Wax Crystals Research Articles

Effects of spherical, disk-like and lamellar nanocomposite particles on cold flow behaviors of waxy crude oil

Wax precipitation and deposition are challenging hurdles in pipeline transportation of waxy crude oil. This study focuses on transforming SiO2, Laponite, and montmorillonite particles into hydrophobic particles through polymer grafting. Chemical structure analysis and grafting ratio calculation were performed. Through thermogravimetric analyzer (TGA) test, Laponite showed the highest grafting ratio. The effects of adding nanocomposites, TDA-2024-22, and commercial pour point depressants on the rheological behavior and wax crystallization behavior of African crude oil were compared. Addition of the nanocomposite PDA-22@SiO2 to crude oil, reduced the viscosity by 85.6% at 20 °C and the thixotropy by 60.2%. Wax crystal size decreased in the order of PDA-22@SiO2, PDA-22@Laponite, KS-10, TDA2024-22, and PDA-22@MMT. The pour point decreased by 6 °C with PDA-22@ SiO2 particles but only 3 °C with Laponite and montmorillonite composites. Spherical nanocomposites are promising flow improvers for waxy crude oil.

Optimizing the low-temperature performance of high-wax-content crude oil with emulsion-based wax inhibitors: Mechanisms and efficacy.

Wax deposition in constant-temperature transportation of waxy oil with high pour points poses a significant challenge for the oil industry. The demand for efficient methods to solve wax deposition has gained attention. To elucidate the impact of emulsion-based wax inhibitors on the performance of crude oils with varying wax content at low temperatures, experiments, rheological analyses, and microscopic analyses were conducted to study their pour point regulation, low-temperature flow improvement, wax prevention effectiveness, and wax crystallization behavior. The results indicate that emulsion-based wax inhibitors significantly reduce the pour point of crude oil, especially for high-wax-content oil, while for low-wax-content oil, the pour point only decreased by 3°C. By penetrating and dispersing wax crystals, the inhibitor reduced the viscosity of crude oil at low temperatures, enhancing its flowability. At 28°C, crude oil transitioned from a shear-thinning non-Newtonian fluid to a Newtonian fluid at wax inhibitor dosages of 500-750ppm. At 1000ppm, the wax prevention rate for high-wax-content oil reached 87.5%, but the effect plateaued beyond this concentration. Additionally, the wax prevention and removal mechanism of the emulsion-based wax inhibitors is discussed. This study confirms that emulsion-based wax inhibitors significantly enhance the low-temperature processability of crude oil, offering a viable strategy for the conveyance and refinement in cold climates.

Application and evaluation of nanohybrid polymer as flow improver for Indian crude oil

Paraffin deposition remains a persistent challenge in the petroleum industry, particularly exacerbated in cold environments, necessitating effective mitigation strategies. Among these, pour point depressants (PPDs) stand out as a superior method. Nanomaterials have advantage of their surface properties due to which they are extensively used in petroleum industry. The present study is aimed at synthesizing a nanohybrid polymer tailored for use as a flow improver for Indian crude oil. The synthesized product was characterized by different analytical techniques and evaluated by pour point studies, rheological studies, Cold finger test, microscopic studies and aging studies. Results underscored the substantial potential of the PPD in reducing both pour point and viscosity of the crude oil. Significantly, the most notable achievement was observed at a concentration of 3000 ppm, yielding a remarkable pour point reduction of 18 °C alongside a substantial 95% decrease in viscosity. Microscopic analyses unveiled the presence of dispersed wax crystals after treatment with the nanohybrid polymer, indicative of its interaction with paraffin crystals, thereby impeding their formation or growth. Furthermore, aging studies elucidated the stability and sustained efficacy of nanohybrid polymer over time, crucial considerations for its practical applicability within the Petroleum industry. Overall, this investigation underscores the promising role of nanohybrid polymers as effective pour point depressants, offering tangible solutions to the persistent challenge of paraffin deposition while enhancing operational efficiencies and cost-effectiveness within the industry.

