KR20180037907A - Tandem neopentyl glycol-based amphiphiles and uses thereof - Google Patents

Abstract

The present invention relates to a tandem neopentyl glycol-based amphipathic compound, a production method thereof, and a method for extracting, solubilizing, stabilizing, or crystallizing a membrane protein using the same. According to the present invention, the use of tandem neopentyl glycol-based amphipathic compound not only derives excellent membrane protein solubilization effects but also allows the membrane protein to be stably stored in an aqueous solution for a long period of time, thereby being applied to functional analysis and structural analysis for the same. Since the structural and functional analysis on the membrane protein is one of the areas of the greatest interest in current biology and chemistry while more than half of currently developed new drugs target the membrane protein, application to studies on structures of the membrane protein closely related to the development of new drugs is possible.

Description

Tandem neopentyl glycol-based amphipathic compounds and their use Tandem neopentyl glycol-based amphiphiles and uses thereof < RTI ID = 0.0 >

The present invention relates to a newly developed amphipathic compound based on tandem neopentyl glycol and a method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein using the same.

Membrane proteins play an important role in biological systems. Because these bio-macromolecules contain hydrophilic and hydrophobic moieties, amphiphilic molecules are needed to extract membrane proteins from the lipid environment and to solubilize and stabilize them in aqueous solutions.

In order to analyze the structure of membrane proteins, it is necessary to obtain good quality membrane protein crystals. For this purpose, structural stability of membrane proteins in aqueous solution should be preceded. The number of existing amphipathic molecules that have been used in membrane protein research is more than 100, but only about 5 have been actively used for membrane protein structure studies. These five ampholytic molecules are OG (n-octyl-β-D-glucopyranoside), NG (n-nonyl-β-D-glucopyranoside) dodecyl-? -D-maltopyranoside, and LDAO (lauryldimethylamine- N- oxide) (Non-Patent Document 1 and Non-Patent Document 2). However, many membrane proteins surrounded by these molecules have considerable limitations in studying the function and structure of membrane proteins that utilize these molecules because their structures are easily denatured or aggregated and rapidly lose their function. This is because conventional molecules have a simple chemical structure and do not exhibit sufficiently diverse properties.

In order to analyze the structure of membrane proteins, it is important to maintain the structural stability of membrane proteins in aqueous solution. Many membrane proteins have not yet been identified and their structural characteristics vary. The number of membrane proteins available has been limited. Therefore, according to the necessity of developing various amphipathic molecules different from the flexible alkyl chains of existing amphipathic molecules, the present inventors repeatedly linked neopentyl glycol having low fluidity to introduce two quaternary carbons into the center of the molecule, The present inventors have completed the present invention by developing a novel amphipathic compound which promotes crystallization of the membrane protein by greatly restricting the fluidity of the whole.

 S. Newstead et al., Protein Sci. 17 (2008) 466-472.  S. Newstead et al., Mol. Membr. Biol. 25 (2008) 631-638.

An object of the present invention is to provide a compound represented by the formula (1).

Another object of the present invention is to provide a composition for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein comprising the above compound.

It is a further object of the present invention to provide a process for the preparation of said compounds.

It is still another object of the present invention to provide a method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein using the above compound.

One embodiment of the present invention provides a compound represented by Formula 1:

[Chemical Formula 1]

In Formula 1,

Wherein R ^1, R ² and Each R ³ may independently be a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;

X ¹ , X ² and X ³ may each independently be a saccharide linked with oxygen;

Y ¹ , Y ² and Y ³ are oxygen (O); And

M may be 0 or 1;

As used herein, the term "saccharide " refers to a compound that is relatively small in carbohydrate and is soluble in water to give a sweet taste. Saccharides are classified into monosaccharides, disaccharides and polysaccharides depending on the number of molecules constituting the sugar.

The saccharide used in the above embodiments may be a monosaccharide or a disaccharide and may specifically be glucose or maltose and more specifically may be maltose but is not limited thereto Do not.

The saccharide may act as a hydrophilic group. In the compound according to one embodiment of the present invention, three saccharides, which are hydrophilic groups, are connected in parallel to minimize the increase in the length of the hydrophilic group while reducing the size of the complex with the membrane protein. If the size of the complex of the compound and the membrane protein is small, high quality membrane protein crystals can be obtained (GG Prive, Methods 2007, 41, 388-397). In particular, amphiphilic molecules with a small hydrophilic group such as glucoside can have an excellent effect on the crystallization of the membrane protein.

The above R ¹ , R ² and R ³ can act as a hydrophobic group. Compounds according to one embodiment of the present invention were introduced into three different alkyl group hydrophobic groups in order to optimize the hydrophile-lipophile balance.

In the compound according to one embodiment of the present invention, a hydrophobic group and a hydrophilic group may be connected by an ether bond. In other words, a linker was introduced to sufficiently secure the fluidity of the alkyl chain while maintaining the rigidity of the center of the molecule. Specifically, the ether linkage may be introduced using a tandem neopentyl glycol linker.

Specifically, R ¹ , R ² and R ³ may be a C ₇ -C ₁₄ alkyl group; Wherein R ^1, R ^2, and R ³ can be the same; Y ¹ , Y ² and Y ³ may be oxygen; X ¹ to X ³ may be maltose linked with oxygen; And m may be 0.

The above R ¹ , R ² and R ³ may be a C ₇ -C ₁₄ alkyl group; Wherein R ^1, R ^2, and R ³ can be the same; Y ¹ , Y ² and Y ³ may be oxygen; X ¹ to X ³ may be maltose linked with oxygen; And m may be 1.

In an embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₉ -C ₁₄ alkyl group; Wherein R ^1, R ^2, and R ³ is the same; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 0 or 1 is referred to as "TNMs (tandem neopentyl glycol-derived maltosides) ".

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₉ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 1 is named "TNM-C9L" and can be represented by the following formula (2).

(2)

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₀ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 1 is referred to as "TNM-C10L ", and can be represented by the following formula (3).

(3)

More specifically, in one embodiment of the present invention, the R^One, R²And R³C₁₁ Alkyl group; Y^One, Y² And Y³Is oxygen; The X^One To X³Is maltose linked with oxygen; The compound wherein m is 1 is referred to as "TNM-C11L ", and can be represented by the following formula (4).

[Chemical Formula 4]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₂ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 1 is named "TNM-C12L" and can be represented by the following formula (5).

[Chemical Formula 5]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₃ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 1 is named "TNM-C13L" and can be represented by the following formula (6).

[Chemical Formula 6]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₄ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 1 is referred to as "TNM-C14L ", and can be represented by the following formula (7).

(7)

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₁ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 0 is referred to as "TNM-C11S ", and can be represented by the following formula (8).

[Chemical Formula 8]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₂ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 0 is named "TNM-C12S" and can be represented by the following formula (9).

[Chemical Formula 9]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₃ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 0 is named "TNM-C13S ", and can be represented by the following formula (10).

[Chemical formula 10]

More specifically, in one embodiment of the present invention, R < ¹ >, R < ² & R ³ is a C ₁₄ alkyl group; Y ¹ , Y ² and Y ³ are oxygen; X ¹ to X ³ are maltose linked with oxygen; The compound wherein m is 0 is named "TNM-C14S ", and can be represented by the following formula (11).

(11)

The compound according to another embodiment of the present invention may be, but is not limited to, an amphipathic molecule for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein.

As used herein, the term "amphipathic molecule" refers to a molecule in which hydrophobic groups and hydrophilic groups are present in a molecule and have affinity for both polar and non-polar solvents. The phospholipid molecules present in the surfactant or cell membrane are amphiphilic molecules having a hydrophilic group at one end and a hydrophobic group at the other end and are characterized in that they form micelles or liposomes in an aqueous solution. Amphiphilic molecules tend to be insoluble in water because hydrophilic groups have polarity but nonpolar groups coexist. However, when the concentration exceeds a certain threshold concentration (critical micelle concentration, CMC), the hydrophobic group aggregates into hydrophobic interactions and micelles are formed on the surface of the hydrophilic group, thereby increasing the solubility in water significantly.

The method for measuring CMC is not particularly limited, but it can be used in a method widely known in the art, and can be measured by, for example, a fluorescence staining method using diphenylhexatriene (DPH).

The compound according to one embodiment of the present invention may have a critical micelle concentration (CMC) in an aqueous solution of 0.0001 mM to 1 mM, specifically 0.0001 mM to 0.01 mM, more specifically 0.0004 mM to 0.001 mM, But is not limited thereto.

In the case of DDM, which has been mainly used for membrane protein research, the TNMs of this specific example have a CMC value much lower than that of DDM, compared with the critical micelle concentration of 0.17 mM. Therefore, it was confirmed that TNMs are more advantageous than DDM because micelles are easily formed in small amounts, and membrane proteins can be effectively analyzed and analyzed using small amounts.

Still another embodiment of the present invention provides a composition for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein comprising the above compound.

The composition may be, but is not limited to, a formulation of micelles, liposomes, emulsions or nanoparticles.

The micelle may have a radius of 2.0 nm to 60.0 nm and may be specifically 3.0 nm to 55.0 nm. More specifically, the micelle formed by the TNMs according to the embodiments of the present invention may have a radius of 3.0 nm to 50.0 nm But is not limited thereto.

The method for measuring the radius of micelles is not particularly limited, but a method well known in the art can be used and can be measured using, for example, dynamic light scattering (DLS) experiments.

It can be seen that the size of the micelle formed by TNMs is wide.

The micelle, liposome, emulsion or nanoparticle may contain a membrane protein therein. That is, the micelles, liposomes, emulsions or nanoparticles can extract and enclose membrane proteins present in the cell membrane. Therefore, it is possible to extract, dissolve, stabilize, crystallize or analyze the membrane protein by the micelle.

The composition may further comprise a buffer, etc., which may aid in the extraction, solubilization, stabilization or analysis of the membrane protein.

The present invention also provides a process for preparing a compound represented by the following general formula (1), which comprises the following steps 1) to 7):

1) Synthesis of 5,5-bis-bromomethyl-2,2-dimethyl- [1,3] dioxane and 5,5-bis-bromomethyl- alcohol to form a dialkylated diol;

2) synthesizing a trialkylated monool by adding 1-bromoalkane to the product of step 1);

3) allylating the alcohol by adding allyl iodide to the product of step 2);

4) hydroboration oxidation of the product of step 3) to synthesize a trialkylated monool;

5) To a solution of 4- (bromomethyl) -1-methyl-2,6,7-trioxabicyclo [2.2.2] -octane (4- (bromomethyl) , 6,7-trioxabicyclo [2.2.2] -octane) to synthesize a trialkylated triol;

6) performing a glycosylation reaction on the product of step 5) to introduce a protecting group-attached saccharide; And

7) a step of performing a deprotection reaction on the product of step 6);

[Chemical Formula 1]

In Formula 1,

Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;

X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;

Y ¹ , Y ² and Y ³ are oxygen (O); And

M is 1.

The compound prepared according to the above method may be a compound represented by the above formulas (2) to (7).

