An animal model of transfusion-related acute lung injury and the role of soluble CD40 ligand.

BACKGROUND AND OBJECTIVES: Transfusion-related acute lung injury (TRALI) is a life-threatening complication of transfusion and is one of leading causes of transfusion-associated fatalities. However, the pathogenesis of TRALI is still unclear. Soluble CD40 ligand (sCD40L) is a proinflammatory cytokine that accumulates during blood component storage and is involved in transfusion reactions. The objective of this study was to establish a clinically relevant TRALI animal model and to evaluate the role of sCD40L in TRALI. MATERIALS AND METHODS: Rats' red-blood-cell (RBC) suspensions were prepared, and the quality of RBC was evaluated. A trauma-haemorrhage-transfusion strategy was applied to build the animal model. Lung oedema was evaluated by histopathology examination, total bronchoalveolar lavage fluid (BALF) protein concentration, Evans blue dye (EBD) leakage and inflammatory cytokines. The sCD40L concentrations were measured. RESULTS: Storage lesions of RBCs gradually increased over time. Obvious histological evidence of lung injury of rats transfused with a 35-day RBC was observed. The total BALF protein concentration, EBD leakage, inflammatory cytokines concentration were increased significantly in the Day 35 group. The sCD40L concentration increased significantly in the storage RBC suspension over time but was slightly elevated in rat plasma. CONCLUSIONS: These findings indicated successful establishment of a TRALI animal model with trauma-haemorrhage-transfusion, in which sCD40L may play a minor role in the development of TRALI.

Direct evidence from in situ FTIR spectroscopy that o-quinonemethide is a key intermediate during the pyrolysis of guaiacol.

Although o-quinonemethide (6-methylene-2,4-cyclohexadien-1-one) has been proposed as a key intermediate in char formation during the pyrolysis of guaiacol (2-methoxyphenol), direct evidence of this (e.g., spectroscopic data) has not yet been provided. Using in situ FTIR spectroscopy, the pyrolysis of guaiacol was investigated from 30 °C to 630 °C at 40 °C/min. The IR profiles showed direct evidence of o-quinonemethide production at about 350 °C, which vanished rapidly at around 420 °C in the vapor phase, indicating char formation. In addition, at 400 °C, salicyl aldehyde was observed, which decomposed slowly at about 500 °C. In combination with the known products of guaiacol pyrolysis, these results allowed the major reaction pathways of guaiacol pyrolysis to be discerned. Density functional theory calculations were performed, and the results were found to be in good agreement with the experimentally obtained IR profiles. These findings provide guidance on how to suppress secondary reactions of guaiacol during lignin pyrolysis. Graphical abstract On-line analysis of pyrolysis process of guaiacol using in situ FTIR.

Population-based survey of refractive error among school-aged children in rural northern China: the Heilongjiang eye study.

BACKGROUND: The prevalence of refractive error in the north of China is unknown. The study aimed to estimate the prevalence and associated factors of refractive error in school-aged children in a rural area of northern China. DESIGN: Cross-sectional study. PARTICIPANTS OR SAMPLES: The cluster random sampling method was used to select the sample. METHODS: A total of 1700 subjects of 5 to 18 years of age were examined. All participants underwent ophthalmic evaluation. Refraction was performed under cycloplegia. Association of refractive errors with age, sex, and education was analysed. MAIN OUTCOME MEASURES: The main outcome measure was prevalence rates of refractive error among school-aged children. RESULTS: Of the 1700 responders, 1675 were eligible. The prevalence of uncorrected, presenting, and best-corrected visual acuity of 20/40 or worse in the better eye was 6.3%, 3.0% and 1.2%, respectively. The prevalence of myopia was 5.0% (84/1675, 95% CI, 4.8%-5.4%) and of hyperopia was 1.6% (27/1675, 95% CI, 1.0%-2.2%). Astigmatism was evident in 2.0% of the subjects. Myopia increased with increasing age, whereas hyperopia and astigmatism were associated with younger age. Myopia, hyperopia and astigmatism were more common in females. We also found that prevalence of refractive error were associated with education. Myopia and astigmatism were more common in those with higher degrees of education. CONCLUSION: This report has provided details of the refractive status in a rural school-aged population. Although the prevalence of refractive errors is lower in the population, the unmet need for spectacle correction remains a significant challenge for refractive eye-care services.