Characterization of morphological and chemical changes using atomic force microscopy and metabolism assays: the relationship between surface wax and skin greasiness in apple fruit.

Skin greasiness occurred on stored apples (Malus domestica Borkh.) is generally believed to result from changes in surface wax components. Previous reports have typically correlated wax changes with greasiness scores to reveal the contributing wax components. A notable limitation of this approach is that greasiness scores are highly subjective and influenced by individual perception. This study aimed to assess skin quality by quantitatively analyzing wax morphology changes in greasy 'Jonagold' apples using Atomic Force Microscopy (AFM) roughness parameters Ra, Rq, Rmax, and Rz, and to correlate these changes with wax composition. AFM results revealed that wax crystals disappeared as skin greasiness increased, accompanied by significant declines in roughness parameters Ra, Rq, Rmax, and Rz, which decreased by 70% to 85%. Chemical analysis showed a significant increase in liquid esters, including linoleate and oleate esters, in the surface waxes, which negatively correlated with the decline in roughness parameters. Key genes related to ester production, such as MdFAD2, MdWSD1, and MdWBC11, exhibited increased expression and were also negatively correlated with decreases in Rq, Ra and Rz. Additionally, 1-Methylcyclopropene (1-MCP) treatment suppressed both the development of greasiness and the associated changes. Our findings suggest that the increased liquid esters contribute to alternations in wax morphology in greasy apples, and that MdFAD2, MdWSD1, and MdWBC11, play crucial roles in ester biosynthesis. These results highlight the effectiveness of AFM roughness parameters Ra, Rq, Rmax, and Rz in quantifying wax morphology changes in apples during skin greasiness development.

Wax crystal dynamics and viscosity abnormal reduction in asphaltene-containing waxy oil at low temperatures

A micro-rheological integration experiment was conducted on model oils without asphaltene and with 0.05 wt% asphaltene. The study examined the viscosity-temperature characteristics of both model oils under different cooling rates and shear rates. Significant changes were observed in the viscosity-temperature curve of the asphaltene-containing model oil. At low shear rates, a distinct wavelet peak was identified in the temperature range of 37-30 °C, where viscosity initially increased, then decreased, and subsequently increased again. This wavelet peak diminished with the increasing shear rate, eventually forming a “buffer platform”. The underlying mechanism was attributed to asphaltene altering the wax crystal behavior from “uniform precipitation network structure formation” to “formation of unstable large aggregates - dissociation into multiple small stable aggregates - growth into larger stable aggregates.” In addition, the wax precipitation rate remained constant in the model oil without asphaltene, while in the asphaltene-containing model oil, the rate significantly accelerated between 34.5 °C and 32.5 °C. And the asphaltenes promoted the formation of small wax crystal particles but inhibited the growth of wax crystal particles.

Study on the formation process of methane hydrates on the surface of wax crystals with pour point depressant

Molecular dynamics simulations were applied to clarify the kinetic pathways and mechanism in the process of methane hydrate formation on the wax crystal surface with ethylene–vinyl acetate copolymer (EVA). The simulation results showed that methane molecules near the surfaces would adsorb on the surfaces of wax-EVA eutectic, while hydrates formed in the bulk of the solution away from the surfaces. The VA side chains on the wax crystal surface inhibit/promote the growth of hydrates by influencing the growth and re-dissolution process of methane nano-bubbles on eutectic surface. When a small amount of VA side chains was introduced on the wax crystal surface, as the simulation proceeded, the generated hydrates would gradually decompose. However, when more VA side chains distribute on the surface of wax crystals, the growth rate of hydrates would increase. In addition, hydrates and wax crystals could connect through a semi-cage structure, in which the VA side chain served as the guest of this semi-cage, making the eutectic and hydrates form a whole and increasing the risk of co-deposition of wax crystals and hydrates.