The present invention also provides a process for preparing a compound represented by the following general formula (1), which comprises the following steps 1) to 8):

1) Synthesis of 5,5-bis-bromomethyl-2,2-dimethyl- [1,3] dioxane (5,5-bis-bromomethyl- ≪ / RTI > to form a dialkylated diol compound;

2) synthesizing a trialkylated monool-type compound by adding 1-bromoalkane to the product of step 1);

3) allylating the alcohol by adding allyl iodide to the product of step 2);

4) hydroboration oxidation of the product of step 3) to synthesize a trialkylated monool compound;

5) synthesizing a trialkylated aldehyde-type compound through an alcohol oxidation reaction of the product of step 4);

6) synthesizing a trialkylated triol compound by adding formaldehyde and water-soluble alkaline ethanol to the product of step 5);

7) performing a glycosylation reaction on the product of step 6) to introduce a protecting group-attached saccharide; And

8) carrying out a deprotection reaction on the product of step 7);

[Chemical Formula 1]

In Formula 1,

Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;

X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;

Y ¹ , Y ² and Y ³ are oxygen (O); And

M is zero.

The compound prepared according to the above method may be a compound represented by the above formulas (8) to (11).

Still another embodiment of the present invention provides a method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein. Specifically, there is provided a method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein, comprising treating a membrane protein with a compound represented by the following formula (1) in an aqueous solution:

[Chemical Formula 1]

In Formula 1,

Wherein R ^1, R ^2, and Each R ³ may independently be a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;

X ¹ , X ² and X ³ may each independently be a saccharide linked with oxygen;

Y ¹ , Y ² and Y ³ are oxygen (O); And

M may be 0 or 1;

Specifically, R ¹ , R ² and R ³ may be a C ₇ -C ₁₄ alkyl group; Wherein R ^1, R ^2, and R ³ can be the same; Y ¹ , Y ² and Y ³ may be oxygen; X ¹ to X ³ may be maltose linked with oxygen; And m may be 0.

The above R ¹ , R ² and R ³ may be a C ₇ -C ₁₄ alkyl group; Wherein R ^1, R ^2, and R ³ can be the same; Y ¹ , Y ² and Y ³ may be oxygen; X ¹ to X ³ may be maltose linked with oxygen; And m may be 1.

The compound may be 11 kinds of compounds represented by the following formula (2) according to an embodiment of the present invention, but is not limited thereto.

As used herein, the term "membrane protein" is a generic term for proteins or glycoproteins that enter the cell membrane lipid bilayer. It exists in various states, such as through the entire cell membrane, in the surface layer, or in the cell membrane. Examples of membrane proteins include, but are not limited to, enzymes, peptide hormones, receptors such as local hormones, acceptors such as sugars, ion channels, and cell membrane antigens.

The membrane protein includes any protein or glycoprotein that is introduced into the cell membrane lipid bilayer, and specifically includes uric acid-xanthine / H ⁺ symporter, MelB (melibiose permease), LeuT (Leucine transporter), GPCRs (G- protein coupled receptors, or a combination of two or more thereof.

As used herein, the term "extraction of membrane protein" means separating the membrane protein from the membrane.

As used herein, the term "solubilization of membrane protein" refers to dissolving a water-insoluble membrane protein in an aqueous solution in micelles.

The term "stabilization of membrane protein " as used herein means to stably preserve the tertiary or quaternary structure so that the structure and function of the membrane protein are not changed.

As used herein, the term "crystallization of membrane protein" means forming crystals of membrane protein in solution.

As used herein, the term "analysis of membrane protein" means analyzing the structure or function of membrane protein. In the above embodiments, the membrane protein can be analyzed by a known method. For example, it is possible to analyze the structure of the membrane protein using, for example, electron microscopy.

Using the tris- or neopentyl glycol-based compounds according to embodiments of the present invention, the membrane protein can be stably stored in an aqueous solution for a long period of time in the aqueous solution, and thus the membrane protein can be utilized for its functional analysis and structural analysis.

Membrane protein structure and function analysis is one of the most interesting fields in current biology and chemistry. Since more than half of the new drugs being developed are targeted to membrane proteins, studies on protein structures that are closely related to the development of new drugs Application is possible.

In particular, the compounds according to the embodiments of the present invention have a small hydrophilic group, which not only has an excellent effect on the crystallization of the membrane protein but also has a problem that the stability of the membrane protein generated when a small hydrophilic group is used is somewhat poor And exhibits better membrane protein stability than conventional compounds.

In addition, the compounds according to the embodiments of the present invention can be synthesized by a simple method from readily available starting materials, so that mass production of a compound for membrane protein research is possible.

Figure 1 shows the synthesis scheme of the TNMs of the present invention.
Figure 2 shows the synthesis scheme of the TNMs of the present invention.
Figure 3 shows the chemical structure of the TNMs of the present invention.
Figure 4 is a dynamic light scattering (DLS) profile of micelles formed by (a) TNM-Ls and (b) TNM-Ss. We obtained information on the size of the micelles based on the number of analyzes.
FIG. 5 shows the structural stability of the LHI-RC complex by (a) TNM-Ls or (b) TNM-Ss used in a concentration of CMC + 0.04 wt% in aqueous solution and monitoring the absorbance at 875 nm .
FIG. 6 shows the structural stability of LHI-RC complex by (a) TNM-Ls or (b) TNM-Ss at a concentration of CMC + 0.2 wt% in aqueous solution as measured by monitoring the absorbance at 875 nm .
FIG. 7 shows the results of measurement of the structural stability of UapA protein by (a) TNM-Ls or (b) TNM-Ss used in a concentration of CMC + 0.2 wt% in aqueous solution using the CPM assay.
FIG. 8 shows the results of measurement of the structural stability of UapA protein by (a) TNM-Ls or (b) TNM-Ss used in a concentration of CMC + 0.04 wt% in aqueous solution using the CPM assay.
FIG. 9 shows the results of measurement of the stability of LeuT protein by TNMs using a scintillation proximity assay (SPA) at a concentration of CMC + 0.04 wt%.
FIG. 10 shows the results of measurement of the stability of LeuT protein by TNMs using CMC + 0.2 wt% concentration using SPA (scintillation proximity assay).
FIG. 11 shows the results of measuring the activity of [ ³ H] -dihydroalprenolol (DHA) at regular intervals while incubating at room temperature for 4 days to measure the effect of β ₂ AR on TNMs stability.
12 shows the results of measuring the activity of [ ³ H] -dihydroalprenolol (DHA) at regular intervals while incubating at room temperature for 4 days to measure the effect of β ₂ AR on TNM-C12L stability .

Hereinafter, the present invention will be described in more detail in the following Examples. It should be noted, however, that the following examples are illustrative only and do not limit or limit the scope of the present invention. It will be understood by those of ordinary skill in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.

< Example  1> TNM - Ls  Synthesis method

The synthesis scheme of TNM-Ls is shown in Fig. Six compounds of TNM-Ls (Tandem Neopentyl Glycol Maltosides) were synthesized according to the synthesis methods of <1-1> to <1-7> shown in FIG.

<1-1> Dialkylated Of dialkylated diols  The general synthetic procedure (step a in figure 1)

Was added dropwise a solution of NaH (4.0 eq.) Is mixed with DMF, under each of the alkyl alcohol (3.0 eq.) 0 ℃, N ₂ atmosphere, followed by stirring for 15 minutes the resulting mixture at room temperature. After the addition of 5,5-bis-bromomethyl-2,2-dimethyl- [1,3] dioxane (A) (1.0 eq.), The reaction mixture was warmed to 120 & . After cooling to room temperature, the reaction was quenched with ice water and the organic material was extracted three times with diethyl ether. The combined organic layers were washed with brine (brine), dried over anhydrous Na ₂ SO ₄ then concentrated by rotary evaporation. After complete evaporation of the solvent, the residue was dissolved in a 1: 1 mixture of CH ₂ Cl ₂ and MeOH, p-toluenesulfonic acid (p-TSA) monohydrate (catalytic amount) was added and stirred at room temperature for 2 hours. Neutralizing the reaction mixture with aqueous saturated NaHCO ₃ solution and reduced the volume of the solvent by rotary evaporation. The reaction mixture was partitioned between CH ₂ Cl ₂ and H ₂ O. After the collected organic layer was washed with brine, dried over anhydrous Na ₂ SO _4, and concentrated in vacuo. The ether-containing diol (B1-B6) of white solid (92-94% (two steps)) was obtained by flash column chromatography (EtOAc / hexane).

<1-2> Trialkylated Monool ( 트리화화  mono- be ) (Step b of Figure 1)

To a suspension of NaH (1.3 eq.) In anhydrous DMF was slowly added a solution of the diol derivative (B1-B6) (1.0 eq) mixed in the anhydrous DMF under argon at 0 ° C. After stirring for 15 minutes, 1-bromoalkane (RBr) (1.3 eq.) Was added to the mixture and the temperature was increased to 100 &lt; 0 &gt; C. After allowing to stand the reaction mixture at this temperature for 4 hours, cooled to room temperature the reaction was terminated with H ₂ O. The reaction mixture was extracted twice with CH ₂ Cl ₂ , washed with brine, and dried over anhydrous Na ₂ SO ₄ . The organic layer was concentrated in vacuo and the resulting residue was purified by silica gel column chromatography (EtOAc / hexanes) to give a trialkyl containing monool (C1-C6) in the oily liquid form (85-90%).

<1-3> General synthesis procedure of allylation (step c in FIG. 1)

To a suspension of NaH (2.0 eq.) In anhydrous DMF was slowly added a solution of the trialkylated monoole derivative (C1-C6) (1.0 eq.) Mixed in the anhydrous DMF under argon at 0 ° C. After stirring for 15 minutes, allyl iodide (1.5 eq) was added to the mixture and the temperature was increased to 60 &lt; 0 &gt; C. After allowing to stand the reaction mixture at this temperature for 4 hours, cooled to room temperature the reaction was terminated with H ₂ O. The reaction mixture was extracted twice with CH ₂ Cl ₂ , washed with brine, and dried over anhydrous Na ₂ SO ₄ . The organic layer was concentrated in vacuo and the resulting residue was purified by silica gel column chromatography (EtOAc / hexanes) to give the trialkylated allyl compound (D1-D6) in the oily liquid form (90-95%).

<1-4> General synthesis procedure for hydroboration (step d in FIG. 1)

Fully distilled anhydrous THF (3 ml) and the trialkylated allyl compound (D1-D6) (1.65 g, 2.77 mmol) were placed in a dry round bottom flask under N ₂ and cooled to 0 ° C in an ice bath. It was slowly added a 1M BH ₃ solution was mixed with THF (3 ml) to the mixture, and the reaction mixture was stirred at 0 ℃ for 10 hours. The reaction mixture was then poured into 3 M NaOH solution (2 ml), stirred for an additional 20 min, and 30% H ₂ O ₂ solution (2 ml) was added. The mixture was stirred at room temperature for 2 hours. After saturation with K ₂ CO ₃ , the reaction mixture was extracted with diethyl ether. The organic layer was dried over anhydrous Na ₂ SO ₄ and concentrated in vacuo. The resulting mixture was purified by silica gel column chromatography (EtOAc / hexanes) to give the trialkylated hydoroborated compound (E1-E6) as an oily liquid (55-60%).

