Before extracting pectin and oligosaccharides (OS) and other biologically active components from plant materials, the raw materials should undergo a defatting process in order to intensify the extraction and isolation processes. Fats and waxes have been used in various formulations for soap-making, skin and hair care, cooking and healing purposes. They give consistency and stability to body butters, healing balms, ointments, lip balms and creams. In addition, because waxes are not absorbed by the skin, they provide an outer layer of protection and a barrier against inclement weather, thereby also helping to retain moisture. Plant waxes embedded into the cuticle are called “intracuticular waxes”, whereas waxes superimposed onto the cuticle are called “epicuticular waxes”. Additional minor amounts of cutinized polysaccharide fiber, including cellulose and hemicellulose, and pectin, link the plant cuticle to the underlying cell wall [1] . Several properties of plant cuticles are mainly based on the waxes. Intracuticular waxes mainly function as water transpiration barriers [2] , whereas epicuticular waxes strongly influence the wettability, self-cleaning behavior and the light reflection at the cuticle interface [3] .

Plant waxes consist of a mixture of aliphatic hydrocarbons and their derivatives with carbon chain lengths between 20 atoms and 40 atoms, and in the case of esters (two connected chains) about 60 atoms. The main component classes are usually primary and secondary alcohols, ketones, fatty acids, and aldehydes. Alkanes are widely distributed but occur usually in low concentrations. However, many plant waxes do not match the chemical definition of true waxes. Triterpenoids, for example, occur in high concentrations in the epicuticular coatings of grapes. Other plant waxes contain polymeric components such as polymerized aldehydes which are only slightly soluble in chloroform. Furthermore, the waxes of conifer needles contain estolides, e.g. oligomeric hydroxy fatty acids [4] . The exudates of some ferns and angiosperms, in particular several members of the Primulaceae, are mainly composed of flavonoids and are termed farinose instead of waxes [5] . Recently an increasing number of publications have reported the discovery of new wax components. Thus, a long list of rare and uncommon ingredients is known, including methyl-branched aliphatics and molecules combining aliphatic and cyclic parts [6] .

Several reviews have addressed the chemical composition of plant waxes [6] [7] [8] , but it must be noticed, that nearly all existing data of the chemical composition was based on solvent-extracted waxes. These are mixtures of epicuticular and intracuticular waxes, which may be chemically different, as shown by Jetter et al. [9] for the waxes of Prunus laurocerasus and recently for the waxes of Taxus baccata [10] . By the development of more selective methods, it is now possible to analyze separately the intra- and epicuticular wax fractions to get a better understanding of the wax chemistry and even the molecular architecture of single epicuticular wax sculptures [11] .

The natural wax on apple fruit contains about fifty individual components belonging to at least half a dozen chemical groups. Two major classes of chemicals are often found. The major cyclic component of apple fruit wax is called ursolic acid and is highly water repellent [12] .

Application of F&W isolation could effectively improving pectin yield and quality in the production technology and bring another valuable byproduct, which the aim of this work.

For this purpose, the isolation of plant waxes is performed either by dissolution in organic solvents or with mechanical methods. Most waxes are easily soluble in organic solvents and are completely dissolved within a few seconds. Some waxes contain components, which are only slightly soluble in organic solvent at ambient temperatures, e.g. aldehydes in sugar cane, or triterpenoids in the wax layer on grapes. Such waxes need to be dissolved in warm chloroform and it may be necessary to verify whether they have been dissolved completely. The complete extraction of the intracuticular waxes may require several minutes. Generally, solvent-extracted waxes contain intracuticular waxes and can be contaminated by other organic solvent soluble components from inside the tissues [6] .

This work combines a number of plant processing technologies, but only focused on a defatting technique applied to the different kinds of plant species: sunflower head residue, fruit wastes, roots of plants and grape seeds. For a quantitative wax extraction, the plant material, e.g. roots, seeds, fruits pulp or pomace are immersed in the solvent. The materials for defatting were then consequently used in pectin production and plant oligosaccharides and polyphenol extraction procedures.

Apricot, grape (Toify, Aleatico and Isabella), pumpkin, apple, and peach were harvested on season and purchased from local market (Dushanbe, Tajikistan). Sunflower head residues were obtained from seedless sunflower heads (Helianthus annuus, Avangard) that were harvested from a suburb of Dushanbe, Tajikistan).

Eremurus h. were planted, cultivated, and harvested by the employees of the Biological Station of the Institute of Botany and Plant Physiology and Genetics, Tajik Academy of Sciences, in 2015 in Syjahkuh Hill, Tajikistan.