Optimizing cold-flow properties and oxidation stability of B40 biodiesel blend with turpentine oil and ethanol: Experimental and quantum chemical approach

This study investigates the effectiveness of turpentine oil and ethanol as cold-flow improvers (CFIs), synergizing as antioxidants for ternary B40 biodiesel blend. Utilizing the response surface methodology (RSM), the effects of these CFIs on the ternary blend, including cloud point (CP), cold filter plugging point (CFPP), filter blocking tendency (FBT), precipitation, and oxidation stability, were comprehensively evaluated. The analysis of variance (ANOVA) presented that a linear model best describes the impact of CFIs on CP, whereas quadratic models more accurately represent CFPP, FBT, and oxidation stability. The results also demonstrate that 2FI was identified as the most suitable model for describing precipitation responses. The optimal concentrations of turpentine oil and ethanol were determined to be 10 % v/v and 0.5 % v/v, respectively, resulting in predicted values for CP, CFPP, FBT, precipitation, and oxidation stability of 10.4 °C, 9.0 °C, 1.31, 9.09 mg/100 mL, and 225 minutes, respectively. Theoretical insights from density functional theory (DFT) calculations confirmed that the primary interaction controlling wax formation in biodiesel blends is the strong hydrogen bonding between monostearin and pinene, a major component of turpentine oil, which effectively disrupts wax crystallization. This study presents a novel approach to enhancing B40 biodiesel cold-flow properties and maintaining its oxidation stability by utilizing the synergistic effects of turpentine oil and ethanol to optimize B40 biodiesel formulation.

Mechanism study on rheological response of thermally pretreated waxy crude oil

Pre-heating treatment significantly influences the low-temperature flow characteristics of waxy crude oil, yet its underlying mechanism warrants further investigation. This study explores the effects of pre-heating temperatures (60–110 °C) on the rheological characteristics of waxy crude oil from Nanyang using 1H NMR, DSC, Microscopic Observation, and Rheological Property Test. Findings reveal that asphaltenes increasingly dissolve at the pre-heating temperature of 70 °C, improving their capacity to act as nucleation sites for wax crystals. Consequently, the wax appearance temperature (WAT) increases. Meanwhile, the rheological parameters of the crude oil decrease and wax crystals exhibit smaller and tighter rod-like structures. Further growing the pre-heating temperature to 80 °C induces a comprehensive effect: alterations in asphaltene polarity, hindrance of asphaltene aggregation by alkyl side chains, and self-aggregation of alkyl side chains. This results in decreased WAT, along with the lowest rheological parameter values, with wax crystals appearing as the smallest and tightest dot-like shapes. At 80 °C, the optimal ratio between asphaltene polarity and alkyl side chain characteristics promotes strong nucleation and co-crystallization between asphaltene and wax crystals, maximizing asphaltene's ability to enhance the crude oil's low-temperature flowability. However, as the pre-heating temperature continues to rise, WAT decreases slightly and the rheological response starts to deteriorate.

Implementation of Cellulose-Based Filtration Aids in Industrial Sunflower Oil Dewaxing (Winterization): Process Monitoring, Prediction, and Optimization.

In the production of refined sunflower oil, waxes are removed during the winterization stage, and wax crystals are separated through filtration assisted by filtration aids. Commonly used filtration aids in oil refining include perlite and diatomaceous earth. After winterization, a significant amount of filter cake remains as a by-product and is treated as waste. Today, natural cellulose fibers are being promoted as filtration aids. Their advantages are numerous, both in the production process and from an environmental perspective. However, their only disadvantage is their higher cost. Therefore, in this study, 57 filtration cycles during the industrial sunflower oil winterization step using cellulose-based filtration aids were monitored. Different process parameters, including the pressure differential on the filter, the flow rate of filtered oil, constant pressure period, the quantity of filtered oil, filtration time, the quantity of pre-coating and dosing filtration aids, the volume of filtered oil, the concentration of dosing filtration aid, as well as the mass of separated waxes, were observed. Additionally, artificial neural networks were applied to predict process parameters, optimize the process, and, above all, determine the dosage of filtration aids, which will make the process more economical. The optimal filtration process is performed at a pressure differential of 3.3 bar, lasting a total of 39 h, with 32 h at constant pressure, resulting in 322,503 kg of filtered oil and 90.41 kg of waxes. The optimal quantity of cellulose-based filtration aids employed for pre-coat was 80 kg, and for dosing, 375 kg, with an optimal concentration of 0.12% w/w.