<1-5> Trialkylated Of trialkylated tri-ol  The general synthesis procedure (step e in Figure 1)

To a solution of the trialkylated hydoroborated compound (E1-E6) (1.0 eq) mixed in DMF (10 ml) at 0 <0> C was added NaH (3.0 eq) under argon and the reaction mixture was stirred for 12 min . 4- (bromomethyl) -1-methyl-2,6,7-trioxabicyclo [2.2.2] -octane (3.0 eq) dissolved in THF (3 ml) was slowly added. The resulting mixture was heated at 100 &lt; 0 &gt; C for 24 hours. After quenching with water, the solvent was removed under reduced pressure. The solid residue was dissolved in diethyl ether, washed with brine, dried over anhydrous Na ₂ SO _4. After concentrating the organic layer with a rotary evaporator, the residue was dissolved in a DCM / MeOH (10 ml / 10 ml) mixture. To this solution was added a few drops of concentrated HCl and the resulting mixture was heated at 50 &lt; 0 &gt; C for 4 hours. After neutralization with NaOH and concentration of the reaction mixture, the resulting residue was purified by column chromatography (EtOAc / Hexane) to give the trialkylated triol (F1-F6) in the oily liquid form (45-50%).

<1-6> General synthesis procedure of glycosycosylation reaction (step f in FIG. 1)

This was in accordance with the synthesis method of Chae , PS et al. (Nat. Methods 2010, 7, 1003.). 2,4,6-collidine (1.0 eq.) Was added to a stirred solution of the trialkylated triol derivative (F1-F6) mixed in anhydrous CH ₂ Cl ₂ under nitrogen. To this mixture was added AgOTf (4.5 eq) at 0 &lt; 0 &gt; C. To this solution was added slowly 4.0 equivalents of the solution of perbenzoylated maltosyl bromide dissolved in CH ₂ Cl ₂ . And the mixture was stirred at room temperature for 10 minutes. After the reaction was complete (confirmation of the reaction was complete by TLC), pyridine was added to the reaction mixture, which was diluted with CH ₂ Cl ₂ and filtered through celite. The filtrate was washed with 1 M aqueous Na ₂ S ₂ O ₃ , 0.1 M aqueous HCl and brine. The organic layer was then dried over anhydrous Na ₂ SO ₄ and the solvent was removed using a rotary evaporator. The residue was purified by silica gel column chromatography (EtOAc / hexane) to give a white solid state glycosylated compound (TNM-La) (45 to 75%).

<1-7> Deprotection vaporization  reaction ( deprotection  The general synthetic procedure for the reaction (step g in Figure 1)

This was in accordance with the synthesis method of Chae , PS et al. (Nat. Methods, 2010, 7, 1003.). Under Zemplen's condition to - O - benzoyl Chemistry (de- O -benzoylation) was carried out. The O- protected compound was dissolved in anhydrous CH ₂ Cl ₂ , then NaOMe, a methanolic solution of MeOH and 0.5 M, was added alternately to prevent precipitation. A 0.5 M methanolic solution of NaOMe was added to the reaction mixture to a final concentration of 0.05M. The reaction mixture was stirred at room temperature for 6 hours. After completion of the reaction, the reaction mixture was neutralized using Amberlite IR-120 (H ⁺ form) resin. The resin was filtered off, washed with MeOH, and the solvent was removed from the filtrate in vacuo . The residue was purified using silica gel chromatography (CH ₂ Cl ₂ / MeOH) to give a white solid compound (90 to 95%). The thus obtained compound is the compound TNM-Ls of the present invention

PREPARATION EXAMPLE 1 Synthesis of TNM-C9L

<1-1> Synthesis of 2,2-bis ((nonyloxy) methyl) propane-1,3-diol (Compound B1)

Compound B1 was synthesized according to the procedure of Example 1-1 with a yield of 92%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.65 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 4.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.56 (quin, J = 4.0 Hz, 4H), 1.28-1.26 (m, 24H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.4, 72.2, 65.7, 44.6, 32.1, 29.8, 29.7, 29.5, 26.3, 22.9, 14.1.

<1-2> 3- ( nonyloxy ) -2,2- bis ((nonyloxy) methyl) propane -One- ol (Compound C1)  synthesis

Compound C1 was synthesized according to the procedure of Example 1-2 with a yield of 87%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.43 (s, 6H), 3.38 (t, J = 8.0 Hz, 6H), 3.17 (t, J = 4.0 Hz, 1H), 1.53 (quin, J = 4.0 Hz, 6H), 1.30-1.26 (m, 36H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.6, 71.3, 66.3, 44.7, 31.6, 29.5, 25.8, 22.6, 14.1.

<1-3> 1- (3- (allyloxy) -2,2-bis ( ( nonyloxy ) methyl) 프로로oxy ) nonane (Compound D1)  synthesis

Compound D1 was synthesized according to the procedure of Example 1-3 with a yield of 92%. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.93-5.83 (m, 1H), 5.28-5.22 (m, 1H), 5.14-5.11 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( s, 2H), 3.38 (s , 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.52 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 36H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.4, 116.1, 72.4, 72.1, 71.9, 71.7, 69.8, 69.7, 45.6, 32.1, 29.8, 29.5, 26.4, 26.3, 22.9, 14.2.

&Lt; 1-4 > 3- (3- ( nonyloxy ) -2,2- bis ((nonyloxy) methyl) propoxy ) propane -One- ol (Compound E1)  synthesis

Compound E1 was synthesized according to the procedure of Example 1-4 with a yield of 55%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.76 (t, J = 8.0 Hz, 2H), 3.60 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.38 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.88 (br s, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.26 (m, 36H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.8, 71.4, 70.2, 62.9, 45.3, 32.1, 31.9, 29.9, 29.7, 29.8, 29.5, 26.4, 22.9, 14.2.

&Lt; 1-5 > 2- ( hidroxymethyl ) -2 - ((3- (3- ( nonyloxy ) -2,2- bis (( nonyloxy ) methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F1)

Compound F1 was synthesized with a yield of 45% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.70 (s, 6H), 3.52 (t, J = 8.0 Hz, 2H) 3.50 (s, 2H), 3.46 (t, J = 8.0 Hz, 2H), 3.38 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 2.88 (br s, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 36H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ73.1, 71.6, 69.9, 69.6, 68.8, 67.7, 64.9, 45.4, 31.9, 29.7, 29.6, 29.5, 29.4, 26.2, 22.7, 14.1.

<1-6> Synthesis of TNM-C9La

According to the general glycation procedure of Example 1-6, TNM-C9La was obtained in 52% yield Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.08 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.92-7.83 (m, 12H), 7.79-7.66 (m , 12H), 7.65-7.20 (m, 69H), 6.10 (t, J = 18.2 Hz, 3H), 5.75-5.62 (m, 3H), 5.43 (t, J = 16.4 Hz, 3H), 5.19-5.10 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.18 (d, J = 8.0 Hz, 3H), 3.70 (t, J = 20.0 Hz, 6H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 14H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H ), 1.31-1.25 (m, 36H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.0, 165.7, 165.5, 165.0, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 29.4, 129.3, 128.9, 128.8, 128.7, 128.6, 128.4, 128.2 , 100.8, 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.9, 68.8, 63.3, 62.3, 45.3, 31.9, 29.7, 29.5, 29.4, 26.2, 22.7, 14.2.

<1-7> Synthesis of TNM-C9L

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 1-7, TNM-C9L was obtained in 90% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 12.0 Hz, 3H), 3.84 J = 12.0 Hz, 2H), 1.56 (quin, J = 12.0 Hz, 6H), 1.87 (m, 1.34-1.29 (m, 36H), 0.89 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ105.1, 103.1, 81.5, 77.9, 76.6, 75.2, 74.8, 74.3, 72.6, 71.5, 70.6, 62.8, 46.7, 46.6, 33.3, 31.0, 30.8, 30.6, 27.6, 23.9, 14.7; HRMS ( EI ) : calcd. for C ₇₆ H ₁₄₂ O ₃₈ [M + Na] ⁺ 1685.9077, found 1685.9081.

PREPARATION EXAMPLE 2 Synthesis of TNM-C10L

<2-1> Synthesis of 2,2-bis ((decyloxy) methyl) propane-1,3-diol (Compound B2)

Compound B2 was synthesized according to the procedure of Example 1-1 with a yield of 92%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.65 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 4.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.56 (quin, J = 4.0 Hz, 4H), 1.28-1.26 (m, 28H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.4, 72.2, 65.7, 44.6, 32.1, 29.8, 29.7, 29.5, 26.3, 22.9, 14.1.

<2-2> 3- ( decyloxy ) -2,2- bis ((decyloxy) methyl) propane -One- be  (Compound C2)

Compound C2 was synthesized with a yield of 90% according to the procedure of Example 1-2. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.44 (s, 6H), 3.36 (t, J = 8.0 Hz, 6H), 3.17 (t, J = 8.0 Hz, 1H), 1.52 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 42H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.6, 71.3, 66.3, 44.7, 31.8, 29.6, 29.1, 26.1, 22.6, 14.1.

<2-3> 1- (3- (allyloxy) -2,2-bis ( ( decyloxy ) methyl) 프로로oxy ) decane (Compound D2)  synthesis

Compound D2 was synthesized with a yield of 92% according to the procedure of Example 1-3. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.93-5.84 (m, 1H), 5.27-5.22 (m, 1H), 5.13-5.10 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( s, 2H), 3.38 (s , 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.52 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 42H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.5, 116.1, 72.4, 72.1, 71.9, 69.8, 69.7, 45.6, 32.1, 29.8, 29.5, 26.4, 26.3, 22.9, 14.2.

&Lt; 2-4 > 3- (3- ( decyloxy ) -2,2- bis ((decyloxy) methyl) propoxy ) propane -One- be  (Compound E2)

Compound E2 was synthesized with a yield of 58% according to the procedure of Example 1-4. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.76 (t, J = 8.0 Hz, 2H), 3.59 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.37 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.88 (brs, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.26 (m, 42H ), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.8, 71.4, 70.2, 62.9, 45.3, 32.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.2.

<2-5> 2- ( hidroxymethyl ) -2 - ((3- (3- ( decyloxy ) -2,2- bis ((decyloxy) methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F2)

Compound F2 was synthesized with a yield of 47% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.51 (t, J = 8.0 Hz, 2H) 3.49 (s, 2H), 3.46 (t, J = 8.0 Hz, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 2.68 (br s, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 42H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ73.1, 71.6, 69.9, 68.8, 67.7, 64.9, 45.4, 31.9, 29.7, 29.6, 29.4, 26.2, 22.7, 14.1.

&Lt; 2-6 > Synthesis of TNM-C10La

According to the general glycation procedures of Examples 1-6, TNM-C10La was obtained at a yield of 45% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.08 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.92-7.83 (m, 12H), 7.79-7.66 (m , 12H), 7.65-7.20 (m, 69H), 6.12 (t, J = 18.2 Hz, 3H), 5.75-5.63 (m, 3H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.11 ( m, 6H), 4.58 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.18 (d, J = 8.0 Hz, 3H), 3.70 (t, J = 20.0 Hz, 6H), 3.36 (quin, J = 18.4 Hz, 12H), 3.24-2.99 (m, 14H), 1.79 (quin, J = 8.0 Hz, 2H), 1.53 (quin, J = 8.0 Hz, 6H ), 1.31-1.25 (m, 42H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.0, 165.7, 165.5, 165.0, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.8, 128.6, 128.4, 128.3, 100.8 , 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.8, 63.3, 62.3, 45.3, 31.9, 29.7, 29.5, 29.4, 26.3, 22.7, 14.2.