Cardioprotective Role of Swertiamarin, a Plant Glycoside Against Experimentally Induced Myocardial Infarction via Antioxidant and Anti-inflammatory Functions.

The study examined the protective effects of swertiamarin on rats with experimentally induced myocardial infarction. Three to six week-old male albino Wistar rats were used in this study and experimental myocardial infarction (MI) was induced using isoproterenol. Our results showed that swertiamarin restored the alteration in heart weight, body weight, and heart weight/tibia length ratio of MI-induced rats to basal levels significantly (p < 0.05). Swertiamarin significantly (p < 0.05) restored the levels of cardiac pathophysiological marker creatine kinase (CKMB), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine transaminase (ALT), and cardiac troponin I (cTn-1) to near normalcy in MI-induced rats. Levels of oxidative stress markers malondialdehyde (MDA), protein carbonyls (PC), and levels of Vitamin C and Vitamin E were significantly (p < 0.05) reverted to near basal levels in MI-induced rats by swertiamarin. Levels of the antioxidant glutathione (GSH) and antioxidant enzymes which include superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione-s-transferase (GST), glutathione reductase (GR), and plasma total antioxidant capacity (TAC) were (p < 0.05) brought to near normalcy in MI-induced rats by swertiamarin. Levels of sodium (Na), potassium (k), and calcium (Ca) ATPases were significantly (p < 0.05) restored to near normalcy in MI-induced rats by swertiamarin. Status of pro-inflammatory cytokines including tumor necrosis factor (TNF-α), interleukin-6 (IL-6), and histological aberrations were also significantly (p < 0.05) restored to near normalcy in MI-induced rats by swertiamarin. Together, our results concluded that swertiamarin exerts significant cardioprotective functions in experimental MI in rats.

Serum PCSK9 is positively correlated with disease activity and Th17 cells, while its short-term decline during treatment reflects desirable outcomes in ankylosing spondylitis patients.

OBJECTIVE: Proprotein convertase subtilisin/kexin type 9 (PCSK9) participates in the autoimmune disease pathology by regulating T helper (Th) cell differentiation, NF-κB pathway, toll-like receptor 4, etc. This study intended to investigate the association of serum PCSK9 with disease activity, Th cells, and treatment response in ankylosing spondylitis (AS) patients. METHODS: Eighty-nine active AS patients were enrolled in this multicenter, prospective study. Serum was collected from AS patients at week (W)0, W4, W8, and W12, as well as from 20 osteoarthritis patients and 20 healthy controls after enrollment to detect PCSK9 by ELISA. Based on the ASAS40 response at W12, AS patients were classified as responders and non-responders. RESULTS: PCSK9 was increased in AS patients versus healthy controls (P < 0.001) and osteoarthritis patients (P = 0.006). In AS patients, PCSK9 was positively linked with C-reactive protein (CRP) (P = 0.003) and ASDAS-CRP (P = 0.017), but not with other clinical properties (P > 0.05). Besides, PCSK9 was negatively correlated with interleukin-4 (P = 0.034), positively associated with Th17 cells (P = 0.005) and interleukin-17A (P = 0.014), but did not relate to Th1 cells, interferon-γ, or Th2 cells (all P > 0.05). Additionally, PCSK9 was decreased from W0 to W12 in general AS patients (P < 0.001) and responders (P < 0.001) but remained unchanged in non-responders (P = 0.129). Moreover, PCSK9 was lower at W4 (P = 0.045), W8 (P = 0.008), and W12 (P = 0.004) in responders versus non-responders. Furthermore, the treatment options did not affect the PCSK9 level (P > 0.05). CONCLUSION: Serum PCSK9 is positively associated with disease activity and Th17 cells, while its short-term decline reflects desirable treatment response in AS patients.

PCSK9 promotes T helper 1 and T helper 17 cell differentiation by activating the nuclear factor-κB pathway in ankylosing spondylitis.