Effects of Different Shade Treatments on the Epidermal Wax Deposition of Hosta Genotypes with Different Glaucousness of Leaf Surface

Epidermal wax is strategically situated at the interface between plants and air; therefore, it plays a key role in plants’ interactions with their surroundings. It is also unstable and susceptible to light intensity. Hosta plants are shade-loving herbs with admirable flowers and leaves. Hosta ‘Halcyon’ and Hosta ensata F. Maek. are two species of Hosta with a glaucous and a glossy appearance, respectively. Light intensity can affect the composition of epicuticular wax on the leaf surface, which influences the leaf color phenotype and ornamental value. In this paper, the crystal micromorphology, content, and components of epicuticular wax on the leaves of two species of Hosta under different light conditions (10%-, 30%-, 50%-, 70%-, and 100%-intensity sunlight, relative light intensity (RLI)) have been studied using pot experiments. The results indicate that the epicuticular wax crystals of H. ‘Halcyon’ and H. ensata are tubular and platelet-like, respectively. The wax crystals of H. ‘Halcyon’ melted and formed a thick crust under 100% RLI, and those of H. ensata melted and formed a thick crust under 70% and 100% RLI conditions. The primary ingredients of the epicuticular wax of the two species of Hosta contained primary alcohols, alkanes, fatty acids, and esters; β-diketones were only detected in H. ‘Halcyon’. The quantity of epicuticular wax of H. ‘Halcyon’ reduced at first and then increased with an RLI increase, achieving its lowest value at 50% RLI, but that of H. ensata declined little by little. The amounts of C28 primary alcohols, C31 alkanes, and C18 fatty acids were significantly higher than those of other carbon atoms in the two genotypes of Hosta. The C31β-diketones content decreased with the increase in light intensity, which caused the white frost phenotype to gradually weaken in H. ‘Halcyon’.

Fabrication and characterization of starch-based bigels under phase control: Structural, physicochemical and 3D printing properties

Edible bigels are being developed to replace plastic fats in the food industry due to their ability to form semi-solids with controllable properties. In this study, biopolymer hydrogels were formed using modified corn starch with chitosan, betaine, and vanillin, whereas oleogels were formed from insect wax in soybean oil. The impact of varying the hydrogel-to-oleogel ratio on the properties of bigels was then examined. The rheological, oil-binding capacity (OBC), water-oil distribution, structural, and 3D printing characteristics of the bigels were evaluated. Higher hydrogel-to-oleogel ratios resulted in improved rheological properties of bigels. Hydrogen bonding, electrostatic interactions, and covalent Schiff base bonds played an important role in the formation and properties of the hydrogels. In contrast, the mechanical strength of the oleogels was mainly attributed to van der Waals attraction between the insect wax crystals. The optimized bigels had good OBC and mechanical strength. Furthermore, increasing the hydrogel-to-oleogel ratio induced a transition from W/O to O/W type structure in the bigels. A study of the potential application of the bigels as edible inks showed that bigels with a high hydrogel-to-oleogel ratio had better 3D printing characteristics. The findings of this study suggest that bigels can be successfully used as solid fat substitutes.