&Lt; 2-7 > Synthesis of TNM-C10L

According to the general synthesis procedure for the deprotection-vaporization reaction of Example 1-7, TNM-C10L was obtained in 95% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.34 (d, J = 8.0 Hz, 3H), 3.97 (d, J = 10.0 Hz, 3H), 3.84 -3.78 (m, 6H), 3.70-3.58 (m, 15H), 3.55-3.22 (m, 45H), 1.79 (quin, J = 12.8 Hz, 2H), 1.54 (quin, J = 12.0 Hz, 6H), 1.34-1.29 (m, 48H), 0.90 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ105.1, 103.1, 81.5, 77.8, 76.6, 75.1, 74.8, 74.2, 72.6, 71.5, 70.6, 70.0, 62.8, 62.2, 46.7, 46.6, 33.3, 31.0, 30.8, 30.7, 27.6, 23.9, 14.7; HRMS (EI) : calcd. for C ₈₂ H ₁₅₄ O ₃₈ [M + Na] ⁺ 1770.0016, found 1770.0011.

PREPARATION EXAMPLE 3 Synthesis of TNM-C11L

<3-1> Synthesis of 2,2-bis ((undecyloxy) methyl) propane-1,3-diol (Compound B3)

Compound B3 was synthesized with a yield of 94% according to the procedure of Example 1-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.65 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 8.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.56 (quin, J = 4.0 Hz, 4H), 1.28-1.25 (m, 32H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.5, 72.3, 65.7, 44.6, 32.1, 29.8, 29.7, 29.6, 29.5, 26.3, 22.9, 14.1.

<3-2> 3- ( undecyloxy ) -2,2- bis ((undecyloxy) methyl) propane -One- be  (Compound C3)

Compound C3 was synthesized according to the procedure of Example 1-2 with a yield of 85%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.43 (s, 6H), 3.38 (t, J = 8.0 Hz, 6H), 3.16 (t, J = 8.0 Hz, 1H), 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.7, 71.4, 62.9, 45.1, 32.1, 29.9, 29.7, 29.5, 26.4, 22.7, 14.2.

<3-3> 1- (3- ( allyloxy ) -2,2- bis ((undecyloxy) methyl) propoxy ) undecane  (Compound D3)

Compound D3 was synthesized with a yield of 94% according to the procedure of Example 1-3. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.95-5.85 (m, 1H), 5.27-5.22 (m, 1H), 5.14-5.11 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( s, 2H), 3.38 (s , 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.52 (quin, J = 8.0 Hz, 6H), 1.29-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.5, 116.1, 72.4, 72.1, 71.9, 70.0, 69.8, 69.7, 45.6, 32.1, 29.8, 29.5, 26.4, 26.3, 22.9, 14.2.

<3-4> 3- (3- ( undecyloxy ) -2,2- bis ((undecyloxy) methyl) propoxy ) propane -One- be  (Compound E3)

Compound E3 was synthesized with a yield of 58% according to the procedure of Example 1-4. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.77 (t, J = 8.0 Hz, 2H), 3.60 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.37 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.90 (brs, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.26 (m, 48H ), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.8, 71.5, 70.2, 63.0, 45.3, 32.1, 29.9, 29.8, 29.6, 29.5, 26.4, 22.9, 14.2.

<3-5> 2- ( hidroxymethyl ) -2 - ((3- (3- ( undecyloxy ) -2,2- bis ((undecyloxy) methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F3)

Compound F3 was synthesized with a yield of 47% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.50 (t, J = 8.0 Hz, 2H) 3.48 (s, 2H), 3.45 (t, J = 8.0 Hz, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 3.29 (brs, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31 -1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ): δ73.1, 71.6, 69.9, 69.6, 68.8, 67.7, 64.9, 45.4, 44.8, 31.9, 29.7, 29.6, 29.4, 26.2, 22.7, 14.2.

<3-6> Synthesis of TNM-C11La

According to the general glycation procedures of Examples 1-6, TNM-C11La was obtained with a yield of 48% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.08 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.92-7.85 (m, 12H), 7.79-7.67 (m , 12H), 7.65-7.22 (m, 69H), 6.11 (t, J = 20 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.10 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.19 (d, J = 8.0 Hz, 3H), 3.70 (t, J = 20.0 Hz, 6H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 14H), 1.79 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H ), 1.30-1.25 (m, 48H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.1, 165.8, 165.5, 165.1, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.8, 128.7, 128.6, 128.4, 128.2 , 100.8, 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.9, 68.8, 63.3, 62.4, 45.3, 31.9, 29.7, 29.5, 29.4, 26.3, 22.7, 14.2.

<3-7> Synthesis of TNM-C11L

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 1-7, TNM-C11L was obtained in 94% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.34 (d, J = 8.0 Hz, 3H), 3.97 (d, J = 10.0 Hz, 3H), 3.84 -3.78 (m, 6H), 3.70-3.58 (m, 15H), 3.55-3.22 (m, 45H), 1.79 (quin, J = 12.8 Hz, 2H), 1.54 (quin, J = 12.0 Hz, 6H), 1.34-1.29 (m, 48H), 0.90 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ105.1, 103.1, 81.5, 77.8, 76.6, 75.1, 74.8, 74.2, 72.6, 71.5, 70.6, 70.0, 62.8, 62.2, 46.7, 46.6, 33.3, 31.0, 30.8, 30.7, 27.6, 23.9, 14.7; HRMS (EI) : calcd. for C ₈₂ H ₁₅₄ O ₃₈ [M + Na] ⁺ 1770.0016, found 1770.0011.

PREPARATION EXAMPLE 4 Synthesis of TNM-C12L

<4-1> Synthesis of 2,2-bis ((dodecyloxy) methyl) propane-1,3-diol (Compound B4)

Compound B4 was synthesized with a yield of 94% according to the procedure of Example 1-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.64 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 8.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.55 (quin, J = 4.0 Hz, 4H), 1.28-1.25 (m, 36H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.6, 72.4, 65.8, 44.6, 32.1, 30.2, 29.9, 29.8, 29.5, 26.3, 22.9, 14.2.

&Lt; 4-2 > 3- ( dodecyloxy ) -2,2- bis ((dodecyloxy) methyl) propane -One- be  (Compound C4)

Compound C4 was synthesized with a yield of 85% according to the procedure of Example 1-2. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.43 (s, 6H), 3.38 (t, J = 8.0 Hz, 6H), 3.17 (t, J = 8.0 Hz, 1H), 1.53 (quin, J = 8.0 Hz, 6H), 1.30-1.26 (m, 54H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.8, 71.4, 70.2, 62.9, 45.1, 32.1, 29.9, 29.7, 29.6, 26.4, 22.7, 14.2.

&Lt; 4-3 > 1- (3- ( allyloxy ) -2,2- bis ((dodecyloxy) methyl) propoxy ) dodecane  (Compound D4)

Compound D4 was synthesized according to the procedure of Example 1-3 with a yield of 94%. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.93-5.83 (m, 1H), 5.27-5.22 (m, 1H), 5.14-5.11 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( s, 2H), 3.38 (s , 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 54H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.6, 116.2, 72.4, 72.1, 71.9, 69.9, 69.7, 45.6, 32.1, 29.8, 29.6, 26.4, 26.3, 22.9, 14.2.

&Lt; 4-4 > 3- (3- ( dodecyloxy ) -2,2- bis ((dodecyloxy) methyl) propoxy ) propane -One- be  (Compound E4)

Compound E4 was synthesized with a yield of 57% according to the procedure of Example 1-4. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.77 (t, J = 8.0 Hz, 2H), 3.60 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.37 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.90 (brs, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.25 (m, 54H ), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.9, 71.4, 70.2, 62.9, 45.3, 32.1, 29.9, 29.8, 29.6, 29.5, 26.4, 22.9, 14.3.

<4-5> 2- ( hidroxymethyl ) -2 - ((3- (3- ( dodecyloxy ) -2,2- bis ((dodecyloxy) methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F4)

Compound F4 was synthesized with a yield of 50% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.70 (s, 6H), 3.52 (t, J = 8.0 Hz, 2H) 3.50 (s, 2H), 3.46 (t, J = 8.0 Hz, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 2.91 (br s, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 54H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.2, 71.6, 69.9, 68.9, 68.8, 67.7, 65.1, 45.4, 44.9, 31.9, 29.7, 29.6, 29.4, 26.2, 22.7, 14.2.

&Lt; 4-6 > Synthesis of TNM-C12La

According to the general glycation procedures of Examples 1-6, TNM-C12La was obtained in 70% yield Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.09 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.92-7.83 (m, 12H), 7.79-7.66 (m , 12H), 7.65-7.20 (m, 69H), 6.11 (t, J = 18.2 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.11 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.18 (d, J = 8 Hz, 3H), 3.70 (t, J = 20.0 Hz, 6H), 3.36 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 14H), 1.79 (quin, J = 8.0 Hz, 2H), 1.53 (quin, J = 8.0 Hz, 6H ), 1.34-1.25 (m, 54H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.0, 165.7, 165.5, 165.0, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.7, 128.6, 128.4, 128.2, 100.8 , 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.9, 68.8, 63.3, 62.3, 45.4, 31.9, 29.8, 29.6, 29.4, 26.3, 22.7, 14.2.

<4-7> Synthesis of TNM-C12L

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 1-7, TNM-C12L was obtained in 95% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 12.0 Hz, 3H), 3.84 J = 12.0 Hz, 2H), 1.55 (quin, J = 12.0 Hz, 6H), 3.80 (m, 1.34-1.29 (m, 54H), 0.90 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ105.1, 103.1, 81.5, 77.8, 76.6, 75.1, 74.9, 74.2, 72.6, 71.5, 70.6, 62.8, 46.7, 46.6, 33.3, 31.0, 30.8, 30.7, 27.6, 23.9, 14.7; HRMS (EI) : calcd. for C ₈₅ H ₁₆₀ O ₃₈ [M + Na] ⁺ 1812.0485, found 1812.0480.

PREPARATION EXAMPLE 5 Synthesis of TNM-C13L

<5-1> Synthesis of 2,2-bis ((tridecyloxy) methyl) propane-1,3-diol (Compound B5)

Compound B5 was synthesized with a yield of 94% according to the procedure of Example 1-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.65 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 8.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.56 (quin, J = 4.0 Hz, 4H), 1.28-1.25 (m, 40H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.6, 72.5, 65.8, 44.6, 32.1, 30.3, 29.9, 29.7, 29.6, 26.3, 22.9, 14.2.

&Lt; 5-2 > 3- ( trideyloxy ) -2,2- bis ((trimethyloxy) methyl) propane -One- be  (Compound C5)

Compound C5 was synthesized according to the procedure of Example 1-2 with a yield of 85%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.43 (s, 6H), 3.38 (t, J = 8.0 Hz, 6H), 3.17 (t, J = 8.0 Hz, 1H), 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.25 (m, 60H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.7, 71.4, 70.2, 62.9, 45.1, 32.1, 29.9, 29.7, 29.6, 26.4, 22.7, 14.2.