OBJECTIVE: Our previous study reveals that proprotein convertase subtilisin/kexin type 9 (PCSK9) is positively related to inflammatory markers, T helper (Th)-17 cells, and treatment response in ankylosing spondylitis (AS) patients. Subsequently, this study aimed to explore the effect of PCSK9 on Th cell differentiation and its potential molecular mechanism in AS. METHODS: Serum PCSK9 was determined by enzyme-linked immunosorbent assay in 20 AS patients and 20 healthy controls (HCs). Then naïve CD4+ T cells were isolated from AS patients and infected with PCSK9 overexpression or knockdown adenovirus followed by polarization assay. Afterward, PMA (an NF-κB activator) was administrated. RESULTS: PCSK9 was increased in AS patients compared to HCs (p < .001), and it was positively related to Th1 cells (p = .050) and Th17 cells (p = .039) in AS patients. PCSK9 overexpression increased the CD4+ IFN-γ+ cells (p < .05), CD4+ IL-17A+ cells (p < .01), IFN-γ (p < .01), and IL-17A (p < .01), while it exhibited no effect on CD4+ IL-4+ cells or IL-4 (both p > .05); its knockdown displayed the opposite function on them. Moreover, PCSK9 overexpression upregulated the p-NF-κB p65/NF-κB p65 (p < .01), while it had no effect on p-ERK/ERK or p-JNK/JNK (both p > .05); its knockdown decreased p-NF-κB p65/NF-κB p65 (p < .01) and p-JNK/JNK (p < .05). Then, PMA upregulates p-NF-κB p65/NF-κB p65 (p < .001) and increased CD4+ IFN-γ+ cells, CD4+ IL-17A+ cells, IFN-γ, and IL-17A (all p < .01), also it alleviated the effect of PCSK9 knockdown on NF-κB inhibition and Th cell differentiation (all p < .01). CONCLUSION: PCSK9 enhances Th1 and Th17 cell differentiation in an NF-κB-dependent manner in AS, while further validation is necessary.

Effect of deep eutectic solvents-regulated lignin structure on subsequent pyrolysis products selectivity.

The chemical structure of lignin has an important effect on the lignin pyrolysis product distributions. Therefore, it is of great significance to regulate the selectivity of pyrolysis products by modifying the lignin structure. Herein, deep eutectic solvents (DESs) including choline chloride/ethylene glycol (CE), zinc chloride/ethylene glycol (ZE) and choline chloride/acetic acid, treatment of softwood kraft lignin (SKL) is demonstrated. Systematic characterization indicate that the DESs are not only highly conducive to increasing the hydrogen to carbon efficient ratio, reducing the molecular weight and ß-O-4 linkage, but also contributes to the maximum degradation rate and thermal stability of SKL. Noticeably, CE and ZE treatment are significantly improved the amount of H-phenols and C-phenols derived lignin pyrolysis, respectively. In addition, DESs pretreatment are also beneficial to the increment of monomer aromatic hydrocarbons. More importantly, the CE pretreatment contributes to the improvement of bio-oil yield and decrease of char content from lignin pyrolysis.

High-efficiency synthesis of 5-hydroxymethylfurfural and 2,5-diformylfuran from fructose over magnetic separable catalysts.

In this work, a sulfonic acid-functionalized magnetic separable solid acid (Fe3O4@SiO2-SO3H) was synthesized, characterized, and tested for fructose conversion to 5-hydroxymethylfurfural (HMF). Results indicated that the prepared catalyst had a good efficacy for fructose dehydration to HMF due to its larger specific surface area, appropriate acid amount and homogeneous acid distribution. The maximum HMF yield of this work was 96.1 mol%. It was obtained at 120 °C for 1.5 h with 100 mol% fructose conversion. More importantly, the produced HMF could be further in-situ oxidized into 2,5-diformylfuran (DFF) after the replacing of the Fe3O4@SiO2-SO3H with a ZnFeRuO4 catalyst, and the highest DFF yield of 90.2 mol% (based on initial fructose) was obtained after reaction another 8.5 h. The production of DFF from fructose through the above two consecutive steps avoids the intermediate HMF separation, which saves time and energy. In addition, both Fe3O4@SiO2-SO3H and ZnFeRuO4 catalysts exhibited satisfied stability in the recycling experiments, which can be reused at least for five times with the HMF and DFF yield loss<5.3% and 3.3%, respectively. Finally, the plausible reaction mechanisms for fructose conversion to HMF or DFF over Fe3O4@SiO2-SO3H or/and ZnFeRuO4 catalysts were also proposed in this work.