Flow assurance of waxy crude oil through thermal methods using nichrome wire and ultrasonic waves effects – Experimental investigation

When crude oil transportation stops for a certain period, the low temperature causes sufficient wax crystallization inside pipelines. During the crystallization, pipelines are exposed to a decrease in the inner diameter of the pipe, which leads to clogging and an increase in pressure drop. The higher pressure drop may require a huge pumping force to break the wax crystals and start the flow, increasing the level of risks and costs. Recently, many thermal and non-thermal methods have been used to alleviate wax deposition, such as surfactants, electric heating, and chemicals. In this study, flow assurance of heavy crude oil was investigated through pipelines using ultrasonic waves, nichrome wires, and combined methods. It includes conducting an experimental investigation on the ability of applying the ultrasonic effect and nichrome wires to improve the transportation performances of waxy crude oil through pipelines in a safe and environmentally friendly manner at the lowest costs while maintaining the characteristics of crude oil at high levels. The cooling rate was also studied at surrounding temperatures close to 30°C, 15°C, and 5°C. An experimental setup was developed to study the efficiency of the combined method in heating the pipeline. It was observed that the combined method was found effective in heating pipelines to higher temperatures in a shorter time. When the surrounding temperature was 30 °C, the combined method was able to raise the temperature of the crude oil to 80 °C in just 9.96 min, while the nichrome wires took 25 min to raise the temperature to 73 °C. As for the ultrasonic, it took 28 min to bring the temperature to 65.6 °C. It has also been proven that the highest cooling rate occurs at the walls of the pipeline. Therefore, the deposition of wax begins on the walls first and then moves successively to the middle.

Synergistic effect of modified ethylene-vinyl acetate and asphaltenes on improving the flow properties of model oil

The effect of alcoholic polyethylene-vinyl acetate (EVA) product ethylene-vinyl alcohol copolymer (EVAL) on the low-temperature flow properties of model oil containing asphaltene (ASP) was investigated. The change of wax crystal microscopic morphology of model oil before and after modification were examined, and the influence of asphaltene mass fraction on the rheological improvement effect of EVAL was analyzed. The composite system of EVAL and asphaltene significantly reduced the pour point, gel point, apparent viscosity, storage modulus and loss modulus of waxy oil at low temperatures. When the EVAL concentration is 400 ppm and the asphaltene mass fraction is 0.5 wt%, the synergistic effect of the two is optimal, which can reduce the pour point by 17 °C and the modulus value by more than 98%. The introduction of EVAL strengthens the interaction between asphaltenes and wax crystals, forming EVAL-ASP aggregates, which promote the adsorption of wax crystals on asphaltenes to form composite particles, and the polar groups prevent the aggregation of wax crystals and reduce the size of wax crystals, thus greatly improving the fluidity of waxy oils.

Morphology of Wax Crystals Affects the Rheological Properties and Thermal Conductivity of Waxy Oils

Morphology of Wax Crystals Affects the Rheological Properties and Thermal Conductivity of Waxy Oils

Herbarium specimens as tools for exploring the evolution of fatty acid-derived natural products in plants.

Plants synthesize natural products via lineage-specific offshoots of their core metabolic pathways, including fatty acid synthesis. Recent studies have shed light on new fatty acid-derived natural products and their biosynthetic pathways in disparate plant species. Inspired by this progress, we set out to develop tools for exploring the evolution of fatty-acid derived products. We sampled multiple species from all major clades of euphyllophytes, including ferns, gymnosperms, and angiosperms (monocots and eudicots), and we show that the compositional profiles (though not necessarily the total amounts) of fatty-acid derived surface waxes from preserved plant specimens are consistent with those obtained from freshly collected tissue in a semi-quantitative and sometimes quantitative manner. We then sampled herbarium specimens representing 57 monocot species to assess the phylogenetic distribution and evolution, of two fatty acid-derived natural products found in that clade: beta-diketones and alkylresorcinols. These chemical data, combined with analyses of 26 monocot genomes, revealed a co-occurrence (though not necessarily a causal relationship) between whole genome duplication and the evolution of diketone synthases from an ancestral alkylresorcinol synthase-like polyketide synthase. Limitations of using herbarium specimen wax profiles as proxies for those of fresh tissue seem likely to include effects from loss of epicuticular wax crystals, effects from preservation techniques, and variation in wax chemical profiles due to genotype or environment. Nevertheless, this work reinforces the widespread utility of herbarium specimens for studying leaf surface waxes (and possibly other chemical classes) and reveals some of the evolutionary history of fatty acid-derived natural products within monocots.