&Lt; 5-3 > 1- (3- ( allyloxy ) -2,2- bis ((tridecyloxy) methyl) propoxy ) tridecane  (Compound D5)

Compound D5 was synthesized according to the procedure of Example 1-3 with a yield of 95%. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.93-5.83 (m, 1H), 5.27-5.22 (m, 1H), 5.14-5.11 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( (s, 2H), 3.38 (s, 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.52 (quin, J = 8.0 Hz, 6H), 1.28-1.26 J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.6, 116.1, 72.4, 72.1, 71.9, 69.8, 69.7, 45.6, 32.1, 29.9, 29.8, 29.7, 29.6, 26.4, 26.3 , 22.9, 14.3.

<5-4> 3- (3- ( tridecyloxy ) -2,2- bis ((tridecyloxy) methyl) propoxy ) propane -1-ol &lt; / RTI &gt; (Compound E5)

Compound E5 was synthesized with a yield of 60% according to the procedure of Example 1-4. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.76 (t, J = 8.0 Hz, 2H), 3.61 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.38 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.90 (brs, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.25 (m, 60H ), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.9, 71.4, 70.2, 62.9, 45.3, 32.1, 31.9, 30.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.3.

<5-5> 2- ( hidroxymethyl ) -2 - ((3- (3- ( tridecyloxy ) -2,2- bis (tridecyloxy)  methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F5)

Compound F5 was synthesized with a yield of 49% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.70 (s, 6H), 3.52 (t, J = 8.0 Hz, 2H) 3.50 (s, 2H), 3.46 (t, J = 8.0 Hz, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 2.67 (brs, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31 -1.25 (m, 60H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.14, 71.6, 69.9, 68.8, 67.7, 65.1, 45.5, 31.9, 29.7, 29.6, 29.4, 26.2, 22.7, 14.2.

<5-6> Synthesis of TNM-C13La

According to the general glycation procedures of Examples 1-6, TNM-C13La was obtained at a yield of 65% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.09 (d, J = 8.0 Hz, 6H), 7.96 (d, J = 7.2Hz, 6H), 7.90-7.83 (m, 12H), 7.79-7.66 (m , 12H), 7.65-7.20 (m, 69H), 6.08 (t, J = 18.2 Hz, 3H), 5.75-5.63 (m, 6H), 5.41 (t, J = 16.4 Hz, 3H), 5.20-5.10 ( m, 6H), 4.56 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.18 (d, J = 8 Hz, 3H), 3.70 (t, J = 20.0 Hz, 6H), 3.36 (quin, J = 18.4 Hz, 12H), 3.20-2.96 (m, 14H), 1.60 (quin, J = 8.0 Hz, 2H), 1.50 (quin, J = 8.0 Hz, 6H ), 1.34-1.21 (m, 60H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.8, 165.5, 165.0, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.8, 128.7, 128.6, 128.4, 128.2 , 100.8, 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.9, 68.8, 63.3, 62.3, 45.4, 32.0, 29.7, 29.5, 29.4, 26.2, 22.8, 14.3.

<5-7> Synthesis of TNM-C13L

According to the general synthesis procedure for the deprotection-vaporization reaction of Example 1-7, TNM-C13L was obtained in 93% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.34 (d, J = 8.0 Hz, 3H), 3.97 (d, J = 10.0 Hz, 3H), 3.86 -3.78 (m, 6H), 3.70-3.57 (m, 15H), 3.55-3.20 (m, 45H), 1.79 (quin, J = 12.4 Hz, 2H), 1.54 (quin, J = 12.0 Hz, 6H), 1.34-1.28 (m, 60H), 0.90 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ105.1, 103.1, 81.5, 77.8, 76.7, 75.1, 74.8, 74.2, 72.6, 71.5, 70.6, 70.0, 62.8, 46.7, 46.6, 33.3, 31.0, 30.8, 30.7, 27.6, 23.9, 14.7; HRMS (EI) : calcd. for C ₈₈ H ₁₆₆ O ₃₈ [M + Na] ⁺ 1855.0989, found 1855.1305.

&Lt; Preparation Example 6 > Synthesis of RMA-C5E

<6-1> Synthesis of 2,2-bis ((tetradecyloxy) methyl) propane-1,3-diol (Compound B6)

Compound B6 was synthesized with a yield of 94% according to the procedure of Example 1-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.65 (d, J = 4.0 Hz, 4H), 3.51 (s, 4H), 3.42 (t, J = 8.0 Hz, 4H), 2.85 (t, J = 4.0 Hz, 2H), 1.56 (quin, J = 4.0 Hz, 4H), 1.28-1.25 (m, 44H), 0.88 (t, J = 7.2 Hz, 6H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.6, 72.5, 65.8, 44.6, 32.1, 30.3, 29.9, 29.7, 29.6, 26.4, 22.9, 14.3.

&Lt; 6-2 > 3- ( tetradecyloxy ) -2,2- bis ((tetradecyloxy) methyl) propane -One- be  (Compound C6)

Compound C6 was synthesized according to the procedure of Example 1-2 with a yield of 85%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.71 (d, J = 8.0 Hz, 2H), 3.43 (s, 6H), 3.38 (t, J = 8.0 Hz, 6H), 3.16 (t, J = 8.0 Hz, 1H), 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.25 (m, 66H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.7, 71.4, 70.3, 62.9, 45.1, 32.1, 31.9, 29.9, 29.7, 29.5, 26.4, 22.7, 14.2.

<6-3> Synthesis of 1- (3- ( allyloxy ) -2,2- bis ((tetradecyloxy) methyl) 프로로oxy ) tetradecane  (Compound D6)

Compound D6 was synthesized with a yield of 93% according to the procedure of Example 1-3. ^{1 H NMR (400 MHz, CDCl} 3): δ 5.93-5.83 (m, 1H), 5.27-5.22 (m, 1H), 5.14-5.11 (m, 1H), 3.95-3.93 (m, 2H), 3.42 ( s, 2H), 3.38 (s , 6H), 3.35 (t, J = 8.0 Hz, 6H), 1.52 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 66H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 135.7, 116.2, 72.5, 72.4, 72.1, 71.9, 71.7, 69.8, 69.7, 45.6, 32.1, 29.8, 29.5, 26.4, 26.4, 23.2, 14.3.

<6- 4> 3 - (3- ( tetradecyloxy ) -2,2- bis ((tetradecyloxy) methyl) 프로로oxy ) propane -1-ol &lt; / RTI &gt; (Compound E6)

Compound E6 was synthesized with a yield of 58% according to the procedure of Example 1-4. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.77 (t, J = 8.0 Hz, 2H), 3.60 (t, J = 8.0 Hz, 2H), 3.43 (s, 2H), 3.37 (s, 6H), 3.35 (t, J = 8.0 Hz , 6H), 2.89 (brs, 1H), 1.80 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.30-1.25 (m, 66H ), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 71.9, 71.4, 70.2, 62.9, 45.3, 32.2, 31.1, 29.9, 29.8, 29.7, 29.5, 26.5, 22.9, 14.3.

<6-5> 2- ( hidroxymethyl ) -2 - ((3- (3- ( tetradecyloxy ) -2,2- bis ((tetrade cyloxy) methyl) propoxy) propoxy) methyl) propane-1,3-diol (Compound F6)

Compound F6 was synthesized with a yield of 50% according to the procedure of Example 1-5. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.70 (s, 6H), 3.52 (t, J = 8.0 Hz, 2H) 3.50 (s, 2H), 3.46 (t, J = 8.0 Hz, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 3.35 (s, 2H), 2.65 (br s, 3H), 1.79 (quin, J = 8.0 Hz, 2H), 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.25 (m, 66H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.14, 71.6, 69.9, 68.8, 67.7, 65.1, 45.5, 32.1, 31.9, 29.7, 29.7, 29.4, 26.2, 22.7, 14.3.

&Lt; 6-6 > Synthesis of TNM-C14La

According to the general glycation procedures of Examples 1-6, TNM-C14La was obtained with a yield of 75% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.08 (d, J = 8.0 Hz, 6H), 7.98 (d, J = 8.0 Hz, 6H), 7.88-7.84 (m, 12H), 7.79-7.66 (m , 12H), 7.65-7.20 (m, 69H), 6.10 (t, J = 18.2 Hz, 3H), 5.75-5.62 (m, 6H), 5.43 (t, J = 16.4 Hz, 3H), 5.19-5.12 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.18 (d, J = 8.0 Hz, 3H), 3.69 (t, J = 20.0 Hz, 6H), 3.32 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 14H), 1.65 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H ), 1.31-1.21 (m, 66H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.8, 165.5, 165.1, 165.0, 164.7, 133.4, 133.1, 129.9, 129.8, 129.7, 129.6, 129.4, 129.3, 128.9, 128.8, 128.7, 128.6, 128.4 , 128.2, 100.8, 95.7, 72.1, 72.0, 71.5, 71.2, 69.7, 68.9, 68.8, 63.3, 62.3, 45.4, 32.0, 29.7, 29.5, 29.4, 26.2, 22.8, 14.3.

<6-7> Synthesis of TNM-C14L

According to the general synthesis procedure for the deprotection-vaporization reaction of Examples 1-7, TNM-C14L was obtained in 95% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.34 (d, J = 7.6 Hz, 3H), 3.97 (d, J = 10.0 Hz, 3H), 3.84 -3.78 (m, 6H), 3.70-3.58 (m, 15H), 3.55-3.22 (m, 45H), 1.79 (quin, J = 12.8 Hz, 2H), 1.54 (quin, J = 12.0 Hz, 6H), 1.34-1.28 (m, 66H), 0.90 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CD ₃ OD):? 103.1, 81.5, 77.9, 74.9, 72.5, 71.5, 70.6, 62.8, 54.9, 46.6, 33.3, 30.8, 30.7, 27.6, 24.3, 23.9, 14.7; HRMS ( EI ) : calcd. for C ₉₁ H ₁₇₂ O ₃₈ [M + Na] ⁺ 1897.1459, found 1897.1396.

Example 2 Synthesis of TNM-Ss

The synthesis scheme of TNM-Ss is shown in Fig. Four compounds of TNM-Ss were synthesized according to the synthesis methods of the following <2-1> to <2-7>, and they are shown in FIG.

TNM-Ss was synthesized from the trialkylated hydroborated compound (E3-E6) produced by the procedures of Examples <1-1> to <1-4> through the following procedure.

<2-1> Alcohol Alcohol oxidation  The general synthesis procedure (step h of Figure 2)

CH ₂ Cl ₂ (E3-E6) (1.0 eq.) Was slowly added to a solution of pyridinium chlorochromate (PCC) (2.0 eq.) And celite in a 1: . The resulting mixture was stirred at room temperature for 1 day. The solution was diluted with CH ₂ Cl ₂ and then filtered over a short pad of celite. The solvent was removed at low pressure and the resulting residue was purified sequentially by silica gel column chromatography (EtOAc / Hexane) to give the trialkylated aldehyde compound (H3-H6) in the form of an oil liquid (90-95%).

<2-2> Trialkylated  Triol ( 트리화화 tri - be  ) (Step i of Figure 2)

Formaldehyde solution was added to a solution of trialkylated aldehyde (H3-H6) mixed with 50% water-soluble alkali EtOH (KOH 30%). The resulting mixture was stirred at room temperature for 2 hours and then heated at 60 &lt; 0 &gt; C for 2 days. After removing EtOH with a rotary evaporator, the resulting mixture was extracted with diethyl ether. The organic layer was dried over anhydrous Na ₂ SO _4, purified by column chromatography on silica gel (EtOAc / hexane) to give the alkylated tree triol compound of the oil liquid (I3-16) (30 to 40%).