Effect of the interaction of phenolic hydroxyl with the benzene rings on lignin pyrolysis.

In this study, lignin with different phenolic hydroxyl contents and five model compounds are pyrolyzed to investigate the effect of the interaction of phenolic hydroxyl with a benzene ring on lignin pyrolysis. The results demonstrated that phenolic hydroxyl can reduce the stability of lignin and promote the elimination of the side chain on lignin during pyrolysis. The repolymerization during lignin pyrolysis, which results in increased activation energy and char yield during pyrolysis, can be mainly attributed to phenolic hydroxyl. Meanwhile, the repolymerization because of phenolic hydroxyl is obviously affected by the electron cloud density of the benzene ring. The repolymerization caused by the phenolic hydroxyl can be effectively reduced by increasing the electron cloud density. Furthermore, regulation of the product distribution obtained via lignin pyrolysis by changing the electron cloud density of the benzene ring and the phenolic hydroxyl content in lignin is proposed.

Effect of pre-acetylation of hydroxyl functional groups by choline chloride/acetic anhydride on subsequent lignin pyrolysis.

In this work, an emerging and efficient strategy for the preparation of lignin samples with different acetylation degree by choline chloride/acetic anhydride (ChCl/Aa) treatment of pine kraft lignin (PKL) is reported, and the effects of efficient pre-acetylation of hydroxyl functional groups on subsequent lignin pyrolysis are also systematically investigated. The results show that the ChCl/Aa displays a high acetylation efficiency towards the aliphatic hydroxyl (~99.1%) and phenolic hydroxyl (~94.0%) of PKL, which enhances the hydrogen to carbon effective ratio of PKL. Noticeably, the ChCl/Aa has a slight effect on ß-O-4 of PKL, and the acetylation of hydroxyl is beneficial for the improvement of the maximum degradation rate of PKL. In addition, the acetylation of hydroxyl is also significantly contributed to the increment of the pyrolysis bio-oil yield. Importantly, the relative content of the H-phenols obtained from acetylated lignins pyrolysis shows a positive correlation with the acetylation degree of hydroxyl.

Structural characterization and comparison of enzymatic and deep eutectic solvents isolated lignin from various green processes: Toward lignin valorization.

In this work, several representative green processes were developed to extract the enzymatic lignin and deep eutectic solvents (DESs) isolated lignin from corn straw. The results revealed that enzymatic lignin and DESs isolated lignin had a relatively low and homogeneous molecular weight and DESs isolated lignin shown a higher purity. Enzymatic and DESs isolated lignin showed good representativeness and similar to original herbal lignin structures accompany few aryl ether linkage cleavages and oxidation phenomenon. Among them, the subcritical CO2-assisted autohydrolysis and ChCl/Lac DESs treatment exhibited a higher severity for lignin preparation, and sequence DESs isolated lignin had a better reactivity. The ß-O-4 ether bonds and carbon-carbon bonds linkage were further broken up during the Lac and DESs sequence treatment. In short, the described processes showed practical significance for lignin extraction and potential valorization, as well as help to develop more novel strategies for the current biorefinery process.

Synergism of organic acid and deep eutectic solvents pretreatment for the co-production of oligosaccharides and enhancing enzymatic saccharification.

A novel pretreatment using organic acid synergism with deep eutectic solvents (DESs) was developed to the co-production of oligosaccharides, especially for the functional oligosaccharides, and enhancement of corn straws enzymatic saccharification. It was found that lactic acid (Lac) pretreatment combined with choline chloride/Lac system could not only selectively convert the hemicellulose to xylo-oligosaccharides (XOS), which account for 89.7% of total xylose in prehydrolysate (the functional XOS (DP < 5) took up about 35%), but also significantly promote the glucose release (33.2 g/100 g material) and well lignin separation (representing 40.9% in whole process), which better than the single organic pretreatment at higher modified severity index (SI). Structural features of various solids were characterized to better comprehend how hemicellulose and lignin removal influenced enzymatic hydrolysis. This work offered the mild and potential method to co-produce fermentable sugars with the effective separation and valorization of lignin.

Direct conversion of cellulose into sorbitol catalyzed by a bifunctional catalyst.

