate workers. After aluminum welding at the shipyard had ceased, the median S-Al concentration decreased by about 50% (P=0.007) within a year, but there was no change (P=0.75) in the corresponding U-Al concentration. CONCLUSIONS: About 1% of aluminum in welding fume appears to be rapidly absorbed from the lungs, whereas an undetermined fraction is retained and forms a lung burden. A higher fractional absorption of aluminum seems possible for aluminum sulfate workers without evidence of a lung burden. After rapid absorption, aluminum is slowly mobilized from the lung burden and dominates the S-Al and U-Al concentrations of aluminum welders. For kinetic reasons, S-Al or U-Al concentrations cannot be used to estimate the accumulation of aluminum in the target organs of toxicity. However, using U-Al analysis to monitor aluminum welders' lung burden seems practical.

Soil biogeochemical characteristics influenced by alum application in a municipal wastewater treatment wetland.

Constructed treatment wetlands are a relatively low-cost alternative used for tertiary treatment of wastewater. Phosphorus (P) removal capacity of these wetlands may decline, however, as P is released from the accrued organic soils. Little research has been done on methods to restore the treatment capacity of aging constructed wetlands. One possibility is the seasonal addition of alum during periods of low productivity and nutrient removal. Our 3-mo mesocosm study investigated the effectiveness of alum in immobilizing P during periods of reduced treatment efficiency, as well as the effects on soil biogeochemistry. Eighteen mesocosms were established, triplicate experimental and control units for Typha sp., Schoenoplectus californicus, and submerged aquatic vegetation (SAV) (Najas guadalupensis dominated). Alum was slowly dripped to the water column of the experimental units at a rate of 0.91 g Al m(-2) d(-1) and water quality parameters were monitored. Soil cores were collected at experiment initiation and completion and sectioned into 0- to 5- and 5- to 10-cm intervals for characterization. The alum floc remained in the 0- to 5-cm surface soil, however, soil pH and microbial parameters were impacted throughout the upper 10 cm with the lowest pH found in the Typha treatment. Plant type did not impact most biogeochemical parameters; however, data were more variable in the SAV mesocosms. Amorphous Al was greater in the surface soil of alum-treated mesocosms, inversely correlated with soil pH and microbial biomass P in both soil layers. Microbial activity was also suppressed in the surface soil of alum-treated mesocosms. This research suggests alum may significantly affect the biogeochemistry of treatment wetlands and needs further investigation.

A study of dual polymer conditioning of aluminum-based drinking water treatment residual.

Conditioning of an aluminum-based water treatment sludge by single and dual polymers was investigated in this study. Capillary suction time (CST), specific resistance to filtration (SRF) and settling rate were used to evaluate the sludge dewatering characteristics. Sludge dewaterability resulting from single and dual polymer conditioning were compared for the purpose of exploring the validity and effectiveness of dual polymer conditioning strategy. Unlike activated sludge derived from wastewater treatment, results from this study have demonstrated that conditioning of the alum sludge by the combined use of an cationic polymer (FO-4140) followed by an anionic polymer (LT-25) does not exhibit considerable advantage in further improvement of sludge dewaterability with comparison of single polymer conditioning. This study supports the view that for aluminum-based water treatment sludge, inter particle bridging seems to be the dominative mechanism and the charge neutralization plays a less important role in the conditioning process. In addition, an intrinsic relationship between CST and SRF was deducted and tested via the experimental data obtained from the study.