<2-3> General synthesis procedure of glycosycosylation reaction (step f in FIG. 2)

This was in accordance with the synthesis method of Chae , PS et al. (Nat. Methods, 2010, 7, 1003.). To the stirred solution of the trialkylated triol compound (I3-I6) mixed in anhydrous CH ₂ Cl ₂ under nitrogen was added 2,4,6-collidine (1.0 eq.). To this mixture was added AgOTf (4.5 eq) at 0 &lt; 0 &gt; C. To this solution was added slowly 4.0 equivalents of the solution of perbenzoylated maltosyl bromide dissolved in CH ₂ Cl ₂ . And the mixture was stirred at room temperature for 10 minutes. After the reaction was complete (confirmation of the reaction was complete by TLC), pyridine was added to the reaction mixture, which was diluted with CH ₂ Cl ₂ and filtered through celite. The filtrate was washed with 1 M aqueous Na ₂ S ₂ O ₃ , 0.1 M aqueous HCl and brine. The organic layer was then dried over anhydrous Na ₂ SO ₄ and the solvent was removed using a rotary evaporator. The residue was purified by silica gel column chromatography (EtOAc / hexane) to give a white solid state glycosylated compound (TNM-Sa) (45 to 75%).

<2-4> Deprotection vaporization  reaction ( deprotection  The general synthetic procedure for the reaction (step g in Figure 2)

This was followed by the synthesis method of Chae and PS (Nat Meth 2010, 7, 1003.). Under Zemplen's condition to - O - benzoyl Chemistry (de- O -benzoylation) was carried out. The O- protected compound was dissolved in anhydrous CH ₂ Cl ₂ , then NaOMe, a methanolic solution of MeOH and 0.5 M, was added alternately to prevent precipitation. A 0.5 M methanolic solution of NaOMe was added to the reaction mixture to a final concentration of 0.05M. The reaction mixture was stirred at room temperature for 6 hours. After completion of the reaction, the reaction mixture was neutralized using Amberlite IR-120 (H ⁺ form) resin. The resin was filtered off, washed with MeOH, and the solvent was removed from the filtrate in vacuo . The residue was purified using silica gel chromatography (CH ₂ Cl ₂ / MeOH) to give a white solid compound (90 to 95%). The thus obtained compound is the compound TNM-Ss of the present invention

PREPARATION EXAMPLE 7 Synthesis of TNM-C11S

&Lt; 7-1 > 3- (3- ( undecyloxy ) -2,2- bis ((undecyloxy) methyl) propoxy ) propanal  Synthesis of Compound H3

Compound H3 was synthesized with a yield of 90% according to the procedure of Example 2-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 9.69 (s, 1H), 3.68 (t, J = 8.0 Hz, 2H), 3.35 (s, 6H), 3.28 (t, J = 8.0 Hz, 6H), 2.54-2.50 (m, 2H) 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ CNMR _{(100 MHz, CDCl 3):} δ202.0, 71.7, 70.4, 69.7, 65.4, 45.5, 44.0, 32.1, 29.9, 29.8, 29.7, 29.5, 26.4, 22.9, 14.3.

&Lt; 7-2 > 2- ( hidroxymethyl ) -2 - ((3- ( undecyloxy ) -2,2- bis (( undecyloxy ) methyl) propoxy) methyl) propane-1,3-diol Synthesis of Compound I3

Compound I3 was synthesized with a yield of 37% according to the procedure of Example 2-2. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.46 (s, 2H) 3.42 (s, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 2.75 (brs, 3H) , 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.5, 72.5, 72.0, 70.6, 65.2, 45.4, 32.1, 29.9, 29.7, 29.5, 26.4, 22.9, 14.3.

<7-3> Synthesis of TNM-C11Sa

According to the general glycation procedure of Example 2-3, TNM-C11Sa was obtained at a yield of 45% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.07 (d, J = 8.0 Hz, 6H), 7.97 (d, J = 8.0 Hz, 6H), 7.90-7.83 (m, 12H), 7.78-7.72 (m , 12H), 7.66-7.22 (m, 69H), 6.11 (t, J = 20.0 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.10 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.34-4.26 (dd, J = 8.0, 12.0 Hz, 9H), 4.19 (d, J = 8.0 Hz, 3H), 3.70 (t, J = 20.0 Hz, 4H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 12H), 1.79 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H ), 1.30-1.25 (m, 48H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.9, 165.6, 165.2, 165.1, 164.8, 133.5, 133.2, 130.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.1, 129.0, 128.9, 128.8, 128.5 , 128.4, 101.2, 96.0, 72.3, 71.5, 71.3, 70.0, 69.5, 69.1, 45.7, 32.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.3.

<7-4> Synthesis of TNM-C11S

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 2-4, TNM-C11Sa was obtained in 94% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 10.0 Hz, 3H), 3.87 J = 12.0 Hz, 6H), 1.34-1.29 (m, 48H), 0.90 (m, 1H), 3.57-3.28 , &Lt; / RTI &gt; J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ 105.2, 103.1, 81.6, 77.9, 76.7, 75.2, 74.9, 74.3, 72.7, 71.6, 70.9, 62.8, 62.3, 33.2, 31.0, 30.9, 30.8, 30.7, 27.6 , 23.9, 14.6; HRMS ( EI ) : calcd. for C ₇₉ H ₁₄₈ O ₃₇ [M + Na] ⁺ 1711.9597, found 1711.9594.

PREPARATION EXAMPLE 8 Synthesis of TNM-C12S

&Lt; 8-1 > 3- (3- ( dodecyloxy ) -2,2- bis ((dodecyloxy) methyl) propoxy ) propanal  (Compound H4)

Compound H4 was synthesized with a yield of 93% according to the procedure of Example 2-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 9.69 (s, 1H), 3.68 (t, J = 8.0 Hz, 2H), 3.35 (s, 6H), 3.28 (t, J = 8.0 Hz, 6H), 2.54-2.50 (m, 2H) 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 2.02, 71.7, 70.4, 69.7, 65.4, 45.5, 44.0, 32.1, 29.9, 29.7, 29.5, 26.4, 22.9, 14.3.

&Lt; 8-2 > 2- ( hidroxymethyl ) -2 - ((3- ( dodecyloxy ) -2,2- bis (( dodecyloxy ) methyl) propoxy) methyl) propane-1,3-diol (Compound I4)

Compound I4 was synthesized according to the procedure of Example 2-2 with a yield of 30%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.46 (s, 2H) 3.42 (s, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 2.82 (brs, 3H) , 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 54H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.5, 72.5, 72.0, 70.6, 65.2, 45.4, 32.1, 29.9, 29.7, 29.5, 26.4, 22.9, 14.3.

<8-3> Synthesis of TNM-C12Sa

According to the general glycation procedures of Example 2-3, TNM-C12Sa was obtained with a yield of 58% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.07 (d, J = 8.0 Hz, 6H), 7.97 (d, J = 8.0 Hz, 6H), 7.90-7.83 (m, 12H), 7.78-7.72 (m , 12H), 7.66-7.22 (m, 69H), 6.11 (t, J = 20.0 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.10 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.35-4.27 (dd, J = 8.0, 12.0 Hz, 9H), 4.19 (d, J = 8.0 Hz, 3H), 3.70 (t, J = 20.0 Hz, 4H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 12H), 1.79 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H ), 1.30-1.25 (m, 54H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.9, 165.6, 165.2, 165.1, 164.8, 133.5, 133.2, 130.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.1, 129.0, 128.9, 128.8, 128.7 , 128.5, 128.4, 101.2, 96.0, 72.3, 71.5, 71.3, 70.0, 69.5, 69.1, 45.7, 32.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.3.

<8-4> Synthesis of TNM-C12S

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 2-4, TNM-C12Sa was obtained in 90% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ 5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 10.0 Hz, 3H), 3.87 J = 12.0 Hz, 6H), 1.34-1.29 (m, 54H), 0.90 (m, 1H), 3.50-3.23 , &Lt; / RTI &gt; J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ 105.2, 103.1, 81.6, 77.9, 76.7, 75.2, 74.9, 74.3, 72.7, 71.6, 70.9, 62.8, 62.3, 33.2, 31.0, 30.9, 30.8, 30.6, 27.6 , 23.9, 14.6; HRMS ( EI ) : calcd. for C ₈₂ H ₁₅₄ O ₃₇ [M + Na] ⁺ 1754.0067, found 1754.0071.

PREPARATION EXAMPLE 9 Synthesis of TNM-C13S

<9-1> 3- (3- ( tridecyloxy ) -2,2- bis ((tridecyloxy) methyl) propoxy ) propanal  (Compound H5)

Compound H5 was synthesized with a yield of 95% according to the procedure of Example 2-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 9.69 (s, 1H), 3.68 (t, J = 8.0 Hz, 2H), 3.36 (s, 6H), 3.28 (t, J = 8.0 Hz, 6H), 2.55-2.51 (m, 2H) 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 2.02, 71.7, 70.4, 69.7, 65.4, 45.5, 44.0, 32.1, 29.9, 29.8, 29.7, 29.5, 26.4, 22.9, 14.3.

&Lt; 9-2 > 2- ( hidroxymethyl ) -2 - ((3- ( tridecyloxy ) -2,2- bis (tridecyloxy)  methyl) propoxy) methyl) propane-1,3-diol (Compound I5)

Compound I5 was synthesized according to the procedure of Example 2-2 with a yield of 35%. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.47 (s, 2H) 3.42 (s, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 2.90 (brs, 3H) , 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 60H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.5, 72.5, 72.0, 70.6, 65.2, 45.4, 32.1, 30.0, 29.9, 29.7, 29.5, 26.4, 22.9, 14.3.

<9-3> Synthesis of TNM-C13Sa

According to the general glycation procedure of Example 2-3, TNM-C13Sa was obtained with a yield of 60% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ 8.07 (d, J = 8.0 Hz, 6H), 7.97 (d, J = 8.0 Hz, 6H), 7.90-7.83 (m, 12H), 7.78-7.72 (m , 12H), 7.66-7.22 (m, 69H), 6.11 (t, J = 20.0 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.10 ( m, 6H), 4.57 (t , J = 3.2 Hz, 6H), 4.35-4.27 (dd, J = 8.0, 12.0 Hz, 9H), 4.19 (d, J = 8.0 Hz, 3H), 3.71 (t, J = 20.0 Hz, 4H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 12H), 1.79 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H ), 1.30-1.25 (m, 60H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.9, 165.6, 165.2, 165.1, 164.9, 133.5, 133.2, 130.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.1, 129.0, 128.9, 128.8, 128.5 , 128.4, 101.2, 96.0, 72.3, 71.5, 71.3, 70.0, 69.5, 69.1, 45.7, 32.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.3.

<9-4> Synthesis of TNM-C13S

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 2-4, TNM-C13Sa was obtained in 93% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 10.0 Hz, 3H), (M, 6H), 3.70-3.58 (m, 13H), 3.55-3.22 (m, 43H), 1.55 (quin, J = 12.0 Hz, 6H), 1.34-1.29 t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ 105.2, 103.1, 81.6, 77.9, 76.7, 75.2, 74.9, 74.3, 72.7, 71.6, 70.9, 62.8, 62.3, 33.2, 31.0, 30.9, 30.8, 30.6, 27.6 , 23.9, 14.6; HRMS ( EI ) : calcd. for C ₈₅ H ₁₆₀ O ₃₇ [M + Na] ⁺ 1796.0536, found 1796.0541.