The conversion of cellulose into sorbitol has been recognized as one of most promising biomass valorization routes for the production of high added-value chemicals. In this work, Sulfonic acid-functionalized carbonized cassava dregs supported ruthenium bifunctional catalyst (Ru/CCD-SO3H) was prepared. The Ru/CCD-SO3H catalyst was employed for direct conversion of cellulose into sorbitol in a neutral aqueous solution. A sorbitol yield of 63.8% could be obtained at 180 °C for 10 h. The acidic sites and metal hydrogenation sites of the Ru/CCD-SO3H catalyst played an important role in hydrolytic hydrogenation of cellulose. Particularly, there presented a strong synergistic effect between sulfonic groups and Ru nanoparticles in conversion of cellulose into sorbitol. This synergistic effect was favorable to enhancing the sorbitol yield. Besides, the stability of Ru/CCD-SO3H catalyst was investigated under the optimal reaction conditions. The catalytic performance of catalyst was decreased after being used five times.

Correlation between pyrolysis behaviors of the components and the overall pyrolysates from pulping spent liquor.

The pyrolysis behaviors of the three major organic components of eucalyptus alkaline peroxide mechanical pulping (APMP) spent liquor including alkali lignin (AL), lignin-carbohydrate complexes (LCC), and polysaccharide (PLS) were studied with emphasis on the effect and contributions of components on the overall pyrolysates of APMP spent liquor solid (ASLS). Profound differences on product properties from each component pyrolysis were illustrated. The results indicated that the bio-oil during ASLS pyrolysis was mainly attributed to AL, while the bio-gas and char were mostly affected by PLS. Small molecule hydrocarbons released during ASLS pyrolysis mainly came from AL and LCC, and CO and CO2 mainly produced from PLS. As for bio-oils, AL generated the largest proportion of monomeric phenols and aromatic hydrocarbons (AHs), while PLS and LCC mainly contributed to the production of ketones, furans and acids. The correlation shown here is of interest for further studies on pulping spent liquor grading utilization.

A promptly approach from monosaccharides of biomass to oligosaccharides via sharp-quenching thermo conversion (SQTC).

In this study, a novel and facile approach of conversion monosaccharides (glucose and xylose) to oligosaccharides (Cello-oligosaccharides and Xylo-oligosaccharides) was demonstrated. The approach did not introduce any chemical reagent and the preparation process could be environmentally friendly. Identification and quantification by ion chromatography (IC) and high performance liquid chromatography (HPLC) showed that the yields of COS and XOS reached to 44.62% (38 s) and 47.09% (30 s) respectively at 500 °C reaction temperature coupled with sharp-quenching method. Structural characterization indicated that such oligosaccharides showed a degree of polymerization (DP) with 2-6, and the units mainly linked by ß-(1 → 4)-glycosidic bond.

[Using HKUST-1 as a Template for Copper Oxides Preparation to Activate Peroxymonosulfate for RhB Degradation].

Using Cu-MOF (HKUST-1) as a template, copper oxide was prepared by heat treatment under different temperatures in this study. The effect of treatment temperature on the composition and morphology of the products were studied. The catalytic abilities of the copper oxide products were tested through a series of experiments of activating peroxymonosulfate (PMS) to degrade RhB under different conditions. The results of XRD, SEM, and XPS show that the product is gradually transformed from a Cu2O/CuO mixture into pure CuO with the increase of heat treatment temperature. The original octahedron structure of HKUST-1 was collapsed. Under neutral pH conditions, when the dosage of PMS and CuO-650 was 1.00 mmol·L-1 and 0.20 g·L-1, respectively, the total degradation of RhB (concentration of 0.10 mmol·L-1) could be completed within 90 min. In addition, CuO-650 also has many advantages, such as it can be applied within a wide range of pH values. In addition, the copper ion dissolution rate was low (1.309 mg·L-1 at pH=3 and 0.987 mg·L-1 at pH=7), and it has great recyclability and stability. These characteristics further proved that CuO-650 can be used as a promising catalyst for PMS activation.

Expression of SIRT1 and P53 in Rat Lens Epithelial cells in Experimentally Induced DM.

PURPOSE: To determine the etiopathogenesis of diabetic cataract by studying changes in relative expressions of silent information regulator protein-1 (SIRT1) and P53 in rat lens epithelial cells (LECs) in experimentally induced diabetes mellitus (DM). METHODS: Six-week-old male SD rats (n = 120) were randomly divided into experimental (n = 80 rats) and control (n = 40 rats) groups. DM was induced in the experimental group (diabetic model) by intraperitoneal (i.p.) injection of 60 mg/kg streptozotocin (STZ). Control group rats were injected similarly with phosphate-buffered saline (PBS). Four and eight weeks after successful induction of DM, relative expressions of SIRT1 and P53 in LECs were analyzed using quantitative real-time (qRT) fluorescence polymerase chain reaction (qRT-PCR) and Western blot analysis. RESULTS: Expression of both SIRT1 and P53 was observed in LECs of control and experimental group rats at 4 and 8 weeks but was significantly greater in experimental compared with control group rats (p < 0.05). CONCLUSIONS: Expression of both SIRT1 and P53 increases in the early stages of diabetic cataract formation, indicating that they play potentially important roles in the pathogenesis of diabetic cataract.

Enhanced enzymatic hydrolysis of eucalyptus by synergy of zinc chloride hydrate pretreatment and bovine serum albumin.

Enhancement of eucalyptus enzymatic saccharification by synergy of ZnCl2 hydrate pretreatment and bovine serum albumin (BSA) was investigated in this study. The result showed that the ZnCl2 hydrate pretreatment could not only selectively extract up to ∼100% of the hemicellulose from eucalyptus, but also convert portion of high crystalline cellulose I into low crystalline cellulose II, which both beneficial for enhancing subsequent pretreated solids enzymatic saccharification. The addition of BSA into enzymatic hydrolysis step could significantly promote the glucose release from pretreated solids, especially, under the low enzyme loading. Furthermore, the material balance indicated that the highest glucose yield of this study was 35.5g/100g raw material, which representing 90.3% of glucose in raw eucalyptus, combined with the xylose yield, 13.9g/100g eucalyptus, it can be concluded that ZnCl2 hydrate pretreatment offered the potential to co-produce xylose and glucose from eucalyptus.

Depolymerization of cellulose into high-value chemicals by using synergy of zinc chloride hydrate and sulfate ion promoted titania catalyst.

Experiments for cellulose depolymerization by synergy of zinc chloride hydrate (ZnCl2·RH2O) and sulfated titania catalyst (SO42-/TiO2) were investigated in this study. The results showed the introduction of sulfate into the TiO2 significantly enhanced the catalyst acid amount, especially for Brønsted acid site, which is beneficial for subsequent cellulose depolymerization. ZnCl2·RH2O hydrate, only a narrow composition range of water, specifically 3.0≤R≤4.0, can dissolve cellulose, which finally resulted the cellulose with low crystallinity and weak intrachain and interchain hydrogen bond network. Coupling of ZnCl2·RH2O hydrate and SO42-/TiO2 catalyst as a mixed reaction system promoted cellulose depolymerization, and the products can be adjusted by the control of reaction conditions, the low temperature (80-100°C) seemed beneficial for glucose formation (maximal yield 50.5%), and the high temperature (120-140°C) favored to produce levulinic acid (maximal yield 43.1%). Besides, the addition of organic co-solvent making HMF as the main product (maximal yield 38.3%).

Evaluation of the effects of isolated lignin on enzymatic hydrolysis of cellulose.

The different physical and chemical properties of lignin might have various effects on the enzymatic hydrolysis of lignocellulosic substrates. In this study, the influence of lignin on enzymatic digestibility of cellulose was assessed. Addition of 20% (4g/L) isolated enzymatic lignin (lignin 2 and 3) and kraft lignin (lignin 4) resulted in 5-20% drop of glucose yield, depending on lignin sources. The inhibitory effect of lignin was abated as the enzyme loading increased from 10 to 20FPU/g dry substrate. However, the increasing lignin amount to 40% (8g/L) did not appear to further decrease the cellulose hydrolysis efficiency. Ethanol lignin (lignin 1) and calcium lignosulfonate (lignin 5) had no negative effect on the enzymatic hydrolysis of cellulose at cellulase loading of 10 or 20FPU/g dry substrate, the increasing lignin content to 40% presented 6.2% increase of glucose yield. The results indicated that different lignin had significantly influence on the enzymatic hydrolysis, which was confirmed by analysis in chemical composition, elemental analysis, functionality, and thermogravimetry.