Higher alcohol acetoacetate aluminum chelates as pour point depressant and impedance reducing agent of biodiesel-diesel blends

The use of additives to improve the shortcomings of biofuels in application is one of the most economical and effective methods. In this study, a new type of tridentate higher alcohol acetoacetate aluminum chelate additive was synthesized originally and then synchronously used as pour point depressant and resistance reducing agent. The low temperature fluidity of biodiesel-diesel blends, including cloud point, cold filter plugging point and pour point could be reduced by the maximum of 6, 7 and 12 °C for B20 (20 vol% biodiesel + 80 vol% diesel), and 5, 5 and 9 °C for biodiesel after the treatment of higher alcohol acetoacetate aluminum chelate additive, named as C16-acac-Al. Furthermore, the impedance modulus determined by electrochemical impedance spectroscopy of the blends were greatly reduced after the treatment of C16-acac-Al, especially noiseless-free in the low frequency region, and the effects of temperature change on electrochemical impedance spectroscopy of treated blends was explored. At length, the mechanism on how the morphology of wax crystals affect electrochemical impedance spectroscopy of treated blends as well as the effect mechanism of the synthetic additives improving the low temperature fluidity of blends were put forward.

3-ketoacyl-CoA synthase 19 contributes to the biosynthesis of seed lipids and cuticular wax in Arabidopsis and abiotic stress tolerance.

Very-long-chain fatty acids (VLCFAs) are essential precursors for plant membrane lipids, cuticular waxes, suberin, and storage oils. Integral to the fatty acid elongase (FAE) complex, 3-ketoacyl-CoA synthases (KCSs) function as crucial enzymes in the VLCFA pathway, determining the chain length of VLCFA. This study explores the in-planta role of the KCS19 gene. KCS19 is predominantly expressed in leaves and stem epidermis, sepals, styles, early silique walls, beaks, pedicels, and mature embryos. Localized in the endoplasmic reticulum, KCS19 interacts with other FAE proteins. kcs19 knockout mutants displayed reduced total wax and wax crystals, particularly alkanes, while KCS19 overexpression increased these components and wax crystals. Moreover, the cuticle permeability was higher for the kcs19 mutants compared to the wild type, rendering them more susceptible to drought and salt stress, whereas KCS19 overexpression enhanced drought and salt tolerance. Disrupting KCS19 increased C18 species and decreased C20 and longer species in seed fatty acids, indicating its role in elongating C18 to C20 VLCFAs, potentially up to C24 for seed storage lipids. Collectively, KCS19-mediated VLCFA synthesis is required for cuticular wax biosynthesis and seed storage lipids, impacting plant responses to abiotic stress.

Relationship between Salmonella enterica attachment and leaf hydrophobicity, roughness, and epicuticular waxes: a focus on 30 baby-leaf salads.

The first step in the contamination of leafy vegetables by human pathogens is their attachment to the leaf surface. The success of this is influenced strongly by the physical and chemical characteristics of the surface itself (number and size of stomata, presence of trichomes and veins, epicuticular waxes, hydrophobicity, etc.). This study evaluated the attachment of Salmonella enterica to 30 baby-leaf salads and tested whether the differences found among them were related to the following leaf traits: hydrophobicity, roughness, and epicuticular waxes. Differences in susceptibility to contamination by S. enterica were found between the 30 baby-leaf salads investigated. The lowest attachment was found in wild lettuce (Lactuca serriola L.) and lamb's lettuce 'Trophy F1' (Valerianella locusta [L.] Laterr.), with values of 1.63 ± 0.39 Log(CFU/cm2) and 1.79 ± 0.54 Log(CFU/cm2), respectively. Attachment was correlated with hydrophobicity (measured as contact angle) (r = -0.39) and epicuticular waxes (r = -0.81) but not with roughness (r = 0.24). The most important wax components for attachment were alcohols and, in particular, the three-dimensional (3D) wax crystals of C26 alcohol, but fatty acids probably also had a role. Both these compounds increased hydrophobicity. The presence of thymol, whose antimicrobial properties are well known, was found in lamb's lettuce. The findings of this study can help to predict and control the attachment and contamination of leafy salads by enterobacteria. They also provide useful information for breeding programs aiming to develop cultivars that are less susceptible to human pathogens, enhancing the food safety of vegetables. © 2024 The Author(s). Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Evaluated the effects of nature α-olefins (limonene, β-caryophyllene and camphene) as additives on the cold flow properties of diesel fuel