PREPARATION EXAMPLE 10 Synthesis of TNM-C14S

<10-1> 3- (3- ( dodecyloxy ) -2,2- bis ((dodecyloxy) methyl) propoxy ) propanal  (Compound H6)

Compound H6 was synthesized with a yield of 95% according to the procedure of Example 2-1. ^{1 H NMR (400 MHz, CDCl} 3): δ 9.69 (s, 1H), 3.68 (t, J = 8.0 Hz, 2H), 3.36 (s, 6H), 3.28 (t, J = 8.0 Hz, 6H), 2.55-2.51 (m, 2H) 1.53 (quin, J = 8.0 Hz, 6H), 1.28-1.26 (m, 48H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 2.02, 71.7, 70.4, 69.7, 65.4, 45.5, 44.0, 32.1, 29.9, 29.8, 29.7, 29.5, 26.4, 22.9, 14.3.

<10-2> 2- ( hidroxymethyl ) -2 - ((3- ( dodecyloxy ) -2,2- bis (( dodecyloxy ) methyl) propoxy) methyl) propane-1,3-diol (Compound I6)

Compound I6 was synthesized with a yield of 40% according to the procedure of Example 2-2. ^{1 H NMR (400 MHz, CDCl} 3): δ 3.67 (s, 6H), 3.47 (s, 2H) 3.42 (s, 2H), 3.36 (quin, J = 8.0 Hz, 12H), 2.85 (brs, 3H) , 1.52 (quin, J = 8.0 Hz, 6H), 1.31-1.26 (m, 66H), 0.88 (t, J = 7.2 Hz, 9H); ¹³ C NMR (100 MHz, CDCl ₃ ):? 73.5, 72.5, 72.0, 70.6, 65.2, 45.4, 32.1, 30.0, 29.9, 29.7, 29.5, 26.4, 22.9, 14.3.

&Lt; 10-3 > Synthesis of TNM-C14Sa

According to the general glycation procedure of Example 2-3, TNM-C14Sa was obtained with a yield of 55% Were synthesized. ^{1 H NMR (400 MHz, CDCl} 3): δ8.07 (d, J = 8.0 Hz, 6H), 7.97 (d, J = 8.0 Hz, 6H), 7.90-7.83 (m, 12H), 7.78-7.72 ( m, 12H), 7.66-7.22 (m , 69H), 6.11 (t, J = 20.0 Hz, 3H), 5.75-5.63 (m, 6H), 5.44 (t, J = 16.4 Hz, 3H), 5.20-5.10 (m, 6H), 4.57 ( t, J = 3.2 Hz, 6H), 4.35-4.27 (dd, J = 8.0, 12.0 Hz, 9H), 4.19 (d, J = 8.0 Hz, 3H), 3.71 (t, J = 20.0 Hz, 4H), 3.37 (quin, J = 18.4 Hz, 12H), 3.23-2.98 (m, 12H), 1.79 (quin, J = 8.0 Hz, 2H), 1.51 (quin, J = 8.0 Hz, 6H), 1.30-1.25 (m, 66H), 0.87 (t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CDCl} 3): δ166.2, 165.9, 165.6, 165.2, 165.1, 164.9, 133.5, 133.2, 130.1, 130.0, 129.9, 129.7, 129.6, 129.5, 129.1, 129.0, 128.8, 128.5, 128.4 , 101.2, 96.0, 72.3, 71.5, 71.3, 70.0, 69.5, 69.1, 45.7, 32.1, 29.9, 29.8, 29.5, 26.4, 22.9, 14.3.

<10-4> Synthesis of TNM-C14S

According to the general synthetic procedure for the deprotection-vaporization reaction of Example 2-4, TNM-C14Sa was obtained in 95% yield Were synthesized. ^{1 H NMR (400 MHz, CD} 3 OD): δ5.15 (d, J = 4.0 Hz, 3H), 4.35 (d, J = 8.0 Hz, 3H), 3.98 (d, J = 10.0 Hz, 3H), (M, 6H), 3.70-3.58 (m, 13H), 3.56-3.23 (m, 43H), 1.55 (quin, J = 12.0 Hz, 6H), 1.34-1.29 t, J = 7.2 Hz, 9H); ^{13 C NMR (100 MHz, CD} 3 OD): δ 105.2, 103.1, 81.6, 77.9, 76.7, 75.2, 74.9, 74.3, 72.7, 71.6, 70.9, 62.8, 62.3, 33.2, 31.0, 30.9, 30.8, 30.6, 27.6 , 23.9, 14.6; HRMS ( EI ) : calcd. _{for C 88 H 166 O 37 [} M + Na] + 1839.1040, found 1839.0858.

&Lt; Example 3 > TNMs characteristic

Example 1 and to determine the properties of the synthesized TNMs according to the synthesis method of 2, the TNMs molecular weight (MW), the critical micelle concentration (critical micellar concentration; CMC) and hydrodynamic radius of the micelles formed (hydrodynamic radii; R _h ) was measured.

Specifically, the critical micelle concentration (CMC) was measured using hydrophobic fluorescent dye, diphenylhexatriene (DPH), and the hydrodynamic radius ( R _h ) of the micelle formed by each preparation was determined by dynamic light scattering (DLS) experiments. The measured results are shown in Table 1 in comparison with DDM, which is a conventional amphipathic molecule (detergent).

Detergent ^Mw CMC (mM) CMC (wt%) ^R _h  ^(nm) TNM-C9L 1646.9 ~ 0.002 ~ 0.00033 4.5 ± 0.2 TNM-C10L 1689.0 ~ 0.0015 ~ 0.00025 4.5 ± 0.1 TNM-C11L 1731.1 ~ 0.001 ~ 0.00017 29.5 ± 1.6 TNM-C12L 1773.2 ~ 0.0008 ~ 0.00016 33.8 ± 0.8 TNM-C13L 1815.2 ~ 0.0006 ~ 0.00011 45.9 ± 2.7 TNM-C14L 1857.3 ~ 0.0004 ~ 0.00007 48.1 ± 1.1 TNM-C11S 1690.0 ~ 0.0025 ~ 0.00051 5.7 ± 0.2 TNM-C12S 1732.1 ~ 0.0015 ~ 0.00035 6.5 ± 0.6 TNM-C13S 1774.2 ~ 0.001 ~ 0.00071 9.8 ± 1.7 TNM-C14S 1816.3 ~ 0.0008 ~ 0.00054 11.6 ± 0.2 DDM 510.1 ~ 0.17 ~ 0.0087 3.4 ± 0.02

The CMC value of TNMs was much smaller than that of DDM. The CMC values of TNM-C14 and TNM-C14S with the longest alkyl chain (C14) were 100 times smaller than those of DDM. Therefore, it was confirmed that TNMs had better solubility than DDM because micelles were easily formed in a small amount.

The size of micelles formed by TNMs was in the range of 4.5 ~ 48.1 nm and it was confirmed that micelles were formed larger than DDM. The size of micelle formed by TNMs increased with increasing alkyl chain length. Further, as a result of analyzing the size distribution based on the number of micelles formed by the TNMs, it was confirmed that TNMs (TNM-Ls and TNM-Ss) form micelles each having one cluster (FIG.

< Example  3> Of TNMs R. capsulatus superassembly  ( LHI - RC ) Structural stabilization ability evaluation (Fig. 4)

TNMs induced Rhodobacter capsulatus ) photosynthetic superassembly structure stabilization ability. Photosynthesis assemblies consist of light-harvesting complex I (LHI) and reaction center complex (RC). The structural stability of LHI-RC was measured by monitoring the structure of the protein for 20 days using UV-Vis spectroscopy. Amphiphilic molecules used all of the TNMs of the present invention and the existing amphiphilic molecules DDM and OG and the concentrations of amphiphilic molecules were measured at CMC + 0.04 wt% (FIG. 5A) and CMC + 0.2 wt% (FIG. 5B) The stability of LHI-RC protein according to the concentration of amphiphilic molecules was examined.

Specifically, the stability of the photosynthetic assembly of R. capsulatus was measured using the method described in our 2008 paper (PS Chae et al., ChemBioChem 2008, 9, 1706-1709). In summary, the inventors used membranes from R. capsulatus , U43 [pUHTM86Bgl] bacteria, which do not have light-harvesting complex II (LHII). R. capsulatus A 10 ml aliquot of the frozen lysate of the membrane was homogenized using a glass homogenizer and incubated at 32 [deg.] C for 30 minutes. The homogenized membrane was treated with 1.0 wt% DDM at 32 &lt; 0 &gt; C for 30 min. Membrane debris was ultracentrifuged at 315,000 g for 30 minutes at 4 ° C to collect pellets. DDM- solubilization (I in buffer containing 10 mM Tris, pH 7.8 - being balance and storage) 200 μL Ni ^{2 +} -NTA resin was added to the supernatant. The mixed solution was allowed to bind to the resin at 4 ° C for 1 hour. The resin was collected and washed twice with 500 μL binding buffer (pH 7.8 Tris buffer solution containing 1 × CMC DDM). After replacement with a new ultracentrifuge tube, the resin was treated twice with 300 μL elution buffer containing 1 M imidazole (the binding buffer at pH 7.8 with 1 M imidazole added). The DDM-purified supermassives were diluted with a solution of CMC + 0.04 wt%, CMC + 0.2 wt% of each TNMs, DDM and OG (pH 7.8, 920 μL). These protein complex samples were collected to monitor protein stability at regular intervals during the incubation period of 20 days (the first 10 days at 15 or 20 &lt; 0 &gt; C and then 10 days at 32 [deg.] C). The UV-Vis absorption spectrum was measured in the range of 650 nm to 950 nm.

All new compounds except TNM-C14S were superior to DDM and OG in stabilizing complexes. Compounds with the shortest alkyl chain (TNM-C9L and TNM-C11S) among the TNMs showed the most excellent effects. Increasing compound concentrations from CMC + 0.04 wt.% To CMC + 0.2 wt.% Resulted in poor protein stability in both DDM and OG (FIGS. 5 and 6), TNMs, especially the longest alkyl chains such as TNM-C14L and TNM- Similar results were obtained with compounds with However, the other TNM molecules at the concentration of this amphipathic molecule (CMC + 0.2 wt%) clearly dominated the effect of stabilizing the LHI-RC complex over DDM.

Example 4 Evaluation of Structure Stabilization Capability of Membrane Protein (UapA) of TNMs (FIG. 5)

Experiments were conducted to measure the structural stability of UapA (uric acid-xanthine / H ⁺ symporter) by TNMs in aqueous solution. The structural stability of UapA was measured by CPM assay. TNMs and DDM concentrations were measured at CMC + 0.04 wt% and CMC + 0.2 wt%.

