**1. Introduction**

Naphthoquinones represent one of the largest families of natural products and are widespread in nature. They were isolated from plants, marine invertebrates, fungi, and bacteria [1]. Naphthoquinones revealed a diverse spectrum of activities: anticancer [2–4], antibacterial [5,6], anti-infective [7], antimalarial [8], and cardioprotective action [9]. The quinone ring contains a system of double bonds conjugated with carbonyl groups: it is easily susceptible to reduction, oxidation, and addition of *O*-, *N*-, and *S*-nucleophiles [10,11]. The high reactivity of naphthoquinones and the well-developed methods

of its chemical modification make this group of compounds attractive for the profound development of new types of substances with high biological activity [12].

Studies are continuing on the antimicrobial activity of 1,4-naphthoquinones in relation to various microbial pathogens that are dangerous as sources of fatal diseases, epidemics, and nosocomial infections. In some cases, not only was the direct effect of new compounds on microbial cells investigated, but also their effect on the viability of biofilms formed by reproducing microorganisms. Thus, a series of new 2-hydroxy-3-phenylsulfanylmethyl-1,4-naphthoquinones were synthesized and evaluated against Gram-negative and Gram-positive bacterial strains and their biofilms to probe for potential lead structures. The structure modification applied in the series resulted in 12 new naphthoquinones with pronounced antimicrobial activity against *Escherichia coli* and *Pseudomonas aeruginosa*. Four molecules showed anti-biofilm activity and inhibited biofilm formation more than 60% with a better profile than standard antibacterial drug, ciprofloxacin [13,14].

Naphthoquinones often possess poor solubility which has hampered their practical use. The conjugation of naphthoquinones with non-toxic carbohydrates is one of the most successful ways for improving their solubility [15–19]. Moreover, the conjugation of naphthoquinones with carbohydrates led to novel structures with new types of biological activity [20,21]. Such naphthoquinone–carbohydrate conjugates include classical *O*- and *S*-glycosides (carbohydrates linked directly to naphthoquinone via glycosidic bond), non-glycosidic conjugates (connection with the carbohydrate component via not glycosidic ether linkage), and *N*-glycosyl triazoles (a triazolic ring connecting the carbohydrate moiety to naphthoquinone). In the course of our drug research project we developed an effective method for preparation of naphthoquinone acetylthioglucosides by the condensation of available substituted chloromethoxynaphthoquinone **1** with per-*O*-acetyl-1-thioderivatives of d-glucose **2a**, d-galactose **2b**, d-xylose **2c**, and l-arabinose **2d** and obtained related naphthoquinone acetylglucosides **3a**–**d** [22]. These acetylglycoside derivatives, **3a**–**d**, were readily deacetylated with MeONa/MeOH and immediately converted to the quinone–sugar tetracyclic conjugates **4a**–**d** in good yields (Scheme 1). The tetracyclic quinone–carbohydrate conjugates **4a**–**d** had a linear planar structure and retained the stereochemistry of the starting sugars.

**Scheme 1.** Synthetic route for the synthesis of fused tetracyclic conjugates **4a**–**d**.

The obtained sugar–quinone tetracycles were converted to acetyl derivatives by treatment with Ac2O/Py. Both synthesized tetracyclic quinone conjugates and their acetylated tetracyclic derivatives were active in vitro against human promyelocytic leukemia HL-60 in 1.0–5.0 μM concentrations, while starting acyclic acetylglycosides were approximately 10–100 times less active [23].

#### **2. Results and Discussion**

The synthesis of tetracyclic oxathiine-fused glycoside naphthoquinone conjugates can provide bioactive compounds, and the variation of naphthoquinone and carbohydrate moieties allows a structure–activity relationship (SAR) study. Therefore, this work aimed to conjugate per-*O*-acetylated 1-thiosugars **2a**–**d** with various substituted 1,4-naphthoquinones. The key intermediates, per-*O*-acetylated 1-thiosugars **2a**–**d**, were prepared from the respective peracetylated glycosyl halides (d-glucose, d-galactose, d-xylose, and l-arabinose) using reducing cleavage of its thiouronium salt with

sodium metabisulfite according to literature procedures [24–26]. Sugar thioderivatives comprise two pairs of structurally related carbohydrates: hexopyranoses of d-glucose and d-galactose, as well as pentopyranoses of d-xylose and l-arabinose, which differ in the configuration of the C4-OH group.