All chemicals were from Sigma-Aldrich (Reachim, Russia) and used as it is.

The isolation of F&W from fruit wastes consisted of two steps. First, the fruit wastes were dried, 50 g for each were then mixed with 300 ml hexane/alcohol (3:2, v/v) in a Soxhlet extractor (Figure 1), and fluxed at following setting:

Extraction time: 5 hours

Temperature of the water bath, 80˚C

Length of each refluxing cycle, 30 min.

Number of cycle, 12

At the end of fluxing-leaching, the solvents were separated from the fruits wastes by filtration, and then evaporated to obtain a mixture of solid and semi- solid residue, which were washed with ether for a few times, then dried in oven at 37˚C. The intermediates thus obtained were referred as total extracts that consist of fats, waxes, polyphenols and sugars compounds.

In the second step, the solid phase was extracted with 10% alcohol-water solution to separate phenolic and sugar compounds in the liquid phase from F&W in solid phase.

F&W contents were calculated by subtracting the amounts of AWSC from the amounts of the total extractant.

The collected root tubers of the Eremurus h. were washed with tap water, dried at room temperature for one day. The dried root tubers (500 g) were cut, using a kitchen knife, into thin sheet-like pieces and spread out on filter paper and left to completely dry at 28˚C - 30˚C for additional 10 days. The dried sheets were milled for 10 minutes in a laboratory mill (Retsch GM 200, Germany) to obtain fine powders. The powders were processed for separating F&W, polyphenols and sugars compounds from the raw materials by the methods described above with slight modification:

Leaching reagent, ethyl acetate

Ratio between solid and liquid, 87.66 g: 750 ml

Length of a reflux cycle, 40 - 45 min

F&W and AWSC were separated from grape pomace (grape cake) and grape seeds from the grapes growing in three different locations in Tajikistan: Toify, Aleatico and Isabella. The grape pomace and seeds were dried at 40˚C, grounded to a particle sizes between 0.8 - 2.0 mm. The extraction and separation were operated under the conditions similar to that used for Eremurus h. roots, except for 1) 12 - 14 refluxing cycles were applied for the extraction of fats, waxes, polyphenols, and sugars, and 2) the ratio of solid to liquid in extraction was 1:10, w/v.

The F&W sample were dried over glass slides to form films, which were analyzed by FT-IR spectroscopy using a Spectrum 65 FT-IR (Perkin Elmer) spectrometer equipped with MIRACLE ATR (ZNSE). Each recorded spectrum is the average of 16 scans in the range 4000 - 600 cm⁻¹ with a 4 cm⁻¹ resolution in transmission on a dried sample, with a background spectrum recorded before each analysis. Three spectra were taken and each one was analyzed and fitted using Perkin Elmer Spectrum, version 10.03.07 software. The shift in spectra in the repeat measurements performed on the same sample was about 2%.

Extracts in hexane/ethanol were the mixtures of fats, waxes, polyphenols, and sugars. After removing the leaching reagents, the residues appeared as highly hygroscopic ointment-like materials, or viscous fluids, or powders, depending the types of the fruits. The time curves of extraction were shown in Figure 2.

For the residues from apple, peach and sunflower processing, the extraction peaks were recorded at around 1 - 1.5 hours after the extraction is started that is much earlier than the record of 3^rd hour for pumpkin and apricot. The differences

could be attributed to the differences in solubility of fats, waxes, polyphenols, and sugars in the hexane/ethanol solution and their diffusion coefficient through cell walls to the environment.

Table 1 shows the yields of extracted materials from fruit wastes. The resulting F&W yield in pumpkin was less in comparison to other studied fruits. F&W quantities increased from pumpkin, peach, sunflower, apple to apricot. The quantities of AWSC isolated from total extracted materials increased as apple < to apricot < pumpkin < sunflower < peach. No correlation between F&W and AWSC contents were found.

The extraction kinetics also expressed as the time-dependence of weight loss of the raw materials. The typical curve was shown in Figure 3, using apple pomace as an example. Figure 3 illustrates a three-stage diffusion kinetics: the first hour was characterized with a sharp weight loss (weight loss, 6.3%) followed by a retarding diffusion that lasted for 3 hours (weight loss, 3.5%), and a slow mass diffusion period in the last 1 hour (weight lost, 0.2%), reaching at a total weight loss of 10% in 5 hours. Such pattern of extraction was likely due to simultaneous isolation of a number of compounds in first hours of extraction having both hydrophobic and hydrophilic chains including phenols and sugar derivatives, which were further isolated from F&W materials.

The F&W occupied a big fraction of 80% of the total materials extracted from