Specifically, the UapA protein is expressed by Aspergillus &lt; RTI ID = 0.0 &gt; uric acid-xanthine / H ⁺ co-transporter (symporter) in the nidulans . Protein stability was measured by fluorescence spectrophotometry using a sulfhydryl-specific fluorophore CPM ( N - [4- (7-Diethylamino-4-methyl-3-coumarinyl) phenyl] maleimide) spectroscopy). The free sulfhydryl group (-SH) of the cysteine residue is in the center of the protein, but is exposed to the surface by protein unfolding and becomes accessible to the solvent. CPM reacts with free thiol to become fluorescent, thereby operating as a release sensor. For thermal stability measurements, UapAG411V &lt; _{RTI ID =} 0.0 _{&gt;# 1-11} &lt; / _{RTI &gt}; was first incubated with saccharomyces cerevisiae S. cerevisiae FGY217 and were isolated in sample buffer (20 mM Tris (pH 7.5), 150 mM NaCl, 0.03% DDM, 1 mM xanthine), as described in J. Leung et al ., Mol . Membr . Biol . 2013, 30, 32-42). UapA protein was concentrated to about 10 mg / ml using a 100 kDa molecular weight cut off filter (Millipore). Concentrated proteins were diluted 1: 150 in a buffer containing DDM or TNMs at concentrations of CMC + 0.04 wt% or CMC + 0.2 wt% in a Greiner 96-well plate. The CPM dye (Invitrogen) stored in DMSO (Sigma) was diluted in dye buffer (20 mM Tris (pH 7.5), 150 mM NaCl, 0.03% DDM, 5 mM EDTA) and 3 μl of diluted dye was tested for each protein Lt; / RTI &gt; The reaction was monitored using a microplate fluorescence spectrometer at 40 ° C for 120 minutes. A sample with a relative maximum fluorescence was used as a control to calculate the percentage of the relative folded protein remaining after 120 minutes at 40 ° C. Relative folded proteins were expressed over time using GraphPad Prism.

The ability to keep the UapA protein folded at CMC + 0.2 wt% was superior to DDM for all TNMs (Figure 7). In particular, TNM-C12L and TNM-C14S were found to be most effective among TNM-Ls and TNM-Ss, respectively (Fig. 7). TNMs showed similar ability at CMC + 0.04 wt% as at CMC + 0.2 wt% (Fig. 8). These results indicate that TNMs can be effectively used to stabilize membrane proteins, since they exert an excellent effect of keeping the UAPA protein extracted from the cell membrane in a structurally stable state in aqueous solution.

Example 5 Evaluation of Structure Stabilization Capability of Membrane Protein (LeuT) of TNMs

The experiment was performed to measure the structural stability of LeuT (leucine transporter) protein by TNMs. LeuT protein activity was measured by SPA (scintillation proximity assay) using protein substrate ([ ³ H] -Leu) and concentration of TNMs or DDM was used as CMC + 0.04 wt% and CMC + 0.2 wt%.

Purification of wild-type LeuT (leucine transporter) from Aquifex aeolicus was carried out according to the method described in G. Deckert et al. ( Nature 1998, 392, 353-358). LeuT is expressed in E. coli C41 (DE3) transformed with pET16b encoding the C-terminal 8xHis-tagged transporter (expression plasmids are provided by Dr E. Gouaux, Vollum Institute, Portland, Oregon, USA ). Briefly, the LeuT protein was isolated and dissolved in 1.0 wt% DDM, then the protein was bound to Ni ^{2 +} -NTA resin (Life Technologies, Denmark) and resuspended in 20 mM Tris-HCl (pH 8.0), 1 mM NaCl , 199 mM KCl, 0.05% DDM and 300 mM imidazole. Then, about 1.5 mg / ml protein sample (stock) was diluted with an equivalent buffer supplemented with DDM and imidazole but with TNMs or DDM (control) to a final concentration of CMC + 0.04 wt% or CMC + 0.2 wt% . Protein samples are stored at room temperature for 10 days, and was centrifuged at a specified time, the protein activity is SPA (scintillation proximity assay) (M. Quick et al., Proc. Natl, Acad. Sci. USA. 2007, 104, 3603- 3608.) was used to measure [ ³ H] -Leu binding. Analysis was performed with buffer containing 450 mM NaCl and each of the above compounds at the final concentration. The SPA reaction was carried out in the presence of 20 nM [ ³ H] -Leu and 1.25 mg / ml copper chelate (His-Tag) YSi beads (both purchased from PerkinElmer, Denmark). [ ³ H] -Leu binding was measured using a MicroBeta liquid scintillation counter (PerkinElmer).

As shown in Figs. 9 and 10, LeuT solubilized by TNM-C10L in TNM-Ls and TNM-11S in TNM-Ss showed the most excellent substrate binding ability, and the protein stabilizing effect Was superior to DDM. In addition, it was confirmed that the TNM-Ss having a shorter linker length exhibited the ability to maintain the activity of the transporter superior to the TNM-Ls having a longer length. The results were similar when the concentration of amphipathic compound increased from CMC + 0.04 wt% to CMC + 0.2 wt%.

As a result, TNM-C11L in TNMs showed the ability to maintain the initial activity of solubilized receptors similar to DDM (FIGS. 9 and 10). However, in the ability of long-term LeuT stabilization, DDM showed a rapid loss of protein activity over time, but TNM-C11L was found to maintain LeuT activity well during 10 days of culture (FIGS. 9 and 10). Therefore, TNM-C10L and TNM-C11S were superior to DDM in maintaining the activity of transporters.

&Lt; Example 6 > ₂ Evaluation of AR protein stabilization ability

Receptors were expressed in Sf9 insect cells infected with baculovirus and solubilized in 1% DDM. The solubilized receptor in DDM was purified by alprenolol sepharose in the presence of 0.01% cholesteryl succinate (CHS). The β ₂ AR purified by DDM was diluted with a buffer containing DDM or TNMs to a final concentration of CMC + 0.2 wt%. The β ₂ AR solubilized in each compound was stored at room temperature for 4 days and incubated at room temperature for 30 minutes with 10 nM of radioactive [ ³ H] -dihydroalprenolol (DHA) Ligand binding ability was measured. The mixture was loaded onto a G-50 column and the supernatant was collected using an aliquot of binding buffer (20 mM HEPES pH 7.5, 100 mM NaCl, supplemented with 0.5 mg / ml BSA). An additional 15 ml scintillation fluid was added. The receptor-bound [ ³ H] -DHA was measured with a scintillation counter (Beckman).

As a result, TNM-C12L among the TNMs showed the ability to maintain the initial activity of solubilized receptors similar to DDM (FIG. 11). However, in the ability to stabilize the receptor over time, DDM showed rapid activity loss over time, but receptors solubilized in TNM-C12L continued to maintain receptor activity during the 4 day incubation (FIG. 12). Therefore, it was concluded that TNM-C12L could be more effective in studying solubilized receptor proteins than DDM.

Claims (16)

A compound represented by the following formula (1):
[Chemical Formula 1]

In Formula 1,
Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;
X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;
Y ¹ , Y ² and Y ³ are oxygen (O); And
M is 0 or 1; The method according to claim 1,
Wherein the saccharide is a monosaccharide or a disaccharide. In the first aspect,
The saccharide is glucose or maltose. The method according to claim 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group; X ¹ to X ³ are glucose or maltose; Y ¹ , Y ² and Y ³ are oxygen (O); And m is 1. The method according to claim 1,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group; X ¹ to X ³ are glucose or maltose; Y ¹ , Y ² and Y ³ are oxygen (O); And m is 0. The method according to claim 1,
Wherein said compound is one of the compounds represented by the following formulas (2) to (11):
(2)

(3)

[Chemical Formula 4]

[Chemical Formula 5]

[Chemical Formula 6]

(7)

[Chemical Formula 8]

[Chemical Formula 9]

[Chemical formula 10]

(11)

The method according to claim 1,
Wherein said compound is an amphipathic molecule for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein. The method according to claim 1,
Wherein the compound has a micelle concentration (CMC) in the aqueous solution of 0.0001 to 1 mM. A composition for the extraction, solubilization, stabilization, crystallization or analysis of a membrane protein comprising a compound according to claim 1. 10. The method of claim 9,
Wherein the composition is a formulation of micelles, liposomes, emulsions or nanoparticles. 1) Synthesis of 5,5-bis-bromomethyl-2,2-dimethyl- [1,3] dioxane and 5,5-bis-bromomethyl- alcohol to form a dialkylated diol;
2) synthesizing a trialkylated monool by adding 1-bromoalkane to the product of step 1);
3) allylating the monool by adding allyl iodide to the product of step 2);
4) hydroboration oxidation of the product of step 3) to synthesize a trialkylated monool;
5) To a solution of 4- (bromomethyl) -1-methyl-2,6,7-trioxabicyclo [2.2.2] -octane (4- (bromomethyl) , 6,7-trioxabicyclo [2.2.2] -octane) to synthesize a trialkylated triol;
6) performing a glycosylation reaction on the product of step 5) to introduce a protecting group-attached saccharide; And
7) a step of performing a deprotection reaction on the product of step 6);
[Chemical Formula 1]

In Formula 1,
Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;
X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;
Y ¹ , Y ² and Y ³ are oxygen (O); And
M is 1. 1) Synthesis of 5,5-bis-bromomethyl-2,2-dimethyl- [1,3] dioxane (5,5-bis-bromomethyl- &Lt; / RTI &gt; to form a dialkylated diol compound;
2) synthesizing a trialkylated monool-type compound by adding 1-bromoalkane to the product of step 1);
3) allylating the monool by adding allyl iodide to the product of step 2);
4) hydroboration oxidation of the product of step 3) to synthesize a trialkylated monool compound;
5) synthesizing a trialkylated aldehyde-type compound through an alcohol oxidation reaction of the product of step 4);
6) synthesizing a trialkylated triol compound by adding formaldehyde and water-soluble alkaline ethanol to the product of step 5);
7) performing a glycosylation reaction on the product of step 6) to introduce a protecting group-attached saccharide; And
8) carrying out a deprotection reaction on the product of step 7);
[Chemical Formula 1]

In Formula 1,
Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;
X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;
Y ¹ , Y ² and Y ³ are oxygen (O); And
M is zero. A method for extracting, solubilizing, stabilizing, crystallizing or analyzing a membrane protein, which comprises treating a membrane protein with a compound represented by the following formula (1) in an aqueous solution:
[Chemical Formula 1]

In Formula 1,
Wherein R ^1, R ^2, and R ³ are each independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group, a substituted or unsubstituted C ₃ -C ₂₀ cycloalkyl group, or a substituted or unsubstituted C ₃ -C ₂₀ aryl group;
X ¹ , X ^2, and X ³ are each independently a saccharide linked with oxygen;
Y ¹ , Y ² and Y ³ are oxygen (O); And
M is 0 or 1; 14. The method of claim 13,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group; X ¹ to X ³ are glucose or maltose; Y ¹ , Y ² and Y ³ are oxygen (O); And m is one. 14. The method of claim 13,
Each of R ¹ to R ³ is independently a substituted or unsubstituted C ₃ -C ₂₀ alkyl group; X ¹ to X ³ are glucose or maltose; Y ¹ , Y ² and Y ³ are oxygen (O); And m is zero. 14. The method of claim 13,
The membrane protein may be selected from the group consisting of LHI-RC complex (light harvesting-I and the reaction center complex), UapA (Uric acid-xanthine / H + symporter), LeuT (Leucine transporter), β2AR (human β2 adrenergic receptor) , Or a combination of two or more thereof.

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