Pour Point Depressants (PPDs) are currently deemed the most efficacious methodology to enhance the low-temperature fluidity of diesel fuel. However, the majority of these PPDs present complications such as high costs, intricate synthesis procedures, and exhibit potential biotoxicity. To address these challenges, natural α-olefins (β-caryophyllene, limonene and camphene) with 1-hexadecene in varying ratios (5, 10, 15, 20, 25:1) were successfully polymerized at ordinary temperature as poly-α-olefine pour point depressant (PAO PPD), and designated as 1-hexadecane-β-caryophyllene (C16-SZX), 1-hexadecane-limonene (C16-NMX), 1-hexadecane-camphene (C16-KX). An in-depth examination was conducted on the cold filter plugging point (CFPP) and solid point (SP) of both untreated and PPD-added diesel. Subsequently, the finding clearly demonstrates that the CFPP and SP were separately reduced 10 and 32 °C in diesel treated with a molar ratio of 10:1 of C16-KX at the dosage of 2000 ppm. Meanwhile, a comprehensive study was undertaken to investigate the influence of natural α-olefins on the reduction of low-temperature fluidity in diesel fuel. The inhibitory impacts of natural molecular formations on wax crystals were meticulously scrutinized, particularly their roles in modifying the growth patterns of these crystals. Eventually, a novel mechanism by which poly-α-olefine alters the crystalline behavior of wax crystals is proposed.

Light-responsive transcription factors VvHYH and VvGATA24 mediate wax terpenoid biosynthesis in Vitis vinifera.

The cuticular wax that covers the surfaces of plants is the first barrier against environmental stresses and increasingly accumulates with light exposure. However, the molecular basis of light-responsive wax biosynthesis remains elusive. In grape (Vitis vinifera), light exposure resulted in higher wax terpenoid content and lower decay and abscission rates than controls kept in darkness. Assay for transposase-accessible chromatin with high-throughput sequencing and RNA-seq data were integrated to draw the chromatin accessibility and cis-elements regulatory map to identify the potential action sites. Terpenoid synthase 12 (VvTPS12) and 3-hydroxy-3-methylglutaryl-CoA reductase 2 (VvHMGR2) were identified as grape wax biosynthesis targets, while VvHYH and VvGATA24 were identified as terpenoid biosynthesis activators, as more abundant wax crystals and higher wax terpenoid content were observed in transiently overexpressed grape berries and Nicotiana benthamiana leaves. The interaction between VvHYH and the open chromatin of VvTPS12 was confirmed qualitatively using a dual luciferase assay and quantitatively using surface plasma resonance, with an equilibrium dissociation constant of 2.81 nm identified via the latter approach. Molecular docking simulation implied the structural nature of this interaction, indicating that 24 amino acid residues of VvHYH, including Arg106A, could bind to the VvTPS12 G-box cis-element. VvGATA24 directly bound to the open chromatin of VvHMGR2, with an equilibrium dissociation constant of 8.59 nm. Twelve amino acid residues of VvGATA24, including Pro218B, interacted with the VvHMGR2 GATA-box cis-element. Our work characterizes the mechanism underlying light-mediated wax terpenoid biosynthesis and provides gene targets for future molecular